CN101955774A

CN101955774A - Nanocrystalline painted plant

Info

Publication number: CN101955774A
Application number: CN2009110002888A
Authority: CN
Inventors: 韩明勇; N·B·穆罕默德·哈纳菲亚
Original assignee: Agency for Science Technology and Research Singapore
Current assignee: Agency for Science Technology and Research Singapore
Priority date: 2008-12-10
Filing date: 2009-12-10
Publication date: 2011-01-26
Also published as: SG162679A1; JP2010150254A

Abstract

The present invention relates to contain at least a nanocrystalline plant, when described plant is exposed to light time of specific wavelength, described plant is by described nanocrystalline a kind of specific color or the multiple color of launching.On the other hand, the present invention relates to carry out painted method with at least a nanocrystalline outside surface to plant.The invention still further relates to at least a nanocrystalline tissue and carry out painted method plant.

Description

Nanocrystalline dyed plants

Technical Field

The invention relates to plants containing at least one nanocrystal through which the plant emits a specific colour or colours when illuminated with light of a certain wavelength. Another aspect of the invention relates to a method of coloring an outer surface of a plant with at least one nanocrystal. The invention also relates to a method for staining the tissue of a plant with at least one nanocrystal.

Background

Semiconductor nanocrystals (quantum dots) have received great attention in both basic and technical contexts, where they can be applied in a variety of technologies, such as: light emitting devices (Colvin et al, Nature 370, 354-. See, for example, Bruchez et al, Science, Vol.281, pp.2013-2015, 2001; chan & Nie, Science, Vol.281, 2016-; U.S. Pat. No.6,207,392 to Klarreich, Nature, Vol.43, pp.450-452, 2001; see also Mitchell, Nature Biotechnology, pp.1013-1017, 2001, and U.S. Pat. Nos. 6,423,551, 6,306,610 and 6,326,144.

The excellent luminescent properties of quantum dots are derived from the quantum size confinement (quantum size confinement) that occurs when the metal and semiconductor core particles (core particles) are about 1 to 5nm smaller than their excitation bohr radii. (Alivisatos, Science, 271, 933-37, 1996; Alivisatos, J.Phys.Chem.100, 13226-39, 1996; Brus, Appl Phys., A53, 465-74, 1991; Wilson et al, Science, 262, 1242-46, 1993).

The most extensively studied semiconductor nanocrystal materials are chalcogenide (chalcogenide) II-VI materials and III-V materials. The main reason for the interest in these semiconductor nanocrystal materials is that photoluminescence can be made across the entire visible spectrum by controlling the particle size.

In view of the above, it is desirable to find further applications for nanocrystals.

Drawings

The invention will be better understood by reference to the detailed description when taken in conjunction with the non-limiting examples and the accompanying drawings.

FIG. 1 shows an orchid, Dendrobii White Fairy, which is dip-coated (dip coat) in a solution containing CdZnSe nanocrystals. The picture shows that the orchid under UV lamp has excitation at 302/345 nm. The orchid flowers appear orange after immersion in the solution, compared to the natural white/purple color of the orchid shown in the smaller picture at the upper left of figure 1.

Fig. 2 the vial shown in the upper left picture contains a solution containing CdZnSe nanocrystals dissolved in octadecene. The top right picture shows white dendrobium candidum before uptake of CdZnSe nanocrystals by the stem (stem) of the plant. The lower panel shows the orchid after uptake of CdZnSe nanocrystals and irradiation with an Ultraviolet (UV) lamp, which has an excitation at 302/345 nm. The colored orchid has the same color as the orchid shown in figure 1, i.e. orange.

Figure 3 shows, in the upper left picture, a vial containing an aqueous solution containing fluorescein. The top right picture shows white dendrobium candidum before fluorogenic yellow uptake by the roots of the plant. The lower panel shows the orchid after taking the fluorescent yellow and illuminated with a UV lamp, which has an excitation at 302/345 nm. The colored orchid appears green, wherein the color is concentrated in vascular bundles (vascular strand), also called veins (vena veils) or vein networks (vena veils).

FIG. 4a shows Dendrobii nobilis albo-marginata after uptake of fluorescein and CdZnSe nanocrystals. In fig. 4a, the orange color of the nanocrystals is visible in the area of the ovary (ovary), slightly visible in the base of the petals and sepals (base), and under the action of the fluorescence of the fluorescein, the veins appear green. Figure 4b shows a cross section of the stem. Fig. 4b shows a true color image based on 32 channel/32 color gamut. The 6 grey spots (more clearly in fig. 4 c) around the center of the section of the stem appeared green under the fluorescence of fluorescein, while the surrounding parts appeared orange due to the luminescence of the CdZnSe nanocrystals. Fig. 4c shows the same cross section of the stem, but with a color filter, only the light emitted by fluorescein (c) is shown, while fig. 4d only shows the light emitted by the CdZnSe nanocrystals.

Detailed Description

In a first aspect, the present invention relates to a plant containing or incorporating at least one nanocrystal (quantum dot). Depending on the nanocrystals used to dye the plant, the plant containing or incorporating the semiconductor nanocrystal material emits light of a particular wavelength.

The term "plant" includes organisms whose cells contain plastids in addition to the nucleus (with a double nuclear membrane and some chromosomes). Plastids exist as chloroplasts, or can be converted to chloroplasts under appropriate conditions. Unlike animals and all other heterotrophs, green plants are phototrophic. The term plant as used herein does not include unicellular organisms (e.g., cyanobacteria) or unicellular cells, which are sometimes also referred to as plants. The plants of the invention also exclude moss, lichens and fungi, which are sometimes classified as plants although they do not have plastids.

In another aspect of the present invention, the nanocrystals may also be contained in or on a dry plant or an artificial plant. This means that the term plant as used herein refers not only to living plants but also to non-living or life-lost plants. Such plants include dry plants and artificial plants. The dry plant may be selected from the plants mentioned further below or parts of the plants described in further detail below. Artificial plants are commonly used for decorative purposes and may be made of polymers or mixtures of polymers, paper or board (laminated or not), metal foil or mixtures of the above materials. The artificial plant may be a replica of one of the plants listed below or a new form of a newly created plant that does not exist in that form in nature.

The artificial plant may be made of silk, rayon (artificial silk made of, for example, rayon (rayon), mercerized cotton, polyester, or a mixture of these materials, or a mixture of rayon and silk), polymer-modified silk (for example, collagen-modified silk), polymer-modified cotton (for example, mercerized cotton), or a pure polymer (for example, nylon, polyamide, or polyester).

A plant contains multiple organs capable of growing outward in large quantities. Such organs include roots, leaves and flowers. In one aspect of the invention, the nanocrystals are contained in or associated with aerial (aerial) parts of a plant, which refers to parts of a plant that are present on the soil in which the plant is growing, such as stems, branches, flowers and leaves. In one example, the aerial parts refer to flowers or leaves of a plant.

In one example, the above-described plants of the invention are flowering plants, i.e. plants that either flower at least once at a particular stage of the plant's life or possess at least one flowering stage. The phrase "flowering plant" generally refers to any of more than 250000 angiosperms (magnolia) having roots, stems, leaves and well-developed conducting tissues (xylem and phloem). Angiosperms are generally distinguished from gymnosperms by the seeds produced being in a closed chamber (ovary) within the flower, but the distinction is not so clear-cut in any case. Angiosperma includes two classes: monocotyledons (monocots) and dicotyledons (dicots). The petals of monocotyledons (flower parts) are multiples of 3 (in threes), the ducts (connecting strands) are dispersed in the stem, and the leaves are mostly parallel veins and are not layered. The petals of dicotyledons are 4 or 5 times, the ducts are arranged in a column (cylinder), the inside of the leaves is a reticular vein, and a cambium is formed. Flowering plants exhibit great diversity in habit, size and shape; flowering plants exceed 300 families and grow in various continents, including antarctica. Flowering plants can adapt almost to various living environments. Most flowering plants are sexually propagated by specific propagation of organs present in all flowers with the aid of seeds.

In one aspect of the invention, the flowering plants include, but are not limited to, plants from: rosaceae (rosaceae), hydrangeaceae (hydrangeaceae), Orchidaceae (orchidaceae), hydrangeaceae (hydrangeaceae), Hyacinaceae (hyacinthaceae), Begoniaceae (begoniaceae), Labiatae (lamiaceae) (including, for example, Lavandula (genus Lavandula)), Compositae (asteraceae) (including, for example, Dalia (genus dahlia) and Sardinia (genus gera)), Geraniaceae (geraniaceae) (including, for example, Geranium (genus Geranium)), Solanaceae (solanaceae) (including, for example, Phaneria (genus), Liliaceae (liaceae) (including, for example, Geranium (genus), Liliaceae) (including, for example, primula (genus), and Pleurotus (genus), Liliaceae (family), Hypomeiaceae) (including, for example, Pleuroidea) (including, and Pleuroidea) (including, Eugenia (genus), Eupatorium (family Cornaceae) (including, family Hypomeiaceae), and Labiatae (family), Violaceae (family), family (family), family Verbenaceae) (including, family (family), family (family), family Primulaceae) (including, genus), family (genus), family Malvaceae) (including, family (genus), family (family), family (Rosaceae) (including, family), family (family), family (, Ericaceae (ericaceae) (including, for example, rhododendron (genus rhododendron) (including azalea (azalea))), theaceae (theaceae) (including, for example, camellia (genus camellia)) or heliothis (family marmalaceae) (including, for example, phyllanthus (genus bougainvillea)).

In one example, the plant includes, but is not limited to: rose, orchid, hydrangea (hydrangea), hyacinth, begonia, lavender, dahlia, geranium, petunia, tulip, lily, primula, viola (viola), gerbera, glorybower (gloxinia), hibiscus, pineapple (broomariad), cyclamen, anthurium (anthurium), azalea, leafflower (bougainvillea) or camellia.

The orchids include the orchidaceae (Orchidoideae) subfamily, the Dracocephalum (Epidendoideae) subfamily, the Vanilla (Vanilloideae) subfamily, the Cypripedia (Cypripedioideae) subfamily, and the Asteroides (Apostasioideae) subfamily. The Cypripedioideae subfamily includes, for example: the family cypripedioae, mexipedia, paphiopedilum americanum (phragmipedioae) and the family celenie shoe blue (selephiediae). The dendron (epindoideneae) subfamily includes, for example: the family of the trees (arethusae), the family of the spoon lip (callpsoaeae), the family of the clairvoya (cryptarrhenae), the family of the fritillaria (coelogyne), the family of the trees (epiplogeae), the family of the tiger tongue (epiplogeae), the family of the gastrodia (gastroodiae), the family of the macrobrachium (Malaxideae), the family of the bird nest (Neottieae), the family of the handle lip (podophilae), the family of the bambusa stem (tropideae) and the family of the ericssole (Xerorchideae). The vanillia (Vanilloideae) subfamily includes, for example: the subfamilies of cymbidium (Pogonineae) and vanilla (Vanilleae). The orchid (Orchidoideae) subfamily includes, for example: the dicerotala (dicerosteleae) family, the codonorchiae (Codonorchideae) family, the clavicle (cratichidae) family, the dieae family, the Diurideae family, and the orchid family.

The above family of the orchidaceae, the labiatae pedunculata (Podichileae), includes, for example: the dendrobii (Dendrobiinae) subfamily, which includes, for example, the genera Kaderi (Cadetia), Dendrobii (Dendrobium), Diclonobium (Diplocybium), Epimedium (Epimenium), Tephram (Flickingeria) and Purpurera (Pseudocerana). For example, an orchid plant, dendrobium candidum (Dendrobii white Fairy), used in one embodiment of the invention belongs to the genus Dendrobium.

The type of nanocrystals used in the present invention may contain or consist of a semiconductor material or a magnetic iron oxide (magnetic oxide) including ternary or higher systems, such as: a metal ferrite. The emission of these nanocrystals (quantum dots) can be tuned by composition and/or size.

The at least one nanocrystal for dyeing a plant may include, but is not limited to:

a rare earth metal-doped metal oxide of the formula MeAl₂O₄Re or Me₄Al₁₄O₂₅R; or

A binary nanocrystal having a chemical formula of M1A or M1O; or

A ternary nanocrystal having a chemical formula of M1M2A or M1AB or M1M 2O; or

A quaternary nanocrystal having a chemical formula of M1M2 AB;

wherein M1 and M2 in the binary, ternary, and quaternary nanocrystals are independently a metal, such as from group II, group III, group IV, or group VIII in the Periodic System of Elements (PSE); and a and B, if present, are, independently of each other, elements from, for example, group VI or group V in the PSE; and is

Wherein Me in the rare earth metal doped metal oxide may include, for example: ca. Sr or Ba; and is

Re in the rare earth metal-doped metal oxide is at least one element, which may include Tb, Dy, Nd, Eu, or Tm.

Rare earth metal doped metal oxides are well known in the art and are described in detail in, for example, US6264855, JPH9-143463, JP 2543852, JP 2697733, JP 2697688, JP H10-273654, JP 2929162. In short, the rare earth metal doped metal oxides as described above may be water resistant and may be prepared, for example, by: the luminescent pigment as a raw material is subjected to an acid treatment with at least one acidic compound, which may be an acid or a compound generating an acid at a pH of not more than 3, and then to a base treatment with at least one basic compound, which may be a base or a compound generating a base at a pH of 4 to 9.

If the luminescent pigment is a luminescent pigment doped with at least one rare earth metal element, in which the oxide matrix (matrix) contains at least one element such as Mg, Ca, Sr, Ba and Zn and at least one element such as B, Al, P or Ga, the luminescent pigment as the object of the acid treatment may be various types of luminescent pigments. The doping amount of the rare earth element is not particularly limited, but 0.0001 to 30 wt% is sufficient, or in one example, 0.1 to 10 wt% is sufficient with respect to the total amount of the luminescent pigment.

Examples of luminescent pigments include those of the formula MeAl₂O₄Re or Me₄Al₁₄O₂₅R. In the formula, Me may be Ca, Sr and Ba; and Re in the rare earth metal-doped metal oxide is at least one element such as Tb, Dy, Nd, Eu, and Tm. The preparation of the above rare earth metal doped metal oxides is described in detail in, for example, US 6264855.

Examples of binary nanocrystals of the formula M1A or M1O are well known in the art. M1 in the above formula may be a group II, III, IV, VII or VIII metal of the periodic System of Elements (periodic System of Elements) according to the convention International Union of theory and applied chemistry (IUPAC). According to the new IUPAC system, the corresponding groups (traditional system in parentheses) are group 2/12 (group II), group 13 (group III), group 14 (group IV), group 7 (group VII). Suitable examples of group II (group 12 of the new IUPAC system) are Cd, Zn and (group 2 of the new IUPAC system) Mg, Sr, Ca and Ba. Suitable examples of group III (group 13 of the new IUPAC regime) are Al, Ga and In. Two illustrative examples of suitable elements of group IV (group 14 of the new IUPAC regime) are Pb and Sn. An illustrative example of a suitable element of group VII (group 7 of the new IUPAC regime) is Mn. Suitable examples of group VIII (group 8 of the new IUPAC regime) elements are Fe, Co, Ni and Ir. In one embodiment of the present invention, the use of Pb and Cd is excluded in the preparation of binary nanocrystals.

Typically, M1, or, where appropriate, M1 and M2, may each be, for example, group II, group III, group IV, group VII, or group VIII, group II such as Cd, Zn, Mg, Sr, Ca, or Ba, group III such as Al, Ga, and In, group IV such as Pb, Sn, group VII such as Mn, group VIII such as Fe, Co, Ni, or Ir.

A in the above formula may be a chalcogen or pnictogen (pnictogen), i.e. an element of group VI or group V in the periodic system of the elements according to the IUPAC nomenclature used in the past, or an element of group 16 or group 15 according to the new IUPAC nomenclature. As such, the reaction mixture used to prepare such binary nanocrystals can include a metal precursor containing metal M1 as well as element a. The metal precursor may include a salt that provides the corresponding metal during the formation of the nanocrystal. The salt may be, for example, an inorganic (e.g., carbonate) or organic (e.g., acetate, stearate or oleate) salt of the corresponding metal. Wherein two metals or metal precursors are used to prepare such binary nanocrystal systems, for example: cadmium and zinc or their oxides, the two metals/precursors being present in any desired ratio.

The elements A and, where appropriate, B may be independently selected from group VI of the periodic system of the elements, such As S, Se, Te, O, or from group V of the periodic system of the elements, such As P, Bi or As.

The metal precursor used to prepare the binary nanocrystal may be a metal oleate. The metal oleate may include, but is not limited to, cadmium oleate, cadmium zinc oleate (cadnium zinc oleate), lead oleate, manganese oleate, magnesium oleate, lead cadmium oleate (cadnium lead oleate), magnesium iron oleate (magnesium iron oleate), manganese iron oleate (manganese iron oleate), calcium iron oleate (calcium iron oleate), zinc iron oleate (zinc iron oleate), strontium iron oleate (strontium iron oleate), cobalt iron oleate (cobalt iron oleate), or nickel iron oleate (nickel iron oleate).

Examples of binary nanocrystals include, but are not limited to, CdSe, CdTe, CdS, PbSe, PbTe, PbS, SnSe, ZnS, ZnSe, or ZnTe. The binary nanocrystal of formula M1O can be, for example, of the formula CdO, PbO, MnO, CoO, ZnO or FeO. It is noted that the expressions M1A and M1O are used herein only to illustrate that the nanocrystals are binary, indicating that they contain two elements. Expressions M1A and M1O do not necessarily represent the stoichiometric ratio of the nanocrystals (although the nanocrystals can be CdSe or CdO, to name just two examples), but expressions M1A and M1O can also include a stoichiometric ratio of MO₂(e.g., MnO)₂)、M₂O₃(e.g., Al)₂O₃) Or M₃O₄(e.g., Fe)₃O₄) The nanocrystals of (1).

Some studies have shown that the luminescence of nanocrystals can be enhanced by encapsulating a core particle (core particle) of tunable size with a lower band gap with a shell of an inorganic material with a higher band gap. For example, CdSe quantum dots passivated with a ZnS layer emit strongly at room temperature and their emission wavelength can be tuned from blue to red by changing the particle size. And the ZnS encapsulating layer passivates non-radiative recombination sites (nnondiaretic recombination sites) on the surface, thereby making the nanocrystals (quantum dots) more stable. (Dabbosi et al, J.Phys.chem.B101, 9463-75, 1997 Kortan et al, J.am.chem.Soc.112, 1327-.

The ternary nanocrystal may have the general formula M1M 2A. M1 and M2 in the above formula are independently metals of group II, group III, group IV, group VII or group VIII of the periodic system of the elements. A may be a group VI or group V element in the periodic system of the elements. In other examples, the ternary nanocrystal may have the formula M1 AB. M1 in the above formula may be a metal of group II, group III, group IV, group VII or group VIII of the periodic system of the elements. A and B independently can be an element of group V or group VI of the periodic system of the elements. In another example, the ternary nanocrystal formed according to the method of the present invention may have the general formula M1M 2O. M1 and M2 independently can be a metal of group II, group III, group IV, group VII or group VIII of the periodic system of the elements. O is oxygen. The ternary nanocrystals can be of various structures. The ternary nanocrystal may, for example, be a layered structure (e.g. a core/shell structure), or the ternary nanocrystal may be homogeneous (homogenes), for example: with a uniform composition or a composition that changes gradually or continuously (steples).

The quaternary nanocrystal formed may have the general formula M1M2 AB. As defined above, M1 and M2 may be metals of group II, group III, group IV, group VII or group VIII of the periodic system of the elements. A and B independently can be an element of group V or group VI of the periodic system of the elements.

Illustrative examples of ternary nanocrystals of the general formula M1M2A include, but are not limited to: the molecular formula is ternary nanocrystalline of ZnCdSe, CdZnS, CdZnSe, CdZnTe, SnPbS, SnPbSe and SnPbTe. Illustrative examples of ternary nanocrystals of formula M1AB can include nanocrystals of the formulae CdSeS, CdSeTe, CdSTe, ZnSeS, ZnSeTe, ZnSTe, SnSeS, SnSeTe, SnSTe, PbSeS, PbSeTe, and PbSTe. Illustrative examples of ternary nanocrystals of the formula M1M2O can include those of the formula M1M2₂O₄For example: ferrite MgFe₂O₄、CaFe₂O₄、SrFe₂O₄、ZnFe₂O₄、NiFe₂O₄、CoFe₂O₄And MnFe₂O₄. It is also noted that the expressions M1M2A, M1AB, and M1M2O are used herein only to illustrate that these nanocrystals are ternary, indicating that they contain three different elements. Accordingly, the stoichiometric ratio of the nanocrystals cannot be derived from the expressions M1M2A, M1AB, and M1M2O, even in some occasional cases, such as CdZnSe, CdSeTe or MgFe₂O₄In embodiments of (a), the stoichiometric ratio of the nanocrystals can be quite accurately mapped into the formula. Thus, the above formulae M1M2A, M1AB and M1M2O also include, for example, the stoichiometric ratio M1_1-xM2_xA (e.g., Cd)_1-xZn_xSe)、M1_xM2_1-xA (e.g. Zn)_xCd_1-xSe) or M1_xA_yB_1-y(e.g., CdSe)_yS_1-y)、M1_xA_1-yB_y(e.g., CdSe)_1-yS_y) Or M1_1-xM2_xO (e.g. Mg)_1-xF_exO) or M1_xM2_1-xO (e.g. Fe)_xMn_1-xO) of a crystalline material.

Illustrative examples of quaternary nanocrystals having the general formula M1M2AB can include nanocrystals of CdZnSeS, CdZnSeTe, CdZnSTe, CdHgSeS, CdHgSeTe, CdHgSTe, HgZnSeS, HgZnSeTe, and HgZnSTe. It is also noted that the expression M1M2AB is used herein only to illustrate that the nanocrystal is quaternary, indicating that the nanocrystal contains four elements. The above expression M1M2AB does not necessarily indicate the stoichiometric ratio of the nanocrystal (even when the nanocrystal is CdZnSeS, to name just one example), but also includes the stoichiometric ratio M1_1-xM2_xA_yB_1-y(e.g., Cd)_1-xZn_xSe_yS_1-y) Or M1_1-xM2_xA_1-yB_y(e.g., Cd)_1-xZn_xSe_1-yS_y) The nanocrystals of (1).

For example, uniform ternary nanocrystals (M1. chem. Soc., 2003125, 8598-8594, J.Am. chem. Soc., 2003125, 13559-13553, or International patent application WO2004/054923, to Zhong et al have been described_1-xM2_xA) Or quaternary nanocrystals (M1)_1-xM2_xA_yB_1-y) Examples of (3).

It should also be noted that whether binary, ternary, or quaternary nanocrystals, when used independently, the nanocrystals can be of various desired configurations. The nanocrystals may be, for example, layered structures such as core/shell structures or core-bulk-shell structures (core-shell structures), (Hines, M.A., & Guyot-Sinonest, P., J.Phys.chem. (1996)100, 468; Dabbosi, B.O., et al, J.Phys.chem.B (1997)101, 9463; Peng, X. et al, J.Am.chem.Soc. (1997)119, 701916-18) or various alloy structures (see, for example, U.S. Pat. No. 7056471), the alloy structure includes an alloy-gradient structure (Foley, J et al, Materials scienform, Vol. 225-, as described in co-pending PCT application PCT/SG2008/000290, "Pprocess Of Forming A catalyst And growing semiconductor continuous confinement Nanocristalline Composite And Nanocristalline Composite Obtained from".

The nanocrystals can be made using a method comprising:

(i) forming a homogeneous reaction mixture containing a metal precursor containing metal M1, element A, and a low boiling point nonpolar solvent, and

(ii) the homogeneous reaction mixture is subjected to an elevated temperature suitable for forming nanocrystals under elevated pressure.

In this process, the low boiling non-polar solvent has a boiling point below about 100 ℃ or 80 ℃ at atmospheric pressure (1013 millibars (mbar)). Examples of such solvents include, but are not limited to: hexane, chloroform, toluene, benzene, heptane, cyclohexane, dichloromethane, pyridine, carbon tetrachloride, carbon disulfide, dioxane, diethyl ether, diisopropyl ether, tetrahydrofuran, and mixtures thereof.

Surfactants may also be added to the process, for example: organic carboxylic acids, organic phosphates, organic phosphonic acids, organic phosphine oxides, organic amines or mixtures thereof. Example 2 gives an example of the preparation of such nanocrystals.

Water-soluble nanocrystal

In one embodiment of the invention, the nanocrystals used are water-soluble. Water-soluble nanocrystals are known in the art and are described, for example, in WO 2006/118543, WO 2006/118542, and WO 2006/075974.

In addition, PCT publication WO 00/17656 discloses core-shell nanocrystals having the formula SH (CH) respectively₂)_n-COOH and SH (CH)₂)_n-SO₃The carboxylic or sulfonic acid compound of H is coated to make the nanocrystal water soluble. Similarly, PCT application WO 00/29617 and british patent application GB 2342651 describe the attachment of organic acids, such as thioglycolic acid or mercaptoundecanoic acid, to the surface of nanocrystals to render them water soluble and suitable for binding to biomolecules, such as proteins or nucleic acids. British patent application 2342651 also describes the use of trioctylphosphine as a covering material (capping material), which is believed to impart water solubility to the nanocrystals.

In another document, PCT publication WO 00/27365 reports the use of a diaminocarboxylic acid or an amino acid as a water-solubilizing agent for nanocrystals (water-solubilizing agent). PCT publication WO00/17655 discloses that nanocrystals are made water-soluble by using a solubilizing agent having a hydrophilic portion and a hydrophobic portion. The solubilizer is linked to the nanocrystal through the hydrophobic group, while the hydrophilic group provides water solubility, such as carboxylic acid or methacrylic acid. Another PCT application (WO02/073155) describes water-soluble semiconductor nanocrystals in which various molecules, such as trioctylphosphine hydroxamate, hydroxamic acid derivatives or multidentate complexing agents (e.g. ethylenediamine) are attached directly to the surface of the nanocrystal to render the nanocrystal water-soluble. These nanocrystals can then be attached to proteins by pulse discharge sintering (EDC). In another approach, PCT application WO 00/58731 discloses nanocrystals for blood cell population (population) analysis in which an amino-derived polysaccharide (amino-derivatized polysaccharide) having a molecular weight of about 3000-3000000 is attached to the nanocrystal.

U.S. patent 6699723 discloses the use of silane-based compounds as linkers to facilitate the attachment of biomolecules, such as biotin and streptavidin, to luminescent nanocrystal probes. U.S. patent application No.2004/0072373a1 describes a biochemical labeling method using silane-based compounds. The silane-attached nanoparticles are bound to the template molecule by molecular imprinting and then polymerized to form a matrix (matrix). Thereafter, the template molecule is removed from the matrix. The cavities formed on the matrix as a result of removing the template molecules have properties that can be used for labeling.

Recently, the use of synthetic polymers to stabilize water-soluble nanocrystals has been reported. U.S. patent application No.2004/0115817a1 describes that amphiphilic diblock polymers can be non-covalently attached by hydrophobic interactions to nanocrystals whose surfaces are covered with agents such as trioctylphosphine or trioctylphosphine oxide. Similarly, Gao et al (Nature Biotechnology, Vol.22, 969-976, 8.2004) disclose water-soluble semiconductor nanocrystals that are encapsulated with amphiphilic triblock copolymers by non-covalent hydrophobic interactions.

1. In one example, to obtain water-soluble nanocrystals, a capping reagent (cappingreagent) is attached to the surface of the nanocrystal. The capping reagent forms a guest-host complex with the water-soluble molecule.

Various nanocrystals can be used, the surface of which can be reacted with a capping reagent having (terminal) groups with affinity for (the surface of) the nanocrystal core, to obtain water-soluble nanocrystals. Accordingly, the capping reagent typically forms covalent bonds with the surface of the nanocrystal. In the case of core-shell nanocrystals, covalent bonds are typically formed between the capping reagent and the shell of the nanocrystal. If uniform ternary or quaternary nanocrystals as described in WO2004/054923 are used, covalent bonds are formed between the surface of the uniform core and the capping reagent. The nature of the capping reagent may be substantially hydrophilic or substantially hydrophobic, depending, for example, on the hydrophobicity (or hydrophilicity) of the internal cavity of the host molecule. In this respect, it should be noted that the term "(substantially) hydrophobic molecule" is also meant to include molecules which may contain hydrophilic moieties in addition to hydrophobic moieties, as long as the hydrophilic moieties do not interfere with the hydrophobic moieties of the molecule (i.e. the capping reagent) forming guest-host complexes with host molecules having a hydrophobic internal cavity. Likewise, the term "(substantially) hydrophilic molecule" includes molecules that may contain hydrophobic moieties in addition to hydrophilic moieties, so long as the hydrophobic moieties do not interfere with the formation of a guest-host complex of the hydrophilic moieties of the molecule (i.e., capping reagent) with a host molecule having a hydrophilic internal cavity.

In one example, the formula of the capping reagent used for "surface capping" is:

H_AX-Y-Z，

wherein X is a terminal group selected from S, N, P or O ═ P, a is an integer from 0 to 3, Y is a group having at least three main chain atoms, and Z is a hydrophobic terminal group which can form a host-guest inclusion complex (host-guest inclusion complex) with a suitable host molecule (host molecule).

Typically, the Y group of the capping reagent contains 3 to 50 backbone atoms. The Y group may contain essentially any suitable group that imparts an outstanding hydrophobic character to the reagent. Examples of suitable groups that may be used in Y include: alkyl radicals (e.g. CH)₂-), cycloalkyl groups (for example: cyclohexyl), ether groups (for example: -OCH₂CH₂-, or an aromatic group (for example: benzene or naphthalene rings) to name only a few. The group Y may be linear or branched and may also have substituents replacing the main chain atoms. Z may be-CH₃Phenyl (-C)₆H₅) -SH, hydroxylAn (OH) group, an acidic group (e.g.: SO)₃H、PO₃H or-COOH), a basic group (e.g.: NH (NH)₂Or NHR₁Wherein R is CH₃or-CH₂-CH₃) Halogen (-Cl, -Br, -I, -F), -OH, -C ≡ CH, -CH ═ CH₂Trimethylsilyl group (-Si (Me))₃) Ferrocenyl or adamantyl (adamantine group), to name a few.

In some examples, for example, the following compounds are used as capping agents: CH (CH)₃(CH₂)_nCH₂SH、CH₃O(CH₂CH₂O)_nCH₂SH、HSCH₂CH₂CH₂(SH)(CH₂)_nCH₃、CH₃(CH₂)_nCH₂NH₂、CH₃O(CH₂CH₂O)_nCH₂NH₂、P((CH₂)_nCH₃)₃、O＝P((CH₂)_nCH₃)₃Wherein n is an integer of 30. gtoreq.n.gtoreq.6. In other embodiments, n is an integer greater than or equal to 30 and greater than or equal to n.gtoreq.8.

In this regard, it should be noted that examples of cover reagents that provide a more hydrophobic or substantially hydrophobic character include, but are not limited to: 1-mercapto-6-phenylhexanoic acid (HS- (CH)₂)₆Ph), 1, 16-dimercapto-hexadecane (HS- (CH)₂)₁₆-SH), 18-mercapto-octadecylamine (HS- (CH)₂)₁₈-NH₂) Trioctylphosphine or 6-mercapto-hexane (HS- (CH)₂)₅-CH₃)。

Typical capping reagents that provide a more hydrophobic or substantially hydrophobic character include, but are not limited to: 6-mercapto-hexanoic acid (HS- (CH)₂)₆-COOH), 16-mercapto-hexadecanoic acid (HS- (CH)₂)₁₆-COOH), 18-mercapto-octadecylamine (HS- (CH)₂)₁₈-NH₂) 6-mercapto-hexylamine (HS- (CH)₂)₆-NH₂) Or 8-hydroxy-octanethiol (HO- (CH)₂)₈-SH)。

Various host molecules may be used as long as the host molecules are capable of reacting with the capping reagent and rendering the complex formed between the capped nanocrystals and the host molecules water soluble. Typically, the host molecule is a water-soluble compound that contains a solvent exposed to polar groups, such as hydroxyl groups, carboxylate groups, sulfonate groups, phosphate groups, amine groups, amide groups (carboxamide groups), and the like.

Examples of suitable host molecules include, but are not limited to: carbohydrates (carbohydrate), cyclic polyamines (cyclic polyamines), cyclic peptides (cyclic peptides), crown ethers, dendrimers (dendrimers), and the like.

Examples of cyclic polyamines that can be used as the main molecule include, for example: tetraazamacrocycle molecules (teterazamacrocycle molecules) and derivatives thereof, tetraazamacrocycle molecules such as 1, 4, 8, 11-tetraazacyclotetradecane (also known as cyclam), derivatives of tetraazamacrocycle molecules such as 1, 4, 7, 11-tetraazacyclotetradecane (isocyclam), 1- (2-aminomethyl) -1, 4, 8, 11-tetraazacyclotetradecane (scorpiond), 1, 4, 8, 11-tetraazacyclotetradecane-6, 13-dicarboxylate, Sroczynski and Grzejdiak in J.Incl.Phenom.Macrocyclic chem.35, 251-260, 1999 or Bernhardt et al in J.Aus.Chem.56, 679-684, 2003; hexaazamacrocycle complexes (Hausmann, j. et al, Chemistry, aeuropen Journal, 2004, 10, 1716; Piotrowski, t. et al, Electroanalysis, 2000, 12, 1397); or octaazamacrocyclic compounds (Kobayashi, K. et al, J.Am.chem.Soc.1992, 114, 1105). The octaazamacrocycles described above by Kobayashi, k. et al are also an example of compounds suitable for accommodating polar guest molecules (e.g., hydrophilic capping reagents). Cyclic polyamines which are water-soluble only to a limited extent can also be used, for example: 5, 5, 7, 12, 14, 14-hexamethyl-1, 4, 8, 11-tetraazacyclotetradecane (Me6cylcam), and the cyclic polyamines can be modified with substituents that provide polar groups, such as carboxylate or sulfonate groups. Other examples of macrocyclic amines that can be used as the primary molecule are the compounds described in Odashima, K., Journal of incorporation phenomena and molecular biology in chemistry, 1998, 32, 165.

Examples of suitable calixarenes include: 4-tert-butyl-calix [4] arene tetraacetic acid tetraethyl ester described by dondondononi et al in chem.eur., j.3, 1774, 1997; tetragalacturonicarene (Davis, AP. et al, angelw. chem. int. edit., 1999, 38, 2979); octaaminoamide resorcinol [4] -arene (octaaminoamide resorcin [4] -arene) (Kazakov, E.K., et al, Eur.J.Org.chem., 2004, 3323); 4-sulfonic acid based calix [ n ] -arene (Yang, w.z., j.pharm. pharmacology, 2004, 56, 703); sulfonated thiacalix [4 or 6] -arene (sulfonated thiacalix [4 or 6] -arene) (Kunsasgi-Mate S., Tetrahedron Letters, 2004, 45, 1387); calixarenes described by Kobayashi et al in J.Am.chem.Soc.116, 6081, 1994 and by Yanagihara et al in J.Am.chem.Soc.114, 10307, 1992.

Examples of cyclic peptides that can be used as host molecules in the present invention include, but are not limited to: calixarenes with bicyclic diacids (dicyclodepiptide) described by Guo, W et al in Tetrahedron Letters, 2002, 43, 5665 or by Peng Li et al in Current Organic Chemistry, 2002, 6.

The crown ether which may be the main molecule may have any ring size, for example: having ring systems containing 8, 9, 10, 12, 14, 15, 16, 18 or 20 atoms, some of which are common heteroatoms such as O or S. Common crown ethers used herein include, but are not limited to: water-soluble 8-crown-4 compounds (wherein 4 represents the number of hetero atoms), 9-crown-3 compounds, 12-crown-4 compounds, 15-crown-5 compounds, 18-crown-6 compounds and 20-crown-8 compounds. Examples of suitable said crown ethers include (18-crown-6) -2, 3, 11, 12-tetracarboxylic acid or 1, 4, 7, 10-tetraazacyclododecane-1, 4, 7, 10-tetracarboxylic acid, to name a few.

In principle, various water-soluble dendrimers can provide hydrophilic or hydrophobic cavities capable of accommodating at least part of the capping reagent used herein (depending on whether the capping reagent used is hydrophobic or hydrophilic). Suitable types of dendrimers include, but are not limited to: polymethylaziridine dendrimers, polyamidoamine dendrimers, polyarylether dendrimers, polylysine dendrimers, saccharide dendrimers and silicon dendrimers (for example as reviewed by Boas and Heegard in chem. soc. rev.33, 43-63, 2004).

In one example, the nanocrystals contain a carbohydrate as a primary molecule. Such a carbohydrate host molecule may be, but is not limited to: oligosaccharide, starch or cyclodextrin molecules (see Davis and Wareham, Angew. chem. int. Edit.38, 2979-.

In some examples where the host molecule is an oligosaccharide, the oligosaccharide may contain 2-20 (e.g., 6) monomeric units in the backbone. These oligomers may be linear or branched. Examples of suitable oligosaccharides include, but are not limited to: 1, 3- (dimethylene) benzenediyl-6, 6 '-O- (2, 2' -oxydiethylene) -bis- (2, 3, 4-tri-O-acetyl-. beta. -D-galactopyranoside), 1, 3- (dimethylene) benzenediyl-6, 6 '-O- (2, 2' -oxydiethylene) -bis- (2, 3, 4-tri-O-methyl-. beta. -D-galactopyranoside (Shizuma et al, J.org.chem.2002, 67, 4795), cyclotris- (1, 2, 3, 4, 5, 6) - [ alpha-D-glucopyranosyl- (1, 2, 3, 4) -alpha-D-glucopyranosyl ] (Cescutti et al, Carbohydre Research, 2000, 329, 647), ethynyl sugars (acetylenosaccharoides) (Burli et al, angelw. chem. int. edit.1997, 36, 1852), or cyclic fructooligosaccharides (fructooligosaccharides) (Takai et al, j.chem. soc. chem. commu., 1993, 53).

If starch is used as the host molecule, the molecular weight of the starch may be about 1000-. In some examples, the starch has a molecular weight of about 4000Da ≧ Mw ≧ about 2000 Da. The starch used may also include amylose, for example: alpha-amylose or beta-amylose.

Examples of cyclodextrins suitable as host molecules include: alpha-cyclodextrin, beta-cyclodextrin, gamma-cyclodextrin, dimethyl-alpha-cyclodextrin, trimethyl-alpha-cyclodextrin, dimethyl-beta-cyclodextrin, trimethyl-beta-cyclodextrin, dimethyl-gamma-cyclodextrin and trimethyl-gamma-cyclodextrin.

Methods of preparing such water-soluble nanocrystals are known in the art and are described, for example, in WO 2006/075974. Briefly, such methods involve reacting a nanocrystal having a nanocrystal core with a capping reagent to attach the capping reagent to the surface of the nanocrystal core, and then contacting the resulting nanocrystal with a host molecule to form a guest-host complex between the reagent and the water-soluble host molecule. The (capping) agent may be hydrophilic or hydrophobic. The same reaction can be carried out to prepare the nanocrystals if pure metal nanocrystals or uniform ternary or quaternary nanocrystals are used. Example 3 provides an example of the preparation of such water-soluble nanocrystals.

2. In another example, at least one of the nanocrystals contains a capping reagent that is attached to the surface of the nanocrystal to form a first layer, the capping reagent having at least one coupling group (coupling group). Furthermore, a polymer forming the second layer and having at least one coupling group is covalently coupled to the at least one coupling group of the capping reagent.

In one embodiment, the polymer comprises at least one coupling moiety reactive with a coupling group of the capping reagent, the polymer having the formula:

wherein J is a coupling moiety reactive with at least one coupling group of the capping reagent and m is an integer of at least 1.

To illustrate this example, if, for example, the first layer has an amino-terminated group, the polymer forming the second layer may have a carboxyl group for covalent coupling with the amino group of the first layer. In fact, it is possible that not all of the coupling moieties (coupling moieties) and coupling groups (coupling groups) present may participate in the covalent coupling. For example, 50% of the carboxyl groups may be polymerized with the amino groups in the first layer

In another example, if the first layer has a carboxyl-terminated surface, the second layer can have an amino group covalently coupled to the carboxyl group of the first layer. It is also possible that not all coupling moieties and coupling groups present may participate in the covalent coupling. For example, 50% of the amino groups may be polymerized with the carboxyl groups in the first layer.

In another example, the polymer contains at least two coupling moieties that react with at least one coupling group of the capping reagent. In this case, the molecular formula of the polymer is:

wherein J and K are coupling moieties, said J and K are the same or different, and m and n are each an integer of at least 1.

Typically, if the capping reagent has both J and K end groups, the polymer may have one or both of K and J for covalent coupling with the capping reagent. For example, if the first layer has both a carboxyl-terminated surface and an amino-terminated surface, the second layer polymer may have only one of amino and carboxyl groups or both amino and carboxyl groups for covalent coupling with the carboxyl and amino groups of the first layer, respectively. It is sufficient that a part of the coupling moiety is covalently coupled to the coupling group without the need for the coupling moiety to be present in a precise stoichiometric ratio with respect to the coupling group.

In another example, the polymer contains at least three coupling moieties that react with at least one coupling group of the capping reagent. In this example, the polymer has the formula:

wherein J, K and L are coupling moieties, the J, K and L are the same or different, and m, n, and p are each integers of at least 1. In further embodiments, the polymer has 3 or more different functional groups (NH)₂COOH, NCO, CHO, etc.) for providing water solubility and a surface for coupling with the first layer.

The polymer forming the second layer will be in contact with the solvent in which the nanocrystals are located. Thus, in order to dissolve the nanocrystal in the solvent (which may contain water), for example, at least one of the coupling moieties J, K or L preferably contains hydrophilic groups that impart water solubility to the water-soluble shell. For this purpose, the polymer may also contain at least one moiety having a hydrophilic group that imparts water solubility to the water-soluble shell. This moiety may be present independently of the coupling moiety or the coupling moiety itself.

In one example, the coupling moieties J, K and L each contain a functional group selected from the group consisting of amino, hydroxyl, carbonyl, carboxyl, nitrile, isocyanate, and halogen groups. If it is desired to obtain homofunctional polymers, the said coupling part of the polymer may be constituted entirely by, for example, hydroxyl, carboxyl or amino groups. In this case, the polymers are polyvinyl alcohol, polycarboxylic acid and polyamine, respectively.

Other types of polymers having more than one monomer may be used in order to obtain nanocrystals having different properties (e.g., solubility in water). For example, a diblock copolymer, a triblock copolymer, or a mixed random polymer may be used as the polymer for forming the second layer. Specific examples include: poly (acrylic acid-b-methyl methacrylate) block copolymers, poly (methyl methacrylate-b-sodium acrylate) block copolymers, poly (t-butyl methacrylate-b-ethylene oxide) block copolymers, poly (methyl methacrylate-b-sodium methacrylate) block copolymers, and poly (methyl methacrylate-b-N, N-dimethylacrylamide) block copolymers.

The coupling moiety J in the above-described polymers of formula (III) may contain various suitable functional groups that react with the coupling groups in the capping reagent. The hydrophilic moiety K may contain various functional groups that impart outstanding hydrophilic properties to the polymer, thereby rendering the polymer water-soluble. Suitable examples of such functional groups include, for example, carboxyl groups, amino groups, hydroxyl groups, amide groups, ester groups, anhydride groups, and aldehyde groups.

In one example, the polymer is selected from the group consisting of polyamine, polyacetyl acid (polyacetyl acid), or polyol (polyol). The molecular weight of the polymer may range from below about 500 (about 400) to over about 1000000. In one embodiment, the molecular weight may range from about 600-. For in vivo applications, a lower limit of about 2000 may be selected to minimize potential toxicity to the human body.

If the capping reagent used contains a polymerizable unsaturated group as a coupling group, such unsaturated polymers may be used to form the second layer of the water-soluble shell, including polyacetylene, polyacrylic acid, polyethyleneimine.

In another example, to obtain water-soluble nanocrystals, at least one nanocrystal contains:

a capping reagent attached to the surface of the nanocrystal to form a first layer, the capping reagent having at least two coupling groups;

a low molecular weight coating reagent (coating reagent) forming a second layer and having at least two coupling moieties covalently coupled to the coating reagent; and

at least one water-solubilizing group for imparting water solubility to the second layer.

The capping reagent may contain an end group having affinity for the surface of the nanocrystal core. The end groups may include, but are not limited to: mercapto, amino, amine oxide (amine-oxide) and phosphino groups. The at least two coupling groups of the capping reagent may be spaced apart from the end group by a hydrophobic region. Each of the at least two coupling groups may contain a functional group independently selected from the group consisting of an amino group, a hydroxyl group, a carbonyl group, a carboxyl group, a nitrile group, a nitro group, an isocyanate group, an epoxy group, an anhydride group, and a halogen group.

The capping reagent may contain two coupling groups spaced apart from the terminal group by a hydrophobic region, the capping reagent being illustrated by the general formula:

in this formula, X represents an end group having an affinity for the surface of the nanocrystal (or sometimes referred to as nanocrystal core). X may be selected from S, N, P or O ═ P. Specific examples of the Hn-X-moiety may include, for example, H-S-, O ═ P-, and H₂N-is any one of the above. Ra is a moiety containing at least 2 backbone carbon atoms and thus has hydrophobic properties. If Ra has an outstanding hydrophobic character (e.g., hydrocarbons), Ra provides a hydrophobic region separating the Z moiety from the nanocrystal core. The Y moiety is selected from N, C, -COO-or-CH₂O-is formed. Z is a group containing at least one couplingPart of the hydrophilic coating agent is used for subsequent polymerization, thus giving the part of the hydrophilic coating agent outstanding hydrophilic properties. Typical polar functional groups include, but are not limited to: -OH, -COOH, -NH₂-CHO, -CONHR, -CN, -NCO, -COR and halogen. The symbols k, n' and m in the above formula represent numbers. K is 0 or 1. The number n is an integer from 0 to 3 and n' is an integer from 0 to 2; n and n' are selected to satisfy the respective valence requirements of X and Y. The number m is an integer from 1 to 3. The number k is 0 or 1. The applicable conditions are as follows: when k is 0, Z will bind to Ra. The value of K ═ 0 satisfies the following condition: the linking moiety Z is directly attached to Ra, where Ra is, for example, a cyclic moiety such as an aliphatic cycloalkane, aromatic hydrocarbon or heterocyclic ring. However, when k is 1, Ra is a cyclic moiety, for example: tertiary amino groups attached to a benzene ring or cyclic hydrocarbon. In the current formula, Z is a functional group selected from the group consisting of an amino group, a hydroxyl group, a carbonyl group, a carboxyl group, a nitrile group, a nitro group, an isocyanate group, an epoxy group, an anhydride group, and a halogen group. Either Y or Z may be used as a coupling group. If Z is present as a coupling group, Y may be used as a structural component (structural component) to which the coupling group Z is attached. If Z is not present, Y may form part of the coupling group.

The Ra moiety in the above formula may contain several tens to several hundreds of main chain atoms. In a particular embodiment, Ra and Z each independently contain from 2 to 50 backbone atoms. Z may contain one or more amide or ester linkages. Suitable examples of moieties that can be used as Ra include alkyl, alkenyl, alkoxy, and aryl.

The term "alkyl" as used herein refers to a branched or unbranched, straight or cyclic saturated hydrocarbon group, typically containing from 2 to 50 carbon atoms, for example: methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, octyl, decyl, tetradecyl, hexadecyl, eicosyl, tetracosyl and cycloalkyl (e.g. cyclopentyl, cyclohexyl). The term "alkenyl" as used herein refers to a branched or unbranched hydrocarbon group, typically containing from 2 to 50 carbon atoms and containing at least one double bond, typically containing from 1 to 6 double bonds, more typically containing from 1 to 2 double bonds, such as: vinyl, n-propenyl, n-butenyl, octenyl, decenyl, and cycloalkenyl (e.g., cyclopentenyl, cyclohexenyl). The term "alkoxy" as used herein refers to the substituent-O-R, where R is alkyl as defined above. The term "aryl" as used herein, unless otherwise defined, refers to an aromatic group containing one or more aromatic rings. The aromatic ring of the aryl group is optionally substituted with one or more inert, non-hydrogen substituents, suitable substituents including, for example: halogen, haloalkyl (preferably halogen-substituted lower alkyl), alkyl (preferably lower alkyl), alkenyl (preferably lower alkenyl), alkynyl (preferably lower alkynyl), alkoxy (preferably lower alkoxy), alkoxycarbonyl (preferably lower alkoxycarbonyl), carboxyl, nitro, cyano and sulfonyl. In all embodiments, Ra can include heteroaryl groups, which typically contain heteroatoms (e.g., nitrogen, oxygen, or sulfur).

In some examples, Ra may include, but is not limited to: ethyl, propyl, butyl and pentyl, cyclopentyl, cyclohexyl, cyclooctyl, ethoxy, propoxy, butoxy, benzyl, purinyl, pyridyl, imidazolyl.

In another example, the at least two coupling groups of the capping reagent can be homo-bifunctional (homo-bifunctional) or heterobifunctional (hetero-bifunctional), meaning that the capping reagent can contain at least two identical coupling groups or two different coupling groups, respectively. Some illustrative examples of suitable capping reagents having two or three coupling groups each have the following structure:

typical capping reagents in which the capping reagent is heterobifunctional (i.e., there are 2 different coupling groups) include, but are not limited to:

and

in another example, the capping reagent and the capping reagent are coupled via various free radical polymerization mechanisms through a polymerizable unsaturated group, such as a C ═ C double bond. Specific examples of such capping reagents include, but are not limited to: omega-thiol-terminated methyl methacrylate, 2-butenethiol, (E) -2-buten-1-thiol, S- (E) -2-butenylthioacetate (S- (E) -2-butenylthioacetate), S-3-methylbutenylthioacetate (S-3-methylbutenylthioacetate), 2-quinolinemethanethiol and S-2-quinolinemethylthioacetate (S-2-quinolinemethylthioacetate).

The second component of the water-soluble shell surrounding the nanocrystal core is formed by coupling a low molecular weight capping reagent bearing a water-soluble group with the capping reagent. Optionally, a coupling agent may be used to activate the coupling groups present in the capping reagent. The coupling agent and the coating agent with the coupling part can be added in sequence, namely the coating agent is added after the activation is finished; alternatively, the coating agent may be added simultaneously with the coupling agent.

The determination of suitable coupling agents is within the knowledge of one skilled in the art. An example of a suitable coupling reagent is 1-ethyl-3- [ 3-dimethylaminopropyl ] carbodiimide (EDC) used in combination with thio-N-hydroxysuccinimide (NHS). Other types of coupling reagents may also be used, including but not limited to: imides (imides) and pyrroles (azoles). Some examples of the diimides that can be used are: carbodiimides, succinimides, and phthalimides (pthalimides). Some notable examples of imides include: 1-ethyl-3- [ 3-dimethylaminopropyl ] carbodiimide (EDC), thio-N-hydroxysuccinimide, N ' -Dicyclohexylcarbodiimide (DCC), N ' -dicyclohexylcarbodiimide, N- (3-dimethylaminopropyl) -N ' -ethylcarbodiimide used in combination with N-hydroxysuccinimide or other activating molecule.

The capping reagent used to form the second layer of the water-soluble shell may contain one or more suitable coupling moieties that can react with activated coupling groups on the capping reagent. Generally, suitable capping reagents have at least 2 coupling moieties, i.e., in some examples, for example, 2, 3, or 4 functional groups that react with activated coupling groups in the capping reagent. When at least two coupling moieties of the capping reagent react with the capping reagent, the capping reagent is covalently bonded ("cross-linked") to the capping reagent by forming, for example, an ester or amide bond, thereby forming a water-soluble shell around the nanocrystal core.

The term "low molecular weight capping agent" as used in forming the second layer of the water-soluble shell includes non-polymeric (small) molecules. The molecular weight of the coating agent may be low or high depending on the type of groups present in the coating agent molecule. The molecular weight of the coating agent may be lower if the coating agent has, for example, small side chains. If there are bulky side chains in the coating agent, the molecular weight of the coating agent molecule will be higher. Accordingly, in some embodiments, the upper limit of the molecular weight of the coating agent is about 200 daltons, about 400 daltons, about 600 daltons, or about 1000 daltons. In other embodiments where a higher molecular weight or larger spatial volume capping reagent is used, the upper limit may be higher, such as about 1200 daltons, about 1500 daltons, or about 2000 daltons. As defined herein, the term "low molecular weight coating reagent" also includes oligomeric compounds, such as oligomeric compounds having a molecular weight of up to about 2000 daltons. The terms "coupling" and "covalent coupling" generally refer to various reactions that join two molecules to form a single molecule, a larger entity (entity), for example: coupling an acid with an alcohol to form an ester, or coupling an acid with an amine to form an amide. Accordingly, various reactions that can couple the coupling group and the coupling moiety present in the capping reagent and the capping reagent fall within the meaning of this term. "coupling" also includes reacting one or more unsaturated groups (e.g., -C ═ C-double bonds) present as coupling groups in the capping reagent with corresponding coupling moieties in the capping reagent, thereby covalently attaching the capping reagent to the capping reagent layer via free radical coupling.

The capping reagent and the capping reagent may each have a functional group that reacts with each other, thereby performing polymerization. In one example, the capping reagent is a water-soluble molecule containing at least 2 coupling moieties, each of the capping reagents having at least one copolymerizable functional group that can react with the coupling group in the capping reagent. In one example, the coating agent may be a water soluble molecule having the formula:

wherein,

t is a moiety for adjusting solubility,

rc is a moiety containing at least 3 backbone carbon atoms,

g is selected from the group consisting of N or C,

z' is a copolymerizable group,

n is an integer of 1 or 2, and

n 'is 0 or 1, where n' is selected to satisfy the valence requirement of G.

A water-soluble shell having the desired properties may be obtained with capping reagents in which the Rc moiety has less than 30, preferably less than 20, or more preferably less than 12 backbone carbon atoms. In a preferred embodiment, Rc contains 3 to 12 backbone carbon atoms. This range provides high coupling efficiency in the synthesis of nanocrystals under specific experimental conditions. The T moiety may be a polar/hydrophilic functional group for adjusting the solubility of the nanocrystal in the environment in which the nanocrystal is located. Accordingly, the group T may impart hydrophilic or hydrophobic properties to the shell, thereby rendering the nanocrystal soluble in aqueous as well as non-aqueous environments. T may be selected from polar groups, for example: hydroxyl, carboxyl, carbonyl, sulfonate, phosphate, amino, carboxamide. To obtain nanocrystals that are insoluble in an aqueous environment, the group T may also be hydrophobic, for example: various aliphatic or aromatic hydrocarbons (e.g., derivatives of fatty acids or benzene), or various other organic groups that are insoluble in water. When T is hydrophobic, it may also be modified by introducing hydrophilic groups after the coating agent has been copolymerized with the capping agent. The group Z' is a copolymerizable group having a functional group that can be copolymerized with the coupling moiety on the capping reagent. Suitable functional groups include, but are not limited to, for example: -NH₂-COOH or-OH, -Br, -C ═ C-. Z' may additionally contain aliphatic or cyclic carbon chains, preferably having at least 2 main chain carbon atoms.

In one example, T may be derived from a cyclodextrin molecule. The cyclodextrin molecules have a large number of hydroxyl groups which increase the water solubility of the resulting copolymer and can also be readily bound to biomolecules for use in biomarkers. Examples of suitable cyclodextrins include: alpha-cyclodextrin, beta-cyclodextrin, gamma-cyclodextrin, dimethyl-alpha-cyclodextrin, trimethyl-alpha-cyclodextrin, dimethyl-beta-cyclodextrin, trimethyl-beta-cyclodextrin, dimethyl-gamma-cyclodextrin and trimethyl-gamma-cyclodextrin.

In another example, the coating reagent is a water soluble molecule selected from an amino acid, preferably selected from a diamino acid (diamino acid) or a dicarboxylic amino acid (dicarboxylic amino acid). Specific examples of presently contemplated diamino acids include: 2, 4-diaminobutyric acid, 2, 3-diaminopropionic acid or 2, 5-diaminopentanoic acid, to name but a few. Dicarboxylic acids contemplated by the present invention include, but are not limited to, aspartic acid and glutamic acid.

In other examples, the coating agent is a water soluble molecule selected from the group consisting of:

wherein CD is cyclodextrin, and

in another example where the capping reagent contains an unsaturated group (e.g., a C ═ C double bond), suitable capping reagents for coupling include dienes and trienes, such as: 1, 4-butadiene, 1, 5-pentadiene and 1, 6-hexadiene.

A method for preparing one of the above water-soluble nanocrystals (item 2) is described, for example, in WO 2006/118543 and WO 2006/118542. Examples 4 and 5 describe some examples of the preparation of such water-soluble nanoparticles.

PCT/SG2008/000356 describes another option for obtaining water-soluble nanocrystals. The nanocrystals described in this application comprise amphiphilic polymers that are placed on the surface of the nanocrystal by non-covalent or covalent interactions.

The amphiphilic polymer may have the following general formula (V):

in the formula, m, o and p are each independently selected from integers of 0 to about 400, including: an integer from 1 to about 400, or from about 2 to about 400, for example: about 0 to about 400, about 0 to about 350, about 0 to about 300, about 3 to about 300, about 0 to about 250, about 0 to about 200, about 2 to about 200, about 0 to about 150, about 2 to about 150, about 0 to about 200, about 1 to about 200, about 3 to about 100, about 2 to about 100, about 0 to about 100, about 3 to about 50, about 2 to about 50, about 1 to about 50, or about 0 to about 50. By way of further illustration, in some examples, m may be selected from about 5 to about 50, for example: from about 10 to about 45, including from about 10 to about 43, and p may be selected, for example, from about 3 to about 40, such as: from about 3 to about 35 or from about 4 to about 30, and p may for example be selected from 0 to about 30, for example: 0 to about 25 or 0 to about 20. The sum of m + o + p is selected from about 10 to about 10000, including: about 10 to about 8000, about 10 to about 6000, about 10 to about 5000, about 10 to about 4000, about 10 to about 2000, about 10 to about 1000, about 10 to about 750, about 10 to about 600, about 10 to about 400, about 10 to about 250, about 10 to about 150, about 10 to about 100, about 15 to about 150, about 20 to about 150, about 15 to about 100, or about 20 to about 100. In some examples, m, o, and p are each independently selected from integers of about 2 to about 300, including: an integer of about 3 to about 300, about 3 to about 250, about 3 to about 200, about 3 to about 150, or about 2 to about 200, about 3 to about 100, about 2 to about 100, about 3 to about 80, about 2 to about 80, about 3 to about 40, or about 2 to about 40, and the sum of (m + o + p) is selected from about 6 to about 400, comprising: about 10 to about 400, about 10 to about 350, about 10 to about 300, about 10 to about 250, about 10 to about 200, about 6 to about 200, about 10 to about 150, about 6 to about 150, about 10 to about 100, about 6 to about 100, about 10 to about 50, or about 6 to about 50. In one embodiment, the sum of (m + o + p) is 32. In another example, the sum of (m + o + p) is 48. The ratio of p/(m + o) may be selected from 0 to about 25, for example: 0 to about 20, 0 to about 15, 0 to about 12, 0 to about 10, 0 to about 8, 0 to about 6, to about 4, to about 3, or to about 2. In one example, the ratio of p/(m + o) is about 1.

R²Is a first aliphatic group as described above having from about 3 to about 20 carbon atoms and from 0 to about 3 heteroatoms in the backbone. The heteroatom is selected from NO, S, Se and Si. R³Is a second aliphatic group as described above having from about 3 to about 80 carbon atoms and from 0 to about 40 heteroatoms in the backbone. The heteroatom is selected from N and O. The second aliphatic group R³Having a copolymerizable group. It will be appreciated that the individual units (individual units) shown in the above formula may be arranged in various orders (including randomly) in addition to being arranged in blocks. Thus, the above formula defines only the presence of m units in the polymer:

an o unit:

and p unit:

thus, the above units comprised by the polymer may be of various sequences. As an illustrative example, an illustrative example of various sequences may include the following units in the order:

amphiphilic polymers can be prepared by the method described in PCT/SG 2008/000356. The amphiphilic polymer is generally at least substantially free of cross-linking. Accordingly, in the amphiphilic polymer, the second aliphatic group R³The copolymerizable groups (before) of (a) can be used for various crosslinking or copolymerization reactions.

For example, in a method of preparing such nanocrystals coated with the amphiphilic polymer, maleic anhydride polymer is used as a reactant to form a hydrophilic backbone on the amphiphilic polymer. The maleic anhydride polymer may be a commercially available poly (isobutylene-alt-maleic anhydride) alternating copolymer having Chemical Abstract registry number 26426-80-2Also known as isobutylene-maleic anhydride copolymers. Other commercial names available are BM 30AE20, Fibersorb^TMSA 7200H, IB 6, KI Gel and

it is also available, for example, from Sigma-Aldrich (St. Louis, MO, USA) or SinoChemex company (Shanghai, PRC). The maleic anhydride polymer may also be a poly (ethylene-alt-maleic anhydride) alternating copolymer with chemical abstractact registry number 106973-21-1, also known as an ethylene-maleic anhydride alternating copolymer. The maleic anhydride polymer is available, for example, from Rutherford Chemicals (Bayonne, NJ) under the trade designation 27109P and under the trade designation

Or

And E60. In formula (V) above, n may be any integer from about 10 to about 10000, for example: about 10 to about 5000, about 10 to about 2000, about 10 to about 1000, about 20 to about 1000, about 10 to about 800, about 20 to about 800, such as about 10 to about 400. In one example, n is 32. The above examples of maleic anhydride polymers should not be understood as limiting, but any maleic anhydride polymer available (also including the maleic anhydride polymers which need to be synthesized), in particular maleic anhydride polymers which can be prepared according to conventional procedures known in the art, are suitable. For example, various maleic anhydride polymers were prepared according to the procedures described in U.S. Pat. Nos. 3846383 and 6316554. The conventional procedures used in the art and involving, for example, the free radical copolymerization of maleic anhydride with olefins in the presence of peroxides are reviewed in the abstract of Frank, H.P., Makromolekulare Chemie (1968)114, 113-121.

Monofunctional compounds and at least difunctional compounds are used as further reactants. During the treatment, said monofunctional compound is converted into the first aliphatic radical R mentioned above². The functional group of the monofunctional compound is capable of forming a bond with an anhydride. The at least bifunctional compound being converted intoThe second aliphatic group R³(before). The at least bifunctional compound may have two, three, four or more functional groups. One of the functional groups is capable of forming a bond with an anhydride. The other functional groups of the at least bifunctional compound are copolymerizable. The reaction of a portion of the anhydride rings with the monofunctional compound results in the formation of hydrophobic side chains that can interact with the hydrophobic surface of the nanoparticles described herein. The other part of the anhydride ring is used to attach the at least bifunctional compound to the backbone. By varying the amount of nucleophile used to react with the polyanhydride chains, the relative number of hydrophobic units and carboxyl groups on the polymer backbone can be controlled. Thus, the monofunctional compounds of the present invention can react with at least about 25%, such as at least about 35%, at least about 50%, at least about 70%, or at least about 75% of the available anhydride rings. The at least difunctional compound of the present invention may be reacted with at least about 5%, such as at least about 8%, at least about 10%, at least about 15%, at least about 20%, or at least about 25% of the available anhydride rings.

The functional group of the monofunctional compound may be selected from, but is not limited to: amino groups, hydroxyl groups, thiol groups, selenohydroxy groups (selenol groups), halogen groups, ether groups, thioether groups, and the like. The alkyl chain of the monofunctional compound has from about 2 to about 20 carbon atoms, for example: about 3 to about 20, including about 5 to about 20, about 5 to about 15, about 7 to about 20, such as 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20. Further, the monofunctional compound has from 0 to about 2 heteroatoms, such as: 1 heteroatom. The heteroatom may be N, O, S, Se or Si, for example. Examples of monofunctional compounds may be, but are not limited to: alkyl amines, wherein alkyl is as defined above. In one embodiment, the alkylamine can be n-propylamine, n-butylamine, n-pentylamine, n-hexylamine, n-octylamine, or n-dodecylamine.

Typically, the at least bifunctional group has two or more different functional groups. Any functional group may be selected as any of the at least difunctional groups so long as one (typically only one) of the at least difunctional groups can form a bond with the anhydride. The first group that reacts with the maleic anhydride group may be selected from, but is not limited to: amino groups, hydroxyl groups, thiol groups, selenohydroxy groups, halogen groups, ether groups, thioether groups, and the like. The second functional group that is not reactive with the maleic anhydride group may be a copolymerizable group. Copolymerizable means that the group can be polymerized with other functional groups. Examples of suitable copolymerizable groups include, but are not limited to: amino, hydroxyl, allyl glycidyl ether, epoxy, oxetane, and C ═ C bonds (e.g., allyl).

The at least bifunctional compound has an alkyl chain of from about 3 to about 80 carbon atoms, comprising: about 3 to about 70 carbon atoms, about 3 to about 60 carbon atoms, about 3 to about 40 carbon atoms, about 10 to about 80 carbon atoms, about 10 to about 60 carbon atoms, about 25 to about 60 carbon atoms, about 10 to about 40 carbon atoms, about 3 to about 20 carbon atoms, or about 3 to about 10 carbon atoms, for example 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, or 44 carbon atoms. Further, the at least bifunctional compound has from 0 to about 45 heteroatoms, including: 0 to about 40 heteroatoms, 1 to about 40 heteroatoms, about 2 to about 30 heteroatoms, or about 0 to about 3 heteroatoms, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, or 43 heteroatoms, e.g., N or O.

In the formation of the above amphiphilic polymer, the reaction between the maleic anhydride polymer of formula (V) (previously described), the monofunctional compound and the at least bifunctional compound may be carried out in the presence of a base. In general, any base suitable for the intended purpose may be used. In one embodiment, the base is a nucleophilic base. Nucleophilic bases are bases having basic as well as nucleophilic properties. Illustrative examples include, but are not limited to: lithium diisopropylamide, lithium tetramethylpiperidine, diisopropylethylamine (Hunig's base), 1, 5-diazabicyclo [4.3.0] -non-5-ene, 1, 8-diazabicyclo [5.4.0] -undec-7-ene, bis (trimethylsilyl) amide, hexamethyldisilazane (hexamethyldisilazane) or magnesium diisopropylideneacetone (bismesityl magnesium).

In forming the amphiphilic polymer, the reaction of the anhydride groups of the maleic anhydride polymer typically forms amide or ester groups, providing non-polar side chains. The side chain includes a moiety formed by the monofunctional compound, such as: an alkyl chain and at least one heteroatom which is a separate atom from the ester group or the amide group to which the moiety formed by the monofunctional compound is attached. The first aliphatic moiety thus comprises the moiety formed by the monofunctional compound and the group COO-, CO-NH-or CO-N-. In a specific embodiment, when the group is CO-N-, a secondary amide is formed, and thus the first aliphatic moiety comprises 2 moieties formed from the monofunctional compound.

In this respect it is to be noted that in the case of the formation of a secondary amide, the first aliphatic moiety may also contain one moiety formed by the monofunctional compound and one moiety formed by the at least bifunctional compound. For the sake of clarity, any aliphatic moiety including moieties formed by at least said at least bifunctional compound is to be understood as second aliphatic moiety. Correspondingly, an aliphatic moiety comprising both a moiety formed by the monofunctional compound and a moiety formed by the at least bifunctional compound is to be understood as a branched second aliphatic moiety.

Further, each anhydride group of the maleic anhydride polymer may be reacted with one or two monofunctional compounds or with one or two at least difunctional compounds (see below). In general, the anhydride groups, whether monofunctional or at least difunctional, are reacted with the largest number of one reagent. As a result, carboxylic acid groups are formed, which usually provide a negative charge in solution. In addition, many anhydride groups are not reacted. These anhydride groups are typically hydrolyzed during the treatment of the present invention, thus further providing carboxylic acid groups. It will be appreciated by those skilled in the art that the method of forming an amphiphilic polymer of the present invention does not require the use of any coupling agent, such as a cross-linking agent.

It has been previously indicated that poly (isobutylene-alt-maleic anhydride) alternating copolymers can be reacted with aliphatic amines (abstract of japanese patent application JP 57016004, Fernandez-argueles, m.t., et al, NanoLetters (2008)7, 9, 2613-2617), and in addition it has been found that di-, tri-and higher functionalized compounds can be used as additional reactants in a one-pot synthesis. The reaction conditions can generally be controlled in such a way that the di-or higher functionalized compounds participate in only one reaction, whereby only one bond is formed on the hydrocarbon backbone of the polymer. The remaining functional groups of the di-or higher functionalized compound can then be used for coupling and crosslinking reactions. In this regard, it should be noted that the hydrocarbon backbone of the amphiphilic polymer may bear polar side groups and side chains having alkyl chains with from about 3 to about 80 carbon atoms and from 0 to about 40 heteroatoms selected from N and O, the side chains having copolymerizable groups. As mentioned above, the copolymerizable group may be an amino group, a hydroxyl group or a group containing a terminal or internal C ═ C or C ≡ C bond (e.g.: terminal-CH ═ CH)₂A radical, a terminal-C ≡ CH radical, an endo-CH ≡ CH-radical or an endo-C ≡ C-radical). As mentioned above, the copolymerizable groups may also include terminal or internal C ═ C or C ≡ C bonds and other functional groups or vicinal (vicinal) or geminal heteroatoms, such as: the general structure-G-CH ═ CH-or G-C ≡ C-, where G is N, O or a-CH ≡ CH-group or a-C ≡ C-group.

The polymerizable group may be the same as or different from the remaining groups of the at least bifunctional compound. In the case where the functional groups are the same, those functional groups that should not react with the maleic anhydride polymer may be covered to prevent these functional groups from participating in the polymerization process. A wide variety of protecting groups well known to those skilled in the art can be used for the various functional groups. As an illustrative example, the hydroxyl group may be protected by an isopropylidene group. Such protecting groups can be removed after polymerization, and the functional groups, which are no longer protected, can thus be used for coupling, crosslinking or copolymerization reactions. For example, the isopropylidene protecting group protecting the hydroxyl group can be removed by acid treatment. It will further be appreciated by those skilled in the art that the protecting groups may be introduced beforehand during the synthesis of the individual at least bifunctional compounds.

PCT/SG2008/000356 describes examples of preparing the amphiphilic polymer and nanocrystals coated with the amphiphilic polymer. Examples are also listed in example 6. Other amphiphilic nanocrystals useful in the present invention are described, for example, in articles by Fernandez-Arg oils, m.t., Yakovlev, a. et al (2007, Nano lett., vol.7, No. 9, p.2613) and by Cheng-An, j.l., Ralph, a. et al (2008, small, vol.4, p.3, p.334). The amphiphilic nanocrystals described by Cheng-An, j.l., Ralph, a. et al are based on a poly (maleic anhydride) backbone. The reaction of a portion of the anhydride ring with the alkyl amine results in the formation of hydrophobic side chains for insertion into the hydrophobic surfactant layer on the surface of the nanoparticle. The other part of the anhydride ring is used to attach a functional organic molecule to the backbone. The amphiphilic nanocrystals described by Fernandez-Arg lues, m.t., Yakovlev, a. et al contain amphiphilic polymers synthesized by linking an amino-functionalized hydrophobic hydrocarbon chain (dodecylamine) to a polar backbone (poly (isobutylene-alt-maleic anhydride) alternating copolymer) under reaction conditions of the anhydride rings of the polymer backbone.

In addition, Dubertret et al (Science (2002)298, 1759) also describe water-soluble nanocrystals. Dubertret and his colleagues have encapsulated nanocrystals in the hydrophobic core of micelles, which is a mixture of 40% 1, 2-dipalmitoyl-sn-glycero-3-phosphoethanolamine-N- [ methoxy (polyethylene glycol) -2000] and 60% 1, 2-dipalmitoyl-sn-glycero-3-phosphocholine. The micelles provide a hydrophobic interface for the nanoparticles and maintain high colloidal stability. The presence of a PEG layer is a prerequisite for solubilization of the nanocrystals.

Other authors reported the feasibility of coating nanocrystals directly with amphiphilic polymers, independent of the formation of polymeric micelles in a process such as Dubertret and his colleagues. Amphiphilic, alkyl-modified (octylamine or isopropylamine) low molecular weight polyacrylic acids were successfully used to coat TOPO-protected nanocrystals and to dissolve the nanocrystals in water (Wu, x. et al, nat. biotechnol. (2003)21, 41; matthakis, l.c. et al, anal.biochem. (2004)327, 200; luccaradin c. et al, Langmuir (2006)22, 2304). In one study, luccarardini et al (supra) reported pH-dependent interactions of nanocrystals coated with lipid membranes. They found that: in biological buffers, the interaction between the polymer-coated nanocrystals and the membrane can be controlled by the pH of the buffer. In the work of Gao et al, a high molecular weight amphiphilic ABC triblock copolymer composed of a polybutylacrylate moiety (hydrophobic), an polyethylacrylate moiety (hydrophobic), and a polymethacrylic acid moiety (hydrophilic) was used to directly encapsulate Quantum Dots (QDs) (Gao x et al, nat. biotechnol. (2004)22, 969).

Another class of polymers used to directly transfer hydrophobic QDs into water are poly (maleic anhydride-alt-1-tetradecene) alternating copolymers (Pellegrino, T et al, Nano Lett. (2004)4, 703; Yu, w.w., et al, j.am. chem. soc. (2007)129, 2871). The stability of the polymer shell is increased by the addition of a bis (6-aminohexyl) amine crosslinker.

In addition to the nanocrystals, the plants of the invention may also contain organic dyes, such as: a fluorophore. Examples of fluorophores include, but are not limited to: hydroxycoumarins (blue), such as 7-hydroxycoumarin-3-carboxylic acid; methoxycoumarin (blue), such as 7-methoxycoumarin-4-acetic acid; cyanine dyes, such as cyanine (Cy2, dark green), indocyanine (indocyanine) (Cy3), Indodicarbocyanine (indocyanine) (Cy 5); aminocoumarins (blue), such as 7-aminocoumarin; FAM (dark green), such as 5-carboxyfluorescein (5-FAM); alexa Fluor, e.g., Alexa Fluor 350, 405,430. 488, 500, 514, 532, 546, 555, 568, 594, 610, 633, 647, 660, 680, 700 and 750; fluorescein (FITC; CAS registry number: 2321-07-5) (light green); HEX fluorophore (pale green), e.g., 6-carboxy-2 ', 4, 4', 5 ', 7, 7' -hexachlorofluorescein; TRITC (yellow), for example: tetramethylrhodamine6-isothiocyanate chloride (6-TRITC) 6; R-Phycoerythrin (PE) (yellow); texas Red (Texas Red) (C)₃₁H₂₉S₂N₂O₆Cl 1; CAS No.: 82354-19-6) (orange); carboxytetramethylrhodamine (TAMARA) (red); 6-carboxy-X-rhodamine (6-Rox) (Red); or allophycocyanin (A-PC) (red), to name just a few. In one example, fluorescein is used as the fluorophore, which can be mixed with the nanocrystal (see fig. 4).

Organic dyes may also include, but are not limited to, chromophores such as: azo compounds, lycopene, beta-carotene, anthocyanin, fluorescein and fluorescein.

Such mixtures of nanocrystals and organic dyes can produce plants of different colors that contain a mixture of at least one nanocrystal and at least one organic dye.

In another aspect, the present invention relates to a method of dyeing a plant with at least one nanocrystal, the method comprising:

providing a solution comprising at least one nanocrystal dissolved in the solution;

contacting said plant with said solution.

In the above method of the present invention, the nanocrystals are first provided in a suitable solvent or mixture of solvents, i.e. the solution is a mixture of a solvent and at least one nanocrystal, wherein the nanocrystals are dissolved in the solvent.

Suitable in this respect means that the nanocrystals should be soluble in the respective solvents. Examples of such solvents are, but are not limited to, aprotic solvents and/or apolar solvents, such as: an aprotic non-polar solvent. The non-polar solvent may include, but is not limited to: mineral oil, hexane, heptane, cyclohexane, benzene, toluene, pyridine, dichloromethane, chloroform, carbon tetrachloride, carbon disulfide, dioxane, diethyl ether, 1-octadecene, 9-octadecene, diisopropyl ether, ethylene glycol monobutyl ether, tetrahydrofuran, or mixtures thereof. In one example, 1-octadecene is used to dissolve the nanocrystals.

In another example, a protic solvent is used. Protic solvents are solvents having, for example, hydrogen atoms attached to oxygen atoms (as in hydroxyl groups) or to nitrogen atoms (as in amino groups). More generally, any compound containing dissociable H⁺The molecular solvent (e.g., hydrogen fluoride) of (a) is referred to as a protic solvent. Molecules of such solvents can provide H⁺(proton). Examples of polar protic solvents include, but are not limited to: water, methanol, ethanol, butanol, tert-butanol, phenol, cyclohexanol, formic acid, acetic acid, and dimethylarsinic acid [ (CH)₃)₂AsO(OH)]Aniline, N-dimethylformamide, N-diisopropylethylamine or chlorophenol. In particular, when water-soluble nanocrystals are used, protic solvents, such as water, may be used.

Other examples of suitable non-polar solvents are non-polar ionic liquids. Examples of non-polar ionic liquids include, but are not limited to: 1-Ethyl-3-methylimidazolium bis [ (trifluoromethyl) sulfonyl ] amide bis (trifluoromethanesulfonyl) amide, 1-ethyl-3-methylimidazolium bis [ (trifluoromethyl) sulfonyl ] amide trifluoroacetate, 1-butyl-3-methylimidazolium hexafluorophosphate, 1-hexyl-3-methylimidazolium bis (trifluoromethylsulfonyl) diimide, 1-butyl-3-methylimidazolium bis (trifluoromethylsulfonyl) imide, trihexyl (tetradecyl) phosphonium bis [ oxalate (2-) ] borate, 1-hexyl-3-methylimidazolium tris (pentafluoroethyl) trifluorophosphate, 1-butyl-3-methyl-imidazolium hexafluorophosphate, tris (pentafluoroethyl) trifluorophosphate, tris (pentafluoroethyl) phosphonium salt, bis (trifluoromethanesulfonyl) imide, bis (trifluoromethyl) amide, 1-ethyl-3-methylimidazolium bis (trifluoromethanesulfonyl) imide, 1-butyl-3-methylimidazoliu, Trihexyl- (tetradecyl) phosphonium, N "-ethyl-N, N' -tetramethylguanidinium salt, 1-butyl-1-methylpyrrolidinium tris (pentafluoroethyl) trifluorophosphate, 1-butyl-1-methylpyrrolidinium bis (trifluoromethylsulfonyl) imide, 1-butyl-3-methylimidazolium hexafluorophosphate, 1-ethyl-3-methylimidazolium bis (trifluoromethylsulfonyl) imide and 1-N-butyl-3-methylimidazolium. Examples of polar aprotic solvents include, but are not limited to: butanone, methyl isobutyl ketone, acetone, cyclohexanone, ethyl acetate, isobutyl isobutyrate, ethylene glycol diacetate, dimethylformamide, acetonitrile, N-dimethylacetamide, nitromethane, acetonitrile, N-methylpyrrolidone, and dimethyl sulfoxide.

In one example, the solvent is a high boiling point solvent. Examples of such solvents include 1-octadecene or 9-octadecene. Their boiling point is higher than 250 ℃. Other examples include, but are not limited to: dow Heat Carrier A (Dowtherm A), which is two very stable compounds (Biphenyl (C)₁₂H₁₀) And diphenyl ether (C)₁₂H₁₀O)) a eutectic mixture having a boiling point of 275.1 ℃; or of the formula C₁₂H_10-xCl_xFor example Therminol 66 with a boiling point of 359 ℃. In another embodiment, low boiling solvents such as hexane, cyclohexane, chloroform, and the like may be used.

The water-soluble nanocrystals can be easily mixed or dissolved in an aqueous solution. The aqueous solution is suitable for uptake by plants via roots or cut stems. When using water-insoluble (i.e., hydrophobic) nanocrystals, it is desirable to dissolve the nanocrystals in a solvent suitable for uptake by plants through roots or crop stalks, such as: a hydrophobic solvent or an amphoteric solvent. In addition to the octadecene used in one of the examples described herein (see example 1), other hydrophobic solvents may also be used. Examples include vegetable oils, such as: soybean oil (soyoil), rapeseed oil (rape oil), sunflower oil, olive oil, to name a few. The hydrophobic nanocrystals may be dissolved in such oils before being absorbed by the plant through the roots or cut stems.

Hydrophobic nanoparticles may also be dispersed in non-hydrophobic solvents. For example, water-oil emulsions may be prepared by dispersing hydrophobic nanocrystals in a polar or aqueous solvent as described above.

In another aspect of the invention, the roots of the plant are immersed in the solution. On the other hand, the stem of the plant is cut and the plant having the cut stem is placed in the solution. In this case, since the plant cell wall and epidermis material is composed of a large number of long chain fatty acids, the emissive nanocrystals are dispersed in long chain hydrocarbons, such as: 1-octadecene or 9-octadecene, to name only two examples.

In one example, an orchid named dendrobium candidum is used to absorb nanocrystals dissolved in octadecene. When transport channels specific for organic media are absent in plants, the nanocrystals diffuse into the flower at a very slow rate. Fig. 2 shows an example of staining a plant by immersing a cut stem of the plant in an organic solvent containing nanocrystals. In another example, a mixture of nanocrystals and fluorophores is dissolved in an organic dye. Then, the cut stem of the plant is immersed in the solution, and the plant shows various colors after being absorbed by the plant (see, for example, fig. 4 a).

In another aspect of the invention, aerial parts of the plant or the whole plant are contacted with a solution comprising at least one nanocrystal or mixture of nanocrystals and organic dye dissolved in a suitable solvent.

The plants are contacted with the solution by dipping or spraying the solution onto aerial parts of the plants or the whole plant. FIG. 1 shows an example of an orchid named Dendrobium candidum which is dip-coated with the solution. The nanocrystals or mixture of nanocrystals and organic dyes are absorbed by the surface of the plant and part of the plant cells of the epidermis/cuticle.

The concentration of the at least one nanocrystal is from about 0.001 to about 1mmol/L, alternatively from about 0.01 to about 1mmol/L, and alternatively from about 0.1 to about 0.5 mmol/L. Organic dyes may be used in concentrations ranging from 0.001 to 1.0mol/L, alternatively from about 0.01 to 1mmol/L, alternatively from about 0.1 to 0.5 mmol/L.

When exposed to light of a particular wavelength, the plants of the present invention may exhibit a color imparted by ingestion or contact with at least one nanocrystal. This can occur in the visible spectrum as well as under UV light. Most nanocrystals emit a specific color when exposed to UV light.

The invention illustratively described herein suitably may be practiced in the absence of any element or elements, limitation or limitations which is not specifically disclosed herein. Thus, for example, the terms "comprising," "including," "containing," and the like are to be construed as open-ended as opposed to closed-ended. Furthermore, the terms and expressions which have been employed herein are used as terms of description and not of limitation, and there is no intention, in the use of such terms and expressions, of excluding any equivalents of the features shown and described or portions thereof, but it is recognized that various modifications are possible within the scope of the invention claimed. Thus, it should be understood that although the present invention has been specifically described by preferred embodiments and optional features, modification and variation of the inventions specifically disclosed herein may be resorted to by those skilled in the art, and that such modifications and variations are within the scope of this invention.

The present invention has been described broadly and generically herein. Every narrow genus and subgeneric group falling within the general disclosure above is also part of the invention. This includes the generic description of the invention with a proviso or negative limitation excluding any subject matter from the genus, regardless of whether or not the excised material is specifically recited herein.

Other embodiments are within the following claims and non-limiting examples. In addition, while features or aspects of the invention are described in terms of Markush groups, it will be understood by those skilled in the art that the invention may also be described in terms of any individual member or subclass of members of the Markush group.

Examples

1.1 surface adsorption/uptake of absorbing emissive nanocrystals

There are various methods for modifying the color of flowers (flora) and leaves (leaf). Compared with gene modification, the physical method is more attractive due to simple technology. Surface adsorption/uptake of the emissive nanocrystals was performed by dip-coating orchid (e.g. dendrobium candidum) in a solution of CdZnSe nanocrystals, which showed highly emissive flowers excited at 302/345nm under UV excitation (fig. 1).

Radical uptake of emissive nanocrystals in octadecene

Since the cell wall and epidermal material of plants is composed primarily of long chain fatty acids, we designed emissive nanocrystals that are dispersed in long chain hydrocarbons (e.g., octadecene). Dendrobium candidum then ingests the same CdZnSe nanocrystals (quantum dots) dissolved in octadecene. In the absence of a dedicated transport channel for organic media, quantum dots diffuse into the flower at a very slow rate. A minimum of 1 hour of ingestion was required to have visible luminescence under UV excitation and 3 hours was required for adequate ingestion. The uptake of nanocrystals along the periphery of the sepals (periphery) was observed at a higher rate compared to pulse-like (vein-like) fluorescent yellow diffusion. Diffusion then begins from the center in a weakly (less) oriented manner towards the edges of the petals and sepals. This can be explained by the high chemical compatibility of octadecene with the plant's cuticle (cuticle) and cell wall. Ingestion for 15 hours resulted in the development of color throughout the flower (FIG. 2, lower panel).

In general, it can also be said that the coloration of plants can be controlled by varying the uptake time of the coloring agent (for example: different types of nanocrystals, or other organic dyes, or mixtures of nanocrystals and organic dyes). In one example, a mixture of CdZnSe nanocrystals and fluorescein may be used. A short intake of the solution containing the two stains allowed the veins to be stained by fluorescein while the ovary, basal and peripheral regions of the flowers and sepals were stained by CdZnSe nanocrystals.

Uptake of emissive quantum dots and dyes in sequence

The sequential uptake of CdZnSe nanocrystals dissolved in octadecene and an aqueous solution of a fluorescent yellow dye is explained next. And the nanocrystalline and the organic dye are taken in sequence to obtain multicolor decoration. The nanocrystals were ingested through the cut stem for 4 hours, followed by 15 minutes of ingestion of the fluorescent yellow into the flower (also through the cut stem). The result is a combined emission of the ovary region as fluorescent yellow-nanocrystals that expands into distinct emissions as opposed to petals and sepals, (fig. 4a) despite minimal changes in the bright field morphology of the flower.

The cross section of the stem corresponding to the above uptake was observed under a confocal microscope (fig. 4 b). In addition to standard confocal imaging, confocal spectroscopic imaging (spectral confocal image) was used to confirm the presence of CdZnSe nanocrystals (quantum dots) and fluorescein yellow. By choosing the appropriate wavelength, sampling increment, and minimizing auto-fluorescence, it is ensured that there is no crosstalk information and the confocal image is spectrally pure.

The specific uptake of dyes and nanocrystals on the cell wall was further investigated on a longitudinal section of the stem. Consistently, clear water-based regions were observed along the vascular bundle while hydrophobic regions were observed around the vascular bundle. Spectral analysis of different regions of the cross-section compared the emission data of fluorescein and nanocrystal. Based on the emission behavior of the above-mentioned dyes and nanocrystals, the swollen fragments were digitally reconstructed into corresponding hydrophobic and hydrophilic cell signals using confocal software.

Dye and nanocrystal uptake was performed using commercially available white dendrobium. The stems were cut open and immersed in a solution of 0.001-1.0mmol/L (or 0.03mol/L in this particular example) of CdZnSe nanocrystals in 1-octadecene, or 0.001-1.0mmol/L (or 0.03mol/L in this particular example) of aqueous fluorescein sodium (Fluka 46960). If it is desired that the entire plant has a visible color, the stem is immersed in a solution containing CdZnSe nanocrystals for about 4 hours and then in a solution containing fluorescein for 15 minutes. The stems were then washed with octadecene or water, respectively, before being wiped dry with KimWipes toilet paper. Images under UV were then taken with a digital camera Sony Cybershot DSC-T1.

The stem section was carefully sectioned with a scalpel for confocal imaging. Each stem section was then carefully transferred to a microscope glass coverslip. A sample placed against the objective lens was imaged using a nikon Clsi laser scanning confocal microscope equipped with unmixed spectroscopic imaging and fluorescent probes.

CdZnSe nanocrystals were prepared by the following method

The liquid oleic acid and the liquid octadecene are used to prepare luminescent or magnetic nanocrystals to prepare nanocrystals. As an example, the following steps are given for the preparation of luminescent nanocrystals: 1.0mmol of cadmium oxide, 0.5mmol of zinc oxide, 3.1mmol of selenium are added to a flask containing 3.1mmol of oleic acid and 10mL of octadecene, which is then degassed with stirring and heated to 300 ℃ until a clear solution is formed. The reaction was continued for 30 minutes and the reaction was allowed to cool to room temperature. The obtained nano-crystal can be directly used for the uptake of flowers.

Soybean oil may also be used instead of oleic acid. Nanocrystals can be formed in these media as described in the previous paragraph or in example 1 above. It is also possible to disperse the prepared nanocrystals into a solution after purifying the nanocrystals prepared by other methods.

2. Synthesis of binary metal oxides in homogeneous mixtures using low-boiling, nonpolar solvents

2.1 Synthesis of binary Metal oxide ZnO

3.0mmol of ZnO was dissolved in 7.5mmol of oleic acid at 260 ℃ to form a clear solution. After cooling to room temperature, 18mL of hexane and 6mmol of oleylamine were added, and the solution was transferred to 100mL of Parr reactor 4950 and reacted with N₂And (5) blowing the gas. The mixture was rapidly heated to 320 ℃ with stirring and held at this temperature for 30 minutes. Then, by simply removing the heatAnd cooled to stop the reaction. The final product is purified by a simple centrifugation and dispersion step, i.e., the precipitated nanocrystals can be collected and dried or redissolved in an organic solvent (e.g., hexane) for storage.

2.2 Synthesis of ternary Quantum dots ZnCdSe

0.3mmol of ZnO and 0.3mmol of CdO were dissolved in 2.4mmol of oleic acid at 320 ℃ to form a clear, homogeneous solution. After cooling the solution to 60 ℃, 2.4mL of Se solution (1M of TOP-Se solution), 5g of Hexadecylamine (HDA) and 2g of trioctylphosphine oxide (TOPO) and 20mL of hexane were added; the solution was then transferred to 100mL Parr reactor 4950 and treated with N₂And (5) blowing the gas. The mixture was rapidly heated to 320 ℃ with stirring and held at this temperature for 30 minutes to 3 hours. The reaction was then stopped by removing the heat and cooling. The resulting nanocrystals are purified by a simple centrifugation/dispersion step, i.e., the precipitated nanocrystals can be collected and dried or the precipitated product redissolved in an organic solvent (e.g., hexane) for storage.

2.3 Synthesis of ternary Quantum dots MgFe₂O₄

0.4mmol magnesium carbonate and 0.4mmol ferrous acetate were dissolved in 3.0mmol oleic acid at 320 ℃ to form a clear homogeneous solution. After cooling the solution to 60 ℃, 5g oleylamine and 20mL of hexane were added; the solution was then transferred to 100mL Parr reactor 4950 and purged with N2 gas. The mixture was rapidly heated to 320 ℃ with stirring and held at this temperature for 30 minutes to 3 hours. The reaction was then stopped by removing the heat and cooling. After centrifugation, the precipitated product is collected and dried or redissolved in an organic solvent (e.g., hexane) for storage.

2.4 Synthesis of NiFe₂O₄Nanocrystal

1.0mmol of nickel acetate and 2.0mmol of iron (III) acetylacetonate (ironacetacetoacetate) were dissolved in 9.0mmol of oleic acid at 150 ℃ to form a homogeneous solution. Cooling the solutionAfter cooling to 60 ℃, 5mL of trioctylamine and 20mL of hexane were added; the solution was then transferred to 100mL Parr reactor 4950 and treated with N₂And (5) blowing the gas. The mixture was rapidly heated to 320 ℃ with stirring and held at this temperature for 30 minutes to 1 hour. The reaction was then stopped by removing the heat and cooling. Therein, 1.0mmol of nickel acetate and 2.0mmol of iron (III) acetylacetonate were dissolved in 9.0mmol of oleic acid and 5mL of trioctylamine and 5mL of ODE at 150 ℃ to form a homogeneous solution. Then, the mixture was rapidly heated to 320 ℃ under 1atm with stirring and held at that temperature for 30 minutes to 1 hour. The reaction was stopped by removing the heat and cooling. The resulting mixture is centrifuged and the obtained nanocrystals are collected and dried or re-dissolved in an organic solvent (e.g., hexane) for storage.

3.1 preparation of TOPO-covered (CdSe) -ZnS nanocrystals (Quantum dots, QD)

CdSe nanocrystals coated with Trioctylphosphine (TOP)/trioctylphosphine oxide (TOPO) were prepared as follows. TOPO (30g) was placed in a flask and dried under vacuum (. about.1 torr) at 180 ℃ for 1 hour. The flask was then charged with nitrogen and heated to 350 ℃. The following injections were prepared in an inert atmosphere glove box: CdMe₂(200mL), 1M TOPSe solution (4.0mL) and TOP (16 mL). The above injections were mixed well, filled into syringes, and removed from the glove box.

The reaction heat was removed and the reaction mixture was transferred to vigorously stirred TOPO using a single continuous injection. The reaction flask was reheated and the temperature was gradually increased to 260-280 ℃. After the reaction was complete, the reaction flask was cooled to-60 ℃ and 20mL butanol was added to prevent the TOPO from freezing. The addition of a large excess of methanol leads to particle flocculation. Separating the flocs from the supernatant by centrifugation; the resulting powder can be dispersed in a variety of organic solvents to produce a clear-looking solution.

The flask containing 5g of TOPO was heated to 190 ℃ for several hours under vacuum, then cooled to 60 ℃ before addition0.5mL Trioctylphosphine (TOP). Approximately 0.1 to 0.4. mu. mol of CdSe dots dispersed in hexane were transferred to the reaction vessel by syringe, and the solvent was withdrawn. Diethyl zinc (ZnEt)₂) And hexamethyldisilathiane ((TMS)₂S) are used as precursors for Zn and S, respectively. An equimolar amount of the precursor was dissolved in 2-4mL TOP in an inert atmosphere glove box. The precursor solution was loaded into a syringe and transferred to an addition funnel attached to the reaction flask. After the addition was complete, the mixture was cooled to 90 ℃ and stirred for several hours. Butanol was added to the mixture to prevent TOPO from freezing when cooled to room temperature.

3.2 preparation of Water-soluble nanocrystals by Forming host-guest complexes with Gamma-Cyclodextrin

The nanocrystal with hydrophobic surface layer using TOP/TOPO obtained in 3.1 was dissolved in 200. mu.L of chloroform/hexane (1: 1) mixture. About 0.5g of the gamma-cyclodextrin and nanocrystal solution was added to 20mL of deionized water. The mixture was refluxed for about 8 hours until a cloudy solution formed. Most of the water was removed by a rotary evaporator and the formed host-guest inclusion complex was separated by centrifugation. The collected solid was further washed with water to remove free cyclodextrin molecules. The thus obtained nanocrystals of the guest-host complex which had been formed by TOP/TOPO with cyclodextrin were stored in solid state. The nanocrystals were dissolved in water by sonication and could be easily transferred to water. It has been found that nanocrystals protected by host/guest complexes can be stable in solid form for a considerable period of time.

The water-soluble gamma-CD modified quantum dots formed by the formation of guest-host complexes can be followed optically. When gamma-cyclodextrin was added to a chloroform solution containing TOP/TOPO capped CdSe/ZnS core-shell nanocrystals, the nanocrystals formed were phase-transferred from organic chloroform into an aqueous solution. Can also use¹H-NMR, FT-IR spectroscopy and XRD testing (data not shown) confirmed the formation of gamma-CD modified quantum dots. Transmission Electron Microscopy (TEM) and fluorescence images show that quantum dots that have formed guest-host complexes with gamma-cyclodextrin form highly monodisperseAnd (3) granules. The CdSe/ZnS core-shell nanocrystals were shown to have a higher coverage (in CHCl) than the unmodified TOP/TOPO-capped core-shell nanocrystals after formation of the host-guest complex (measured in water)₃Medium) but no change in the maximum emission wavelength. Photoluminescence tests showed that CdSe/ZnS core-shell nanocrystals, which have formed a guest-host complex with γ -cyclodextrin, are very stable in PBS buffer at pH 7.4 (i.e., under physiological conditions), and show satisfactory stability even in aqueous solutions at pH 5.0 (triangles pointing upwards) and 3.0 (triangles pointing downwards), respectively. Finally, the CdSe/ZnS core-shell nanocrystals after forming guest-host complexes with γ -cyclodextrin showed good thermal stability in aqueous solution heated to 50 ℃.

4.1 preparation of Polymer-coupled Water-soluble nanocrystals in aqueous solution

Quantum dots coated with TOPO were prepared as described in 3.1. The TOPO coated quantum dots were then dissolved in chloroform together with a large amount of mercaptoethylamine (aminoethylthiol). The mixture was sonicated for 2 hours and then allowed to stand at room temperature until precipitation was complete. The resulting solid was washed several times with chloroform and collected by centrifugation. Then, the amino group-covered quantum dots were dissolved in a buffer solution having a pH of 8, followed by dropwise addition to a solution of a poly (acrylic acid) polymer (average molecular weight: 2000 based on GPC), EDC and thio-NHS were used as coupling agents to activate the coupling groups on the covering agent, and stirring was carried out at room temperature for 30 minutes.

The reaction mixture was first stirred at 0 ℃ for 4 hours and then reacted at room temperature overnight. The resulting solution was dialyzed overnight and stored after degassing with nitrogen. Further purification was performed by first washing the reaction solution twice with ether and centrifuging the above acidic (pH adjusted to about 4-5) solution coated with polymer nanocrystals. The collected nanocrystals were then redissolved in water by adjusting the pH (to 7-8).

The physico-chemical properties of the polymer shell nanocrystals of the invention are correlated with the mere capping with mercaptopropionic acid (MCA) or aminoethanethiol (aminoethanethiol)The physico-chemical properties of the (CdSe) -ZnS core-shell nanocrystals of hol) (AET) were compared as follows: addition of H to an aqueous solution of nanocrystals₂O₂And is caused to H₂O₂The final concentration of (2) reached 0.15mol/l and the chemical behavior was followed by photon spectroscopy (Photoscopial). For nanocrystals coated with MCA or AET only, oxidation of the nanocrystals was immediately detected and the nanocrystals precipitated out within 30 minutes. In contrast, the nano-crystalline surface with the shell is significantly more stable to chemical oxidation, which occurs only slowly.

4.2 preparation of Water-soluble nanocrystals with coupled polymers in organic solution

Nanocrystals capped with TOPO were prepared as in example 4.1 and dissolved in chloroform together with an excess of 3-mercaptopropionic acid. The mixture was first sonicated for about 1 hour and then allowed to stand overnight at room temperature until a large precipitate formed in the solution. The precipitate was collected by centrifugation and the free 3-mercaptopropionic acid was removed by washing several times with acetone. The resulting quantum dots covered with 3-mercaptopropionic acid were simply dried with argon and then dissolved in anhydrous DMF. To this solution, excess EDC and NHS were added, followed by stirring at room temperature for about 30 minutes to activate and form a cross-linking interface. Polyaziridine (Sigma-Aldrich Pte Ltd) with a molecular weight of 1200 (usually Molecular Weight (MW) 400-60000 is suitable) dissolved in anhydrous DMF was added dropwise through an addition funnel with vigorous stirring. After the polyethylenimine solution was added in its entirety, the reaction was continued overnight at room temperature for coupling the polymeric second layer to the capping reagent. The DMF solvent was then removed by rotary evaporation under reduced pressure, followed by dissolution in water. The polymer coated quantum dots were further purified by washing twice with ether.

5.1 preparation of Water-soluble nanocrystals with Cross-linked shells in aqueous solution

Quantum dots coated with TOPO were prepared using the method described in 2.1. The (CdSe) -ZnS core-shell nanocrystals thus formed were dissolved in chloroform with a large excess of 3-mercaptopropionic acid and a few drops of pyridine. The mixture was sonicated for about 2 hours and stirred at room temperature overnight. The precipitate formed was collected by centrifugation and washed with acetone to remove excess acid. The residue was dried briefly with argon. The resulting nanocrystals are then dissolved in water or a buffer solution, the nanocrystals being coated with carboxylic acid molecules that form a first layer covering/surrounding the nanocrystal core. Before use, the nanocrystals in aqueous solution were centrifuged again, filtered through a 0.2 μm filter, degassed with argon and stored at 25 ℃.

The above-mentioned nanocrystals covered with carboxylic acid are dissolved in an aqueous buffer system in order to form a cross-linking interface and subsequent polymerization with the coating agent layer contained in the second layer. EDC (1-ethyl-3- [ 3-dimethylaminopropyl ] -carbodiimide) and thioNHS (thio-N-hydroxysuccinimide) as crosslinking agents were added to the nanocrystal solution in a 1000-fold excess of 500-. The resulting solution was stirred at room temperature for 30 minutes, thereby activating the functional groups involved in the formation of the cross-linking interface. With stirring, the mixture containing the carboxylic acid-coated nanocrystals, EDC and thionhs was added dropwise to a diamino-carboxylmethyl ester (diamino-carbonyl methyl ester) solution in the same buffer. The above mixture was stirred at room temperature for 2 hours and then left overnight at 4 ℃ to form a cross-linking interface and covalently couple the coating agent contained in the second layer to the first layer. To release the water-soluble carboxyl group (i.e., hydrolyze the methyl ester bond) of the diamino-carboxyl ester (diamido-carboxylester) and thereby form a water-soluble second layer, 0.1N NaOH and ethanol were then added, and the solution was stirred at room temperature for an additional 6 hours. The solution was centrifuged to remove various solids and stored as an aqueous solution at 4 ℃ as a stock solution.

The obtained quantum dots can also be purified by organic solvent extraction. After the reaction (formation of the cross-linked interface and covalent coupling of the coating agent contained in the second layer to the first layer) is completed, the solution is extracted with ethyl acetate, thereby extracting the quantum dots with the polymer shell having an ester surface from the organic solvent. The organic solvents thus obtained were combined and dried, and then the organic solvent was removed by a rotary evaporator and dissolved in ethanol and 0.1N NaOH to hydrolyze ester bonds and form water-soluble nanocrystals. The solution was kept stirring at room temperature for 4 hours and then neutralized. The resulting clear solution was centrifuged to remove various traces of solids and stored as an aqueous solution at room temperature after degassing.

The physicochemical properties of the nanocrystals obtained with a crosslinked water-soluble shell of the invention were compared with those of (CdSe) -ZnS core-shell nanocrystals coated with only mercaptopropionic acid (MCA) or Aminoethanethiol (AET) as follows: addition of H to an aqueous solution of nanocrystals₂O₂And is caused to H₂O₂The final concentration of (2) reached 0.15mol/L and the chemical behaviour was followed by photon spectroscopy. For nanocrystals coated with MCA or AET only, oxidation of the nanocrystals was immediately detected and the nanocrystals precipitated within 30 minutes. In contrast, the nano-crystalline surface of the present invention with a shell is significantly more stable to chemical oxidation, which occurs only slowly.

In further experiments (data not shown), when 0.1M CdSO was added₄When the solution is added to (CdSe) -ZnS core-shell nanocrystals covered with MCA only or the shelled nanocrystals of the invention, the MCA-covered nanocrystals precipitate out of the solution very quickly. In contrast, the nanocrystals of the present invention are stably present in solution, which means that the addition of cadmium ions has no significant effect on the stability of the nanocrystals.

Similarly, the photochemical stability of the shelled nanocrystals was also significantly improved compared to the MCA-coated nanocrystals (data not shown). When exposed to UV light at a wavelength of 254nm, the above MCA-coated nanocrystals were found to precipitate out of solution within 48 hours, whereas the shell-containing nanocrystals of the present invention remained stable for 4 days. It was also found that the fluorescence intensity remained stable for a long time.

5.2 preparation of Water-soluble nanocrystals with crosslinked shells in organic solution

Nanocrystals covered with TOPO were prepared as in example 4.1 and dissolved in chloroform together with excess (3-N-ethanethiol) -1, 5-pentanediamine (pentane- (3-N-ethyl thiol) -1, 5-diamine) for forming the first layer. The mixture was left at room temperature overnight. The precipitate formed was collected by centrifugation, then washed with methanol and briefly dried with argon. The resulting nanocrystals were dissolved in anhydrous DMF (50 mL).

In another flask, 3-diethyl-carboxylate-1, 5-pentanedicarboxylic acid (pentane-3, 3-diethyl-carboxylic acid ester-1, 5-dicarboxylic acid) (contained as a capping agent in the second layer) was dissolved in DMF together with 5 equivalents of EDC and NHS and stirred at room temperature for 20 minutes under nitrogen. This solution is slowly added to the nanocrystal solution, thereby covalently coupling the nanocrystals to the capping agent. After stirring the resulting solution at room temperature for 2 hours, the DMF solvent was evaporated under reduced pressure using a rotary evaporation system. The resulting slurry was dissolved in 5mL of water, then 5mL of a 1M solution of EtONa/EtOH was added and stirred at room temperature for an additional 2 hours to form water-soluble bonds exposed to the solvent in the second layer. The resulting solution was washed twice with ether (5 mL. times.2) to remove various traces of additives or unreacted starting materials. Then neutralized with 0.1N aqueous HCl for storage. Further purification is performed by centrifuging the polymer coated nanocrystals in an acidic solution and redissolving the nanocrystals in water by adjusting the pH of the solution.

6.1 preparation of amphiphilic polymers

To 1g of dried CHCl at 300mL₃To a solution of poly (isobutylene-alt-maleic anhydride) in (b), the respective amine was added followed by DIPEA (1mL), and the mixture was stirred at 50 ℃ for 16 hours. In the evaporation of CHCl₃The above material was then suspended in water with a slight excess of NaOH with respect to the carboxyl groups on the polymer backbone. After evaporation of the water and DIPEA, the residue was dissolved in water and dialyzed against water for several days. After basification of the solution to pH 11 with 1M NaOH and subsequent evaporation, the product is obtained. Said poly (isobutylene-alt-maleic anhydride) spontaneously and in high yield with a monofunctional compound and at least bisThe functional compound (here the second component) reacts.

Typical data for forming the amphiphilic polymers 1-5 of the series of interest are provided below. The amphiphilic polymer was synthesized according to the previous scheme by grafting n-octylamine onto poly (isobutylene-alt-maleic anhydride) in different molar ratios.

Polymer 1

Poly (isobutylene-alt-maleic anhydride) 6000 was reacted with octylamine (0.05g, 0.4mmol) to give crystalline product 1(0.86g, 59%).¹H-NMR(400MHz，D₂O) δ: 3.25-2.90(m, 9H), 2.59(bs, 39H), 2.08(bs, 39H), 1.95-1.35(m, 81H), 1.34(bs, 62H), 1.13-0.65(m, 238H). The target composition is: 97% of carboxyl and 3% of caprylamide; the detected composition was: 96% of carboxyl and 4% of caprylamide.

Polymer 2

Poly (isobutylene-alt-maleic anhydride) 6000 was reacted with octylamine (0.15g, 1.2mmol) to give crystalline product 2(0.97g, 64%).¹H-NMR(400MHz，D₂O) δ: 3.30-2.90(m, 23H), 2.59(bs, 38H), 2.42-1.40(m, 152H), 1.23(bs, 116H), 1.15-0.70(m, 238H). The target composition is: 91% of carboxyl and 9% of caprylamide; the detected composition was: 89% of carboxyl and 11% of caprylamide.

Polymer 3

Poly (isobutylene-alt-maleic anhydride) 6000 was reacted with octylamine (0.42g, 3.2mmol) to give crystalline product 3(1.06g, 63%).¹H-NMR(400MHz，D₂O) δ: 3.28-2.85(m, 47H), 2.60(bs, 37H), 2.41-1.60(m, 81H), 1.47(bs, 68H), 1.24(bs, 253H), 1.12-0.75(m, 286H). The target composition is: 75% of carboxyl and 25% of caprylamide; the detected composition was: 75% of carboxyl and 25% of caprylamide.

Polymer 4

Poly (isobutylene-alt-maleic anhydride) 6000 was reacted with octylamine (0.84g, 6.5mmol) to give crystalline product 4(1.63g, 88%).¹H-NMR(400MHz，D₂O) δ: 3.25-2.85(m, 72H), 2.60(bs, 34H), 2.40-1.65(m, 73H), 1.48(bs, 87H), 1.24(bs, 379H), 1.12-0.65(m, 317H). The target composition is: 50% of carboxyl and 50% of caprylamide; the detected composition was: 61% of carboxyl and 39% of caprylamide.

Polymer 5

6.2 Synthesis of nanocrystalline (Quantum dot (QD)/Polymer Assembly)

2mg of purified QD was dissolved in THF (2mL), 0.5mL of 1.4mM polymer solution was added, followed by 5mL of water. The mixture was concentrated to 1mL using a rotary evaporator. The turbid aqueous suspension was filtered through a 0.8 μm filter, which was washed with 5mL of pure water. The supernatant was filtered again using a hydrophilic filter (0.2 μm) following the same procedure to give a clear aqueous QD solution. Excess water was removed using a rotary evaporator to give the desired concentration.

The above polymers 1-5 were used to transfer nanocrystals into water. Based on the absorption of the QD/polymer assembly, it is estimated that 10-80% of the quantum dots pass through the suspension step. However, the yield strongly depends on the polymer used, even more on the purity of the QDs (excess hydrophobic ligand leads to the formation of aggregates). The suspension efficiency of polymers 1-5 was then examined. The same initial amount of QDs in THF was used initially, and the same concentration of polymers 1-5 in water was used, but different concentrations of QDs in water were obtained (calculated by optical absorption). The final concentration of QDs in water depends on the ability of a given polymer to suspend the QDs in water. It was found that polymers with more n-octyl groups attached to the polymer backbone are more advantageous and this follows a general observation that the greater the number of binding sites (here hydrophobic n-octyl chains) based on hydrophobic-hydrophobic interactions, the higher the stability imparted to the assembly. Polymer 5 has a molar mass value 10 times higher than polymers 1-4 and the QD concentration in the final aqueous solution is lower, probably due to the formation of aggregates (few quantum dots are attracted to the same polymer chain) and problems associated with the filtration step. Careful purification to remove TOPO, TOP and hexadecylamine from the QD solution is critical to efficiently form stable QD/polymer assemblies. Residual alkyl ligands are likely to hinder the formation of monodisperse micelles and promote flocculation. Coating the QDs with a polymer and transferring the QDs into water does not have any significant effect on the basic light emitting properties of the QDs. Although the absorption spectrum shows a slight variation, the wavelength at maximum emission and the width of the emission spectrum remain substantially unchanged.

6.2.1 coating of nanocrystals

The hydrophobic nanocrystals (quantum dots (QDs)) were coated by suspending them in THF and then adding an aqueous solution of the polymer (e.g., TOPO coated CdSe/ZnS quantum dots). THF was removed by evaporation to give a stable colloidal solution of polymer-coated quantum dots. The resulting solution was clear, transparent, and exhibited long-term optical and colloidal stability. It should be noted that the above coating process can be performed on any nanoparticle substance in general, regardless of the surface of the nanoparticles.

6.3 example 4: NIPAM Quantum dot copolymerization

Polymerization is carried out by directly mixing the acrylic monomer and the polymer coated QD in water or by adding one component to a solution of the other component in the presence of a suitable initiator. Depending on the polymer used and the reaction conditions (e.g. temperature, initiator, concentration of ingredients) microspheres of different sizes may be obtained, said microspheres being embedded with different numbers of QDs covalently attached to the polymer chain.

Typical polymer procedure:

in a 50mL three-necked flask, the NIPAM (0.0503g), the solution coated with QD in the form of an acrylate polymer-ammonium salt (8mg), and 20mL of water were deoxygenated by 15 vacuums/argon evacuations. The temperature is raised to 70 ℃ and initiator K is added₂S₂O₈(0.0319g, in 1mL of H₂In O). Followed by addition of 4mL of H over 75 minutes₂Solutions of the crosslinkers MBAAM (0.0285g) and NIPAM (0.2526g) in O. After the addition was complete, the mixture was held at 71 ℃ for an additional 4 hours and cooled to room temperature. Purification was performed by dialysis (50kD membrane) and centrifugation to remove larger particles. Thus, an aqueous solution of submicron particles of PNIPAM with QDs was obtained.

Direct cross-linking polymerization of quantum dots

By adding an initiator (e.g. K) to an aqueous solution of quantum dots coated with a polymerisable amphiphilic polymer₂S₂O₈) Thereby carrying out polymerization. The crude reaction mixture was dialyzed to give a water-processable and suspendable solid polymer membrane loaded with a high level of QDs.

Typical polymerization procedure:

in a 50mL flask, the mixture was poured into a flaskThe coated QD in the form of the acrylamide polymer-sodium salt (8mg) was deoxygenated with 20mL of water by 15 vacuums/argon evacuations. The temperature is raised to 70 ℃ and initiator K is added₂S₂O₈(0.0319g, in 1mL of H₂In O). The reaction temperature was maintained for 6 hours. Purification was performed by dialysis (50kD membrane). Purification yields a water-soluble polymer network with a high concentration of QDs.

6.4 microemulsion polymerization and copolymerization of QDs

[00206] Polymeric materials with polymerizable functional groups (as described in example 5.3) with coated QDs (e.g., as described in examples 5.1-5.2) are used for self polymerization and copolymerization with other water-soluble monomers in a reverse microemulsion process. Microspheres with controllable diameters in the range of 400nm-10 μm are obtained. The luminescence time of the microspheres is prolonged.

Typical polymerization procedure for microemulsion polymerization:

argon gas was passed through a solution of sorbitan monooleate (Span 80, 0.23g) in 10mL of paraffin oil (parafin oil) for 1 hour. A1 mL volume aqueous solution containing NIPAM (0.15g, 1.32mmol), N' -methylenebisacrylamide (0.013g, 0.06mmol), 8mg of QD and 10mg of polymer (similar to 1.3.2) was added to the oil phase and bubbling with argon was continued for an additional 1 hour. The emulsion was sonicated at 40 ℃ for 1 minute and then shaken at room temperature for 1 hour using a laboratory vortex shaker. The stable and homogeneous emulsion was cast onto Petri dishes (Petri dish), irradiated with UV lamps at a temperature of 5-15 ℃ for 4 hours in a nitrogen-fed crosslinking apparatus, and then left without irradiation in an inert atmosphere for 16 hours. Purification of the PNIPAM latex was performed by multiple washes and centrifuges with hexane followed by multiple washes and centrifuges with water.

Claims

1. A plant comprising at least one nanocrystal.

2. The plant according to claim 1, wherein the at least one nanocrystal is selected from the group consisting of nanocrystals of the following types:

a rare earth metal doped metal oxide of the formula MeAl₂O₄Re or Me₄Al₁₄O₂₅:R；

A binary nanocrystal having a chemical formula of M1A or M1O;

a ternary nanocrystal having a chemical formula of M1M2A, M1AB, or M1M 2O;

and

a quaternary nanocrystal having a chemical formula of M1M2 AB;

wherein M1 and M2 in the binary, ternary and quaternary nanocrystals are independently a metal selected from group II, group III, group IV and group VIII of the periodic system of elements; and, if present, A and B are, independently of one another, an element from group VI or group V of the periodic system of the elements; and

wherein Me in the rare earth metal-doped metal oxide is selected from the group consisting of Ca, Sr, and Ba; and

re in the rare earth metal-doped metal oxide is at least one element selected from the group consisting of Tb, Dy, Nd, Eu and Tm.

3. The plant according to claim 1 or 2, wherein said at least one nanocrystal does not comprise cadmium.

4. The plant according to any one of claims 1-3, wherein said at least one nanocrystal is water soluble.

5. The plant of claim 4, wherein a capping reagent is attached to the surface of the nanocrystal, and wherein the capping reagent forms a guest-host complex with a water-soluble molecule.

6. The plant of claim 5, wherein the covering agent is a molecule having the following molecular formula:

H_aX-Y-Z，

wherein,

x is an end group selected from S, N, P or O ═ P,

a is an integer of 0 to 3,

y is a group having at least three main chain atoms, and

z is a hydrophobic end group.

7. The plant of claim 5, wherein said water soluble molecule is a compound containing a polar group exposed to a solvent.

8. The plant according to claim 7, wherein said water soluble molecule is selected from the group consisting of carbohydrates, cyclic polyamines, cyclic peptides, calixarenes, crown ethers and dendrimers.

9. The plant of claim 4, wherein the at least one nanocrystal comprises:

a low molecular weight capping reagent forming a second layer and having at least two coupling moieties covalently coupled to the capping reagent; and

at least one water-solubilizing group for imparting water solubility to said second layer.

10. The plant of claim 4, wherein the at least one nanocrystal comprises:

a capping reagent attached to the surface of the nanocrystal to form a first layer, the capping reagent having at least one coupling group;

a polymer forming a second layer and having at least one coupling moiety covalently coupled to at least one coupling group of the capping reagent.

11. The plant of claim 9 or 10, wherein the mulching agent is a molecule having the following molecular formula:

wherein,

x is an end group selected from S, N, P or O ═ P,

ra is a moiety containing at least 2 backbone carbon atoms,

y is selected from N, C, -COO-or-CH₂O-，

Z is a moiety containing a polar functional group,

k is a number of 0 or 1,

m is an integer of 1 to 3,

n is an integer of 0 to 3, and

n 'is an integer from 0 to 2, where n' is selected to satisfy the valence requirement of Y.

12. The plant of claim 10, wherein said polymer has the formula:

wherein,

j is a coupling moiety reactive with at least one coupling group of the capping reagent, and

m is an integer of at least 1.

13. The plant of claim 9, wherein the low molecular weight coating agent contained in the second layer comprises a water soluble molecule having the general formula:

wherein,

t is a hydrophilic moiety which is a hydrophilic moiety,

rc is a moiety containing at least 2 backbone carbon atoms,

g is selected from the group consisting of N, P, C or Si,

z 'is a coupling moiety, and Z' is a linker moiety,

m' is a number of 2 or 3,

n is 1 or 2, and

n 'is 0 or 1, where n' is selected to satisfy the valence requirement of G.

14. The plant of claim 4, wherein the surface of at least one of said at least one nanocrystal comprises an amphiphilic polymer that interacts non-covalently or covalently, said amphiphilic polymer having the general formula:

wherein m, o and p are each independently integers selected from about 3 to 400, and wherein the sum of m + o + p is selected from within the range of about 10 to 10000,

R²is a first aliphatic group having a main chain of about 3-20 carbon atoms and 0 to about 3 heteroatoms selected from the group consisting of N, O, S, Se and Si, and

R³is a second aliphatic group having a backbone of from about 3 to about 80 carbon atoms and from 0 to about 40 heteroatoms selected from N and O,

wherein R is³Having a copolymerizable group.

15. The plant according to any one of claims 1 to 14, wherein said nanocrystals are contained on the aerial part or all of the external surface of said plant.

16. The plant of claim 15, wherein said nanocrystals are contained in aerial parts of said plant.

17. The plant according to claim 15 or 16, wherein the aerial parts of the plant are flowers and/or leaves of the plant.

18. The plant according to any one of claims 1 to 15 or 17, wherein said plant is an artificial plant made of at least one polymer, paper, metal foil or a mixture of the above materials or a dry plant.

19. The plant of any one of claims 1-18, wherein said plant is a flowering plant.

20. The plant of claim 19, wherein said plant is selected from the group consisting of plants of Rosaceae, Sparganiaceae, Orchidaceae, Sparganiaceae, hyacinth, Malabaceae, Labiatae, Compositae, Geraniaceae, Solanaceae, Liliaceae, Primulaceae, Epilobii, Violaceae, Sonchusaceae, Malvaceae, Anacardiaceae, Araceae, Ericaceae, Theaceae, and Mirabilis.

21. The plant of claim 19, wherein said plant is selected from the group consisting of roses, orchids, hydrangeas, hyacinths, begonias, lavender, dahlia, geranium, petunia, tulip, lily, primrose, viola, gerbera, erythrina, hibiscus, pineapple, cyclamen, anthurium, rhododendron, leafflower, and camellia.

22. The plant according to claim 20, wherein said orchid is selected from a subfamily of the orchid family selected from the group consisting of orchidaceae, dendrolimiidae, vanillioideae, aryorchidaceae and pseudoorchidaceae.

23. The plant according to any one of claims 1 to 22, wherein the plant further comprises an organic dye.

24. The plant according to claim 23, wherein said organic dye is a fluorophore.

25. The plant according to claim 23, wherein said organic dye is selected from the group consisting of inks, chromophores such as azo compounds, lycopene, beta-carotene and anthocyanins, fluorescein and fluorescein.

26. A method of coloring a plant with at least one nanocrystal, the method comprising:

providing a solution containing at least one nanocrystal dissolved in the solution;

contacting said plant with said solution.

27. The method of claim 26, wherein the method comprises:

immersing the roots of the plant in the solution, or cutting the stem of the plant and immersing the ends of the cut stem in the solution.

28. The method of claim 26, wherein the method comprises:

contacting aerial parts of said plant or the whole plant with said solution.

29. The method of claim 28, wherein said contacting comprises dipping or spraying said solution on aerial parts of plants or whole plants.

30. The method according to claim 26 or 27, wherein the at least one nanocrystal is a nanocrystal or a mixture of nanocrystals as defined in any one of claims 1 to 14 and an organic dye as defined in any one of claims 23 to 25.

31. The method of any one of claims 26-30, wherein the solution comprises a suitable solvent or mixture of solvents in which the at least one nanocrystal is dissolved.

32. The method of claim 31, wherein the solvent is an aprotic solvent and/or a non-polar solvent or a protic solvent.

33. The method of claim 32, wherein the non-polar solvent is selected from the group consisting of mineral oil, hexane, heptane, cyclohexane, benzene, toluene, pyridine, methylene chloride, chloroform, carbon tetrachloride, carbon disulfide, dioxane, diethyl ether, 1-octadecene, 9-octadecene, diisopropyl ether, ethylene glycol monobutyl ether, tetrahydrofuran, and mixtures thereof.

34. The method of any one of claims 26-33, wherein the concentration of the at least one nanocrystal is from about 0.001 to 1 mol/L.

35. The method of any one of claims 26-34, wherein the solution further comprises an organic dye.

36. The method of claim 35, wherein the organic dye is a fluorophore.

37. The method of claim 35, wherein the organic dye is selected from the group consisting of inks, chromophores such as azo compounds, lycopene, beta-carotene, and anthocyanins, fluorescein, and fluorescein.

38. The method of claim 31, wherein the solvent is a high boiling point solvent.

39. The method of claim 38, wherein the solvent is selected from the group consisting of 1-octadecene and 9-octadecene.

40. Use of a nanocrystal or a mixture of nanocrystals as defined in any one of claims 1 to 14 with an organic dye as defined in any one of claims 23 to 25 for dyeing plants.

41. The use of claim 40, wherein aerial parts of the plant are stained.

42. Use according to claim 41, wherein the aerial parts are dyed by contacting the plants with a solution containing nanocrystals dissolved in a suitable solvent.

43. Use according to claim 42, wherein the contacting comprises immersing the roots of the plant in the solution or cutting the stem of the plant and immersing the ends of the cut stem in the solution.

44. The use according to claim 43, wherein the contacting comprises dipping or spraying the solution on aerial parts of the plant or on the whole plant.