DE102004059704A1 - Spectroscopic detection of inorganic, organic or biological material in sample comprises adding host system for material to sample to give guest-host system, exciting system and comparing spectrum obtained with reference spectrum - Google Patents

Abstract

Spectroscopic detection (SD) of an inorganic, organic or biological material (I) in a sample comprises: (i) adding a host system for (I) to the sample; (ii) exciting the system comprising the sample and host system; (iii) determining the spectrum of the system based on a signal shift, and (iv) detecting the presence of (I) by comparing the spectrum with at least one reference spectrum. Independent claims are also included for: (A) use of a three-dimensional receptor as a host system for spectroscopic detection of chemical materials in a sample, especially using a procedure as above; (B) a measuring cell especially for carrying out a procedure as above; (C) a measuring module comprising at least one measuring cell as in (B), where the module has an arrangement (15) for securing the module onto the body of a spectroscopic device; (D) a method of determining the spectrum of a sample, especially in a measuring cell as in (B).

Description

The The present invention relates to the determination and detection of inorganic, organic or biological substances. The measuring method according to the invention allows in particular the presence and concentration of substances by means of spectroscopic devices, in particular UV and luminescence detectors to determine.

To the Detection of substances without their own spectroscopic properties, for example, a variety of metals and heavy metals belong, are conventional spectroscopic devices, like UV / Vis or fluorescence devices not suitable. Furthermore, the detection of individual substances in spectroscopic Mixtures, e.g. biological degradation agents, or the detection of substances in very low concentration with such devices extremely difficult or even impossible. To measure such substances were therefore consuming measurement methods as the mass spectroscopy develops, the more specialized spectroscopic Properties or other physical or chemical properties as a basis for measurement use. These methods include, in particular, atomic absorption spectroscopy (AAS) and Inductively Coupled Plasma Mass Spectroscopy (ICPMS). These methods are the standard methods, especially for the measurement of metals and heavy metals.

It is therefore an object of the present invention, a novel Procedure for Provide the provisions and evidence of substances. These Provision should be independent from the own spectroscopic properties of the to be proved Fabrics possible be. Furthermore, it is an object of the invention, a method with which the qualitative or quantitative determination of substances, in particular of organic substances and metals or Heavy metals or their salts by known spectroscopic Equipment, in particular UV and luminescence detectors, is possible.

These Tasks are solved with the features of the claims.

The The present invention is based on the basic idea of a host system for the to use substance to be determined by an interaction of the host system with the substance the spectrum of the system in one Way that is characteristic of the substance to be determined is changed, and thus one qualitative and by the intensity quantitative analysis of the Stoffs allows becomes. To determine the spectrum of the system of sample and host system the system is stimulated, for example with electromagnetic radiation irradiated, and the spectrum of the system depending on the wavelength or the signal shift determined. The wavelength of the electromagnetic Radiation is here, e.g. at UV / Vis, preferably between about 200 nm and about 800 nm. The spectrum obtained is then with compared to a reference spectrum. The reference spectrum can hereby the spectrum of the host system be. In particular, at least one maximum of innatency can be used to detect the substance be determined in the spectrum and its location with the location in Reference spectrum are compared.

requirement for a suitable host system is that the Interaction of the host system with the substance to be determined if possible immediately at room temperature and without any addition of catalysts or other reactants. The resulting system from host system and sample should be individually detectable spectroscopically and for Each substance to be detected should be distinguishable. Furthermore, that should Host system be very selective to the substance to be determined, ie preferably do not interact with other substances.

When Wirt system suitable systems include biopolymers based systems, porphyrin based systems, on phthalocyanines based systems, catenane-based systems or rotaxanes based systems. In particular, it has been found that these Systems different interactions with different molecules, atoms and ionize. The type of interactions is from the chemical Properties, such as the type of atoms and the electron density, and the geometric structure of the molecular systems. In many cases to lead the interactions due to π-π bonds, hydrogen bonds or metallic bonds to form aggregates or complexes. The substance to be determined may be any inorganic, organic or be biological substance. However, the method according to the invention is particular for the Detection of organic molecules, Metals and heavy metals and their salts.

The The invention is particularly based on the use of a three-dimensional receptor as a host system for spectroscopic Detection of chemical substances in a sample directed, in particular the use of any of the systems described above.

The The invention further provides a device for spectroscopic Detection of an inorganic, organic or biological substance in a sample for performing the method according to the invention ready. So can the host systems, for example, in a mobile device, such as on a test strip or in a test ampule, which enables the visual verification of a substance. Furthermore, by the invention is a measuring cell and a measuring cell having measuring module provided in the form of an add-on module to the market spectroscopic devices the performing the method according to the invention also by means of such conventional equipment allowed.

The Invention will now be described with reference to the accompanying drawings explained in more detail.

1 (a) to 1 (d) : show examples of host systems for use in accordance with the present invention.

2 Figure 4 shows preferred porphyrin systems (a) and a polyrotaxane system (b).

3 Figure 4 shows another preferred porphyrin system (a) and various possible interaction systems between the porphyrin system and metal cations (b) to (d).

4 : shows the UV-Vis spectrum of the host system 1d 2 (a) or of 2 B) with concentration-dependent E values.

5 : shows the spectrum of the host system 1d 2 with nickel ions.

6 : shows spectra of host system 1d 2 with sodium or rubidium ions (a) and spectra of the host system 87 3 (a) with Ag ⁺ (b) and other metals (c) to (e).

7 : shows schematically a measuring module according to the invention.

8th : shows schematically a measuring cell according to the invention.

Examples of molecular systems which can be used as host systems in the process according to the invention are described in US Pat 1 (a) to 1 (d) shown. In 1 (a) Examples of systems based on biopolymers are shown. 1 (b) shows porphyrin based systems. Examples of catenane-based systems are in 1 (c) shown. 1 (d) shows rotaxane-based systems. For the properties of the host systems which are of paramount importance in the context of the present invention, not only the chemical properties but mainly those properties which result from the geometrical structure of the systems. In particular, the nature of the interactions of the host systems with different molecules, atoms or ions depends on the chemical properties, for example the nature of the atoms and the electron density, but also on the geometric structure of the molecular systems. Of particular interest are the weak, non-covalent bonds. Through π-π bonds, hydrogen bonds, or metallic bonds, these interactions often lead to the formation of aggregates or complexes.

For the embodiments of the method according to the invention for the detection of metals and heavy metals described below, in particular those disclosed in US Pat 1 (b) . 2 (a) and 3 (a) shown on porphyrin-based systems of interest.

The systems described below are for the determination of metals, but also for the determination of aromatics suitable.

In the context of the present invention, water-soluble three-dimensional porphyrin systems, which are capable of forming electron or energy transfer systems by complexing with metals, are preferably used for the determination of metals. In principle, every substance to be determined, in particular every metal to be determined, is regarded as a guest, and a corresponding measure is made designed host in the form of a three-dimensional receptor. In particular, the host system should also interact at room temperature and without any addition of catalysts and other reactants with the substance to be determined. The resulting system should then be individually detectable spectroscopically and distinguishable for each individual metal. This principle is also called "finger print" principle. The host system should interact as selectively as possible with the substance to be determined, that is as possible have no interaction with other substances.

In the 2 (a) Due to their three-dimensional structures, porphyrin systems 1a to e shown form good host-guest bonds or metal complexes. Furthermore, this class of compounds additionally has good solubility due to the 8-fold phosphonium groups. To represent such three-dimensional receptors, several methods are available. In the most general method, molecular scaffolds are clamped together from their components by multiple linkages.

3 (a) shows another example of a porphyrin system for use according to the invention. In the 3 (b) to 3 (d) possible complexes are shown by the interaction between metal ions and the in 3 (a) shown porphyrin system arises. Metal ions can be in the center, as in 3 (b) , above and below the cage structure, as in 3 (c) shown, or a combination of these two structures, as shown in 3 (d) is shown.

While the spectra of metal ions in the ultraviolet or visual range or in fluorescence show indefinable signals, the spectra have the in 2 (a) shown porphyrin systems very characteristic and intense signals. An example of such a spectrum of the than 1d in 2 (a) shown system shows the 4 (a) , In particular, the spectrum of the dissolved in methanol porphyrin system 1d for wavelengths between 200 and 800 nm is shown. Significant intensity maxima can be seen in particular in the range of 301 nm, 411 nm, 523 nm and 582 nm.

If an interaction between the porphyrin systems and metal ions takes place, lead these due to the change the electron density of the formation of new bonds to shifts or changes the characteristic signals of the porphyrin systems. Because the different ones Metal ions lead to different interactions, giving each interaction an individual and metal-dependent spectrum. Thus, can an identification of the different metals due to the change of the Spectrum accessible become. The intensities The new signals are proportional to the metal concentration.

Since porphyrins and bipyridines have different complexing properties, systems such as those in 2 (a) 1d, which contains both porphyrins and bipyridines, of particular interest. The interaction and thus the formation of porphyrin-metal complexes can be observed, inter alia, by the shift of the porphyrin-typical Soret and Q bands in the UV / Vis spectra. The bipyridine forms the bipyridine metal complexes with metals through the participation of its two pairs of nitrogen electrons. The number of bipyridine moieties that can complex with a metal ion depends on the number of metal coordinates, the geometry of the complex, and steric hindrance. In addition to the two porphyrin rings, the porphyrin system 1d has four bipyridine units and thus six possible positions at which the metals can interact with the porphyrin system.

Both the in 3 (a) shown porphyrin system 87 as well as in 2 The porphyrin systems 1a to 1e shown were examined for their property as a three-dimensional receptor by their reaction with various metal ions of the periodic table. Since the porphyrin systems are well soluble in both aqueous and organic solutions, the investigations of their interactions with the various metal compounds in various solvents were tested.

at investigating the interactions of the porpyhrin system 1d with The metal ions were the UV / Vis spectra used, since these even with minimal amounts of substance can detect interactions. The intense color of the porphyrin system facilitates the examination in very dilute solution and makes the process even more sensitive to any kind of interactions.

For the solution of the porphyrin system 1d in the different solutions, the wavelengths at which intensity maxima in the spectrum show - the λ _max values - were determined. In the following examples some metallic compounds of the first and the fourth main group, the iron and the copper group were reacted with the solutions and the λ _max values were determined.

example 1

5 shows a comparison of the spectra of a DMF solution containing nickel ions, a DMF solution of the porphyrin system 1d, a DMF solution of the propyrin system 1d after the reaction with a nickel ion solution and an aqueous solution of the propyrin system 1d after the reaction with a nickel ion solution. The DMF solution containing nickel ions shows an indefinable spectrum, whereas the porphyrin system 1d in the DMF solution shows a porphyrin-type spectrum. The addition of the nickel solution to the solution of the porphyrin system shows a strong shift of the signal. This shift is characteristic of and identifiable by the presence of nickel ions in the DMF. Repeating the experiment in aqueous solution also observed a shift in the signals. Here, the shift of signals for the presence of nickel ions in water was characteristic and identifiable.

Furthermore, the detection of silver ions by their interaction with the system 87 from 3 (a) examined. The porphyrin double-decker was dissolved in an ethanol / water mixture and then mixed with silver acetate. The solution was stirred for about 20 minutes under light or with exclusion of light and the UV / Vis spectrum of the in 6 (d) System 87 measured with or without silver acetate in both light and absence of light. The interaction of the Ag ⁺ with the system 87 forms complexes such as those in 3 (b) to 3 (d) . shown Again, the signal shift clearly shows. Table 1 shows the shifts of the signals. It can be seen from the spectra shown or the numerical values shown in Table 1 that the silver ion causes a characteristic shift of the signals of the propyrin system 87.

Tab. 1: λ _max [nm] of 87 in ethanol / water before and after the displacement with Ag ⁺ in the absence of light and in the presence of light:

Example 2

Furthermore, the interaction between the 4,4'-bisbrom-methyl-1,1'-biphenyl porphyrin double-decker 87 and metal ions was investigated.

The porphyridine double-decker 87 was dissolved in EtOH / H ₂ O (1: 1) and reacted with metal salts at room temperature. The interaction between the porphyrin double-decker 87 and the metal cations is examined by the UV spectrum of the solutions given in Table 2.

Tab. 2: UV / VIS spectroscopic data of 87 before and after the displacement with Hg (OAc) ₂ , Pb (OAc) ₂ , Ni (OAc) ₂ , Co (OAc) ₂ , Cu (OAc) ₂

The corresponding spectra are in the 6 (c) to 6 (e) shown. The spectroscopic data show that the signals for solutions 4 and 5 are slightly different compared to the stock solution, which suggests that small metal ions Ni ²⁺ and Co ²⁺ , respectively, have occurred (see 6 (c) ).

The UV spectra for solutions 2 and 3 show significant changes in the signals compared to stock solution 1, indicating a change in the structures or very strong interactions between the porphyrin double-decker 87 and the metal cations Hg ²⁺ and Ph ²⁺ in neutral Solution can close (see 6 (d) and 6 (e) ).

The Mercury (II) cation and the lead (II) cation have large radii of 1.10 Å or 1.20 Å. Therefore, the binding of these cations to the nitrogen of the pyrroles unlikely in the center of the porphyrin ring at the same level.

Other macrocyclic compounds that can be used in particular by their interaction for the detection of aromatics in both UV / Vis and NMR or DSC are compounds from the group of polyrotaxanes. Exemplary here is the interaction of bis (1,5-naphtho) -38-crown-10 with the oligomer 2,7-di- (3,3'-sulfonic acid-4,4'-biphenyl-benzo [lmn] [3 , 8] phenanthroline-1,3,6,8-tetrazone in DMSO-d ₆ (see 2 B) ). The corresponding spectrum is in 4 (b) shown. Thus, the method of the invention is also for the detection of larger materials, such as aromatics, usable. Overall, it should be noted that organic, inorganic and biological substances can be detected spectroscopically by their interaction with a suitable host system.

In 7 a preferred embodiment of a measuring cell according to the invention is shown, which is suitable for carrying out the method according to the invention. The measuring cell is designed as an additional module for existing in the market spectroscopic devices. The measuring cell is the unit in which the host system comes together with the substance to be determined and in which the spectrum of the system formed by the interaction between host system and substance is measured. The measuring cell according to the invention can be used for all spectroscopic methods in which the radiation path length through the sample is a parameter, ie in particular in UV / VIS and fluorescence methods. The dimensions of a measuring module having a measuring cell according to the invention may be variable so that the module can be universally grown to different spectroscopic devices from different manufacturers. Through a software program and a corresponding interface between the module and a computer, the module can be operated and controlled from the computer of the spectroscopic device. The module has a sample feed 1 and a reference feed 2 on. The sample is taken through the sample introduction 1 by means of a flüssigkeitsför reducing pump 3 in the liquid reservoir 5 guided. Preferably, the liquid reservoir has a volume which is the volume of the measuring cell 9 equivalent. The measuring cell 9 is variable in length and has a heatable outer shell 8th for heating the measuring cell 9 on. To the length of the measuring cell 9 to make variable, the measuring cell has a movable rear cell wall 7 . 19 on, which is preferably made of quartz. Furthermore, a flexible cover strip 6 provided in the 7 and 8th rolled up. The measuring module has a housing 4 on and can by means of screws 15 be attached to the spectroscopic device.

Commercially available spectroscopic Cells or cuvettes have a fixed length. To the maximum intensity signal in the spectrum to limit to a value in the measuring range of the detector must be in In general, the concentration of the substance to be determined to be changed. higher Concentrations lead to an excess of the maximum measurable Intensity value, while very low concentrations the measurable range, so the sensitivity limit of the device, below.

In contrast, the measuring cell 9 of the present invention a variable cell length 20 on. Thus, the light path traveled by the electromagnetic radiation through the sample can be varied. Thus, with a very dilute solution, the optical path can be lengthened, and with a highly concentrated solution, the optical path can be shortened, so that in each case the maximum measured signal lies within the measuring range of the detector. Thus, regardless of the concentration of the sample, an optimal measurement is made possible in a single measuring operation and without a change or aftertreatment of the concentration of the sample.

According to a preferred method according to the invention, the measurement starts with a cell length for this purpose 20 , which corresponds approximately to the mean of the possible cell lengths. Subsequently, a fast scan is performed with a coarse pitch for the entire spectroscopic waveband and the maximum value of the signal is determined by software. For a small range around the wavelength at which the maximum signal was determined, one or more scans are performed. A stepper motor 10 , which is controlled by the software, varies the cell length 20 until an optimal cell length is reached. The optimum cell length can be determined, for example, so that the maximum signal is about 80% of the maximum measurable value, that is, the OD value of the spectroscopic detector. With this optimal cell length, the entire spectrum of the sample is then measured accurately, ie with a fine stepwise.

Then you can the cell temperature increases be, preferably at about 80 to 120 ° C, and again the entire spectrum be measured exactly. The measurements at higher temperature allow the Release of the substance to be determined from inclusions, for example from a matrix or collections, often in nature or in nature Solutions, especially with metal ions, occur or are formed.

By comparing the spectra at room temperature or at the elevated temperature either with stored in the software or measured in parallel in a second measuring cell reference spectra of the substance to be determined can be identified. As both the concentration of the host system and the cell length 20 is known, the software can continue to measure the concentration of the substance to be determined by comparing the measured spectrum with stored concentration curves. After the measurement process, the sample or the reference through the output 14 led out of the measuring cell and the cell automatically cleaned. The measuring module also has an input 12 and an exit 13 for a cooling liquid in order to cool the measuring cell, optionally after heating again or to bring to a predetermined temperature can.

In 8th a more detailed view of the variable measuring cell is shown. In addition to the already with reference to the 7 features described 8th the import opening 16 for the sample or reference. The cover 17 the measuring cell is preferably made of PTFE. Furthermore, the rigid front 18 the measuring cell 9 and by the stepper motor 10 over the movement arm 11 movable rear wall 19 the measuring cell 9 shown. Preferably, both the front and the rear wall of the measuring cell 9 made of quartz or glass. The arrows 21 indicate the electromagnetic radiation passing through the sample. Furthermore, a temperature sensor 22 located.

The Measuring cell according to the invention preferred for Measurements according to the method of the invention used. The measuring cell according to the invention can but also for normal spectroscopic measurements are used. It works as a cuvette with variable length.

Claims (25)

Method for the spectroscopic detection of a inorganic, organic or biological substance in a sample by Add a host system for the substance to be detected to the sample to be examined, Stimulating the system from sample and host system and determining a spectrum of the system in dependence the singal shift and Proof of the substance by comparison of the spectrum with at least one reference spectrum. The method of claim 1, wherein the system is characterized by Irradiation with electromagnetic radiation is stimulated and a Spectrum in dependence the wavelength is determined. The method of claim 1 or 2, wherein the sample gaseous, liquid or is fixed. The method of claim 1, 2 or 3, wherein the host system is a three-dimensional receptor. The method of claim 4, wherein the host system is a porphyrin-based system. The method of claim 5, wherein the host system a 5,5'-dimethyl-2,2'-bipyridine porphyrin double-decker 4,4'-Bismethyl-1,1'-biphenyl-Porphyrindoppeldecker or a 6,6'-dimethyl-2,2'-bipyridine porphyrin double-decker is. Method according to one of the preceding claims, wherein the substance to be detected is a metal or its salt. Method according to one of claims 1 to 6, wherein the to be detected Fabric is an Aromat. Method according to one of the preceding claims, wherein the host system with the substance to be detected by interaction forms a guest-host system. Method according to one of the preceding claims, wherein the wavelengths the electromagnetic radiation between about 200 nm and about 800 nm lie. Method according to one of the preceding claims, wherein an absorption spectrum, a transmission spectrum and / or a Flureszenzspektrum is determined. Method according to one of the preceding claims, wherein the reference spectrum is the spectrum of the host system. Method according to one of the preceding claims, wherein to detect the substance, at least one intensity maximum in the spectrum is determined becomes. The method of claim 13, wherein the chemical Substance by determining the shift of an intensity maximum in the spectrum of the system of sample and host system over the Spectrum of the host system is detected. Using a three-dimensional receptor as Wirt system for the spectroscopic detection of chemical substances in a sample, especially according to the method according to any one of the preceding claims. Use according to claim 14, wherein the three-dimensional Receptor is a three-dimensional porphyrin system. Use according to claim 14 or 15, wherein the host system a 5,5'-dimethyl-2,2'-bipyridine porphyrin double-decker, a 4,4'-bismethyl-1,1'-biphenyl porphyrin double-decker or a 6,6'-dimethyl-2,2'-bipyridine porphyrin double-decker is. Use according to any one of claims 13 to 16, wherein the host system with the substance to be determined by interaction a guest-host system forms. Measuring cell, in particular for performing the method according to one of claims 1 to 14, wherein the measuring cell has a sample volume for receiving a sample with an inorganic, organic or biological substance and a host system for the substance to be detected, wherein the system of sample and host System by a front ( 18 ) and a back wall ( 19 ), which limit the measuring volume, is irradiated with electromagnetic radiation, wherein at least one wall ( 18 . 19 ) of the measuring cell is movable so that the length of the path of the light can be changed by the measuring cell. A measuring cell according to claim 19, wherein the measuring cell further comprises a stepping motor ( 10 ) for changing the position of the movable wall ( 19 ) of the measuring cell. A measuring cell according to claim 19 or 20, wherein the measuring cell comprises a device ( 8th ) for heating the measuring cell. Measuring module having at least one measuring cell according to claim 19 or 20, wherein the measuring module comprises a device ( 15 ) for attaching the measuring module to a housing of a spectroscopic device. Measuring module according to claim 21, wherein the measuring module is an import ( 1 ) and an export ( 14 ) for filling or emptying the measuring cell. Method for determining a spectrum of a sample, especially in a measuring cell according to one the claims 19 to 22, with the steps: (a) Determining the spectrum in a predetermined wavelength range with a coarse wavelength step size, (B) Determining the wavelength, where the maximum signal occurs (c) determining the spectrum in a range around the wavelength, where the maximum signal occurs with a wavelength step size, which is smaller than the coarse wavelength step size in step (a) is, (d) varying the length of the path of the electromagnetic Radiation through the sample until an optimal length is reached, and (E) Measuring the spectrum in the predetermined wavelength range with a wavelength step size, which is smaller than the coarse wavelength step size in step (a) is. Method according to the claim 24, with the optimal path length then reaches the electromagnetic radiation through the sample is when the maximum signal is about 80% of the maximum determinable Signal corresponds.