JP6724638B2 - Immersion oil for microscope - Google Patents

Description

The present invention relates to immersion oils for microscopes.

Immersion oils are very commonly used in the microscopic field. The use of the immersion oil not only provides substantially less spherical aberration than the case where the immersion oil is not used, but also increases the numerical aperture of the objective lens to improve the resolution of the microscope.
As such an immersion oil, for example, as shown in Patent Document 1, a liquid immersion oil for microscopes containing a liquid diene polymer and an aromatic compound such as a diarylalkane and an alkylbenzene has been proposed.
On the other hand, in the microscope observation, the observation may be performed for a long time, and the immersion oil used is required to have a small change in viscosity during the period. However, the immersion oil for microscopes described in Patent Document 1 does not always meet the above requirements, and further improvement is required.

JP, 2008-38001, A

The present disclosure relates to a liquid immersion oil for a microscope, which contains a liquid diene polymer, an aromatic compound, and a higher alcohol.

The immersion oil for a microscope will be described in detail below. In addition, in this specification, "-" is used in the meaning including the numerical values described before and after it as the lower limit and the upper limit.
The immersion oil for microscopes contains a liquid diene polymer, an aromatic compound, and a higher alcohol. In particular, it is presumed that the inclusion of the higher alcohol improves the compatibility between the components, further suppresses the volatilization of the components, and consequently suppresses the viscosity change of the immersion oil for microscopes. The immersion oil for a microscope has a small change in refractive index with time and has low fluorescence.
Below, first, each component contained in the immersion oil for microscopes will be described in detail.

<Liquid diene polymer>
Liquid immersion oils for microscopes include liquid diene polymers. This polymer contributes to increasing the refractive index and viscosity of the immersion oil for microscopes.
The liquid diene polymer is a diene polymer in a liquid state at room temperature (25° C.). Although the melting point of the liquid diene-based polymer is not particularly limited, it is preferably −10° C. or lower from the viewpoint of handleability.
Further, the diene polymer is a polymer containing a repeating unit (structural unit) derived from a diene compound as a main component. Examples of the diene compound include conjugated diene compounds such as 1,3-butadiene and isoprene. Moreover, the said main component intends that the repeating unit derived from a diene compound is more than 50 mol% with respect to all the repeating units in a diene polymer. The content of the repeating unit derived from the diene compound in the diene polymer is preferably 70 mol% or more based on all repeating units in the diene polymer. The upper limit is not particularly limited, but may be 100 mol%.
In addition, the diene polymer may contain a repeating unit derived from a monomer other than the diene compound. Examples of other monomers include olefin compounds, acrylate compounds, methacrylate compounds, acrylonitrile and the like.
The type of liquid diene-based polymer is not particularly limited, and known liquid diene-based polymers can be used. Examples of the liquid diene-based polymer include diene homopolymers composed of diene monomers (diene compounds) having 4 to 12 carbon atoms, diene copolymers, and addition of these diene monomers and α-olefins having 2 to 22 carbon atoms. Examples thereof include copolymers with a polymerizable monomer (olefin compound). More specific examples include liquid polyisoprene, liquid polybutadiene, butadiene-isoprene copolymer, butadiene-acrylonitrile copolymer, and butadiene-2-hexyl acrylate copolymer.
The liquid diene-based polymer may have a functional group such as a hydroxyl group (for example, a polar group) in the molecule (for example, a molecular end).

As a preferred embodiment of the liquid diene polymer, an embodiment in which two or more kinds of liquid diene polymers are used in that the change in viscosity of the immersion oil for microscopes is further suppressed can be mentioned. That is, it is preferable that the immersion oil for a microscope contains two or more kinds of liquid diene-based polymers. The upper limit of the number of types of liquid diene polymer is not particularly limited, but is preferably 3 or less. Among them, 2 types are more preferable.

Examples of the above-mentioned two or more liquid diene polymers include, for example, a mode in which two or more liquid diene polymers having the same basic structure but different molecular weights are used, and two or more liquid diene polymers having different basic structures. Examples include a mode in which a system polymer is used.
It is preferable to use two or more types of liquid diene-based polymers having different basic structures because the above-described effect (the change in viscosity of the immersion oil for a microscope can be further suppressed) can be obtained. The reason is that the entanglement of each component changes due to the difference in the shape of the main chain and the position of the double bond between two or more liquid diene polymers having different basic structures, and the volatilization of the components is suppressed. It is presumed that it was done.
As the two or more liquid diene polymers, a combination of liquid polyisoprene and liquid polybutadiene is preferable.

When liquid immersion oil for microscopes contains liquid polyisoprene and liquid polybutadiene, the mass ratio of both (mass of liquid polyisoprene/mass of liquid polybutadiene) is not particularly limited, but is preferably 0.16 to 0.30. , 0.17 to 0.20 is more preferable.

The liquid diene polymer may be used alone or in combination of two or more.
The number average molecular weight of the liquid diene polymer is not particularly limited, but from the viewpoint of easily adjusting the viscosity of the immersion oil for microscopes, 1,000 to 100,000 is preferable, and 10,000 to 50,000 is more preferable. The above number average molecular weight is a polystyrene equivalent value measured by gel permeation chromatography (GPC) method.

<Aromatic compound>
The immersion oil for a microscope contains an aromatic compound. This aromatic compound contributes to increase the refractive index and decrease the viscosity of the immersion oil for microscopes.
The aromatic compound is an organic compound having an aromatic ring. Examples of the aromatic ring include an aromatic hydrocarbon ring and an aromatic heterocycle. The aromatic ring may be either a monocycle or a condensed ring, but a monocycle is preferred.
Examples of the aromatic hydrocarbon ring include a benzene ring, a biphenyl ring, a naphthalene ring, a terphenyl ring, an azulene ring, an anthracene ring, a phenanthrene ring, and a pyrene ring. As the aromatic heterocycle, for example, furan ring, thiophene ring, oxazole ring, pyrrole ring, pyridine ring, pyridazine ring, pyrimidine ring, pyrazine ring, triazine ring, oxadiazole ring, triazole ring, imidazole ring, pyrazole ring, Examples thereof include a thiazole ring, an indole ring, and an indazole ring.
The aromatic compound is preferably a so-called liquid aromatic compound. The liquid aromatic compound is an aromatic compound in a liquid state at room temperature (25° C.).

As a preferred embodiment of the aromatic compound, the compound represented by the general formula (1) can be mentioned in that the change in viscosity of the immersion oil for microscopes is further suppressed.

In general formula (1), Ar represents an aromatic hydrocarbon ring. The aromatic hydrocarbon ring may be monocyclic or polycyclic. Specific examples of the aromatic hydrocarbon ring are as described above. Of these, a benzene ring is preferable because the change in viscosity of the immersion oil for microscopes is further suppressed.
R ¹ represents a hydrocarbon group which may have a substituent. The number of carbon atoms contained in the hydrocarbon group is not particularly limited, and is preferably 1 to 30 and more preferably 1 to 10, for example. Examples of the hydrocarbon group include an alkyl group, a cycloalkyl group, an alkenyl group, an alkynyl group, and an aryl group. The alkyl group may be linear or branched.
The kind of the substituent that may be substituted on the hydrocarbon group is not particularly limited, and examples thereof include a halogen atom, an alkyl group, an aryl group, an alkoxy group, an oxyaryl group, a benzyloxy group, an acyl group, an aldehyde group, and a carboxyl group. Examples include groups, hydroxyl groups, and combinations thereof.
m represents an integer of 0 to 4. m is preferably 0 to 1 and more preferably 1 in terms of further suppressing the change in viscosity of the immersion oil for microscopes.
When m is 2 or more, a plurality of R ¹ may be the same or different. Further, two R ^1's may combine with each other to form a ring.

As a preferred embodiment of the aromatic compound, the compound represented by the general formula (2) can be mentioned in that the change in viscosity of the immersion oil for microscopes is further suppressed.

In formula (2), R ² and R ³ each independently represent an alkyl group or an alkoxy group. The number of carbon atoms contained in the alkyl group and the alkoxy group is not particularly limited, but is preferably 1 to 3.
p and q each independently represent 0 to 2. 0 is preferable for p and q.

The aromatic compounds may be used alone or in combination of two or more.

<Higher alcohol>
Immersion oil for microscopes contains higher alcohols. This higher alcohol contributes to improving the compatibility between the components.
The higher alcohol is an alcohol having 8 or more carbon atoms.
The number of carbon atoms contained in the higher alcohol is preferably 10 to 25, more preferably 10 to 20 and even more preferably 15 to 20 from the viewpoint of further suppressing the change in viscosity of the immersion oil for microscopes.
The alkyl group contained in the higher alcohol may be linear or branched, but is preferably branched.
Although the valence of the higher alcohol is not particularly limited, it is preferably monovalent.
Specific examples of higher alcohols include lauryl alcohol, myristyl alcohol, cetyl alcohol (cetanol), 2-hexyldecanol, cetostearyl alcohol, stearyl alcohol, isostearyl alcohol, and oleyl alcohol.
The higher alcohols may be used alone or in combination of two or more.

The above-mentioned immersion oil for microscopes contains known additives used for immersion oils for microscopes such as immersion oils for ordinary fluorescent microscopes, as long as the effect as the original immersion oil is not impaired. Good.

<Immersion oil for microscope and its manufacturing method>
The immersion oil for a microscope contains the above-mentioned components.
The content of the liquid diene polymer in the immersion oil for microscopes is not particularly limited, but it is 30 with respect to the total weight of the immersion oil for microscopes in that the viscosity change of the immersion oil for microscopes is further suppressed. 〜70 mass% is preferred, and 40-60 mass% is more preferred.
When two or more liquid diene-based polymers are used, the total content is preferably within the above range.

The content of the aromatic compound in the immersion oil for microscopes is not particularly limited, but is 20 to 60 relative to the total mass of the immersion oil for microscopes in that the change in viscosity of the immersion oil for microscopes is further suppressed. Mass% is preferable, and 30 to 50 mass% is more preferable.
When two or more aromatic compounds are used, the total content is preferably within the above range.

The content of the higher alcohol in the microscope immersion oil is not particularly limited, but is 0.1 to the total weight of the microscope immersion oil, in that the change in viscosity of the microscope immersion oil is further suppressed. 10 mass% is preferable, and 1 to 5 mass% is more preferable.
When two or more higher alcohols are used, the total content is preferably within the above range.

The mass ratio represented by {mass of higher alcohol/(mass of aromatic compound/mass of liquid diene-based polymer)} is not particularly limited and is often 4.0 to 20. In the point that the change in viscosity of the oil is further suppressed, 4.4 to 15 is preferable, and 4.5 to 8.0 is more preferable.
The mass ratio of the higher alcohol and the aromatic compound (mass of the higher alcohol/mass of the aromatic compound) is not particularly limited and is often 0.01 to 0.5, but the viscosity change of the immersion oil for microscopes is From the point of being further suppressed, 0.08 to 0.4 is preferable, and 0.08 to 0.15 is more preferable.
The mass ratio of the higher alcohol to the liquid diene-based polymer (mass of higher alcohol/mass of liquid diene-based polymer) is not particularly limited and is often 0.01 to 0.3. 0.05 to 0.25 is preferable, and 0.06 to 0.2 is more preferable, from the viewpoint that the change in viscosity is further suppressed.

The method for preparing the immersion oil for microscopes is not particularly limited, and a known method can be adopted. For example, the immersion oil for microscopes can be prepared by mixing the above-mentioned components by stirring. When mixing, the components to be used may be mixed at once, or the components may be mixed stepwise in various orders.

The above-mentioned immersion oil for microscopes can be suitably used as an immersion oil for ordinary microscopes, particularly as an immersion oil for fluorescent microscopes.

In addition, as preferable physical properties of the immersion oil for a microscope, the refractive index (wavelength 546 nm) is about 1.5170 to 1.5190, the Abbe number (wavelength 546 nm) is about 39 to 47, and the viscosity (temperature: 23° C.) is 50 to 1670 (mPa·s), and the transmittance (wavelength 380 to 900 nm) is 90% or more.

Hereinafter, the immersion oil for a microscope will be described in more detail with reference to Examples and Comparative Examples, but the present invention is not limited thereto.
The properties of the immersion oil for microscopes were evaluated according to the following methods.

<Evaluation method of immersion oil for microscopes>
(Evaluation of viscosity change)
A rheometer (“MCR301” manufactured by Anton Paar, parallel plate PP-25 (diameter 25 mm)) was used to measure the change in viscosity of the immersion oil for microscopes, which is the measurement object. More specifically, in a 40° C. environment, shearing is performed for 72 hours at a shear rate of 10 s ⁻¹ , and the viscosity of the immersion oil for microscopes before shearing and the immersion for microscopes after 72 hours of treatment have passed. The viscosity of each oil is calculated, and the rate of change in viscosity {(absolute value of difference between viscosity of immersion oil for microscope after 72-hour treatment and viscosity of immersion oil for microscope before shearing)/72} (unit : MPa·s/h) was determined. The obtained value was evaluated according to the following criteria. Practically, it is preferable that the evaluation is “Δ” or more.
"○": When the rate of change in viscosity is 10 or less "△": When the rate of change in viscosity is more than 10 and 15 or less "X": When the rate of change in viscosity is more than 15 In addition, the above evaluation was performed by observing with a microscope for a long time. It is carried out assuming the environmental conditions when performing it.

(Evaluation of change in refractive index)
Using a digital refractometer RX-9000α (manufactured by ATAGO Co., Ltd.), the change in the refractive index of the immersion oil for a microscope, which is a measurement object, was measured. More specifically, the refractive index of the microscope immersion oil before the shearing treatment used in the above (viscosity change evaluation) and the refractive index of the microscope immersion oil after 72 hours of treatment are measured, A change in refractive index (absolute value of difference between refractive index of immersion oil for microscope after 72 hours treatment and refractive index of immersion oil for microscope before shearing treatment) was obtained. The obtained value was evaluated according to the following criteria. Practically, the evaluation of “Δ” is preferable.
"○": When the change in refractive index was 0.0030 or less "△": When the change in refractive index was more than 0.0030 and not more than 0.0040 "X": When the change in refractive index was more than 0.0040 Is performed under the assumption of the environmental conditions when performing microscope observation for a long time.
Further, the above-mentioned refractive index is a refractive index at a wavelength of 589.3 nm.

(Evaluation of fluorescence intensity)
A fluorescence spectrophotometer F-2500 (manufactured by Hitachi High-Tech Co., Ltd.) was used to measure the fluorescence intensity of the immersion oil for a microscope, which is a measurement target. More specifically, under the environment of 22.5° C., the fluorescence intensity X at a wavelength of 450 nm of the fluorescence spectrum obtained at the excitation of a wavelength of 365 nm, and the fluorescence intensity at the wavelength of 485 nm of the emission spectrum obtained at the excitation of a wavelength of 405 nm. I asked for Y. The obtained value was evaluated according to the following criteria. Practically, it is preferable that the evaluation is “◯”.
"O": When the fluorescence intensity X is 3.78 or less and the fluorescence intensity Y is 1.38 or less "X": The fluorescence intensity X is more than 3.78, or the fluorescence intensity Y is When it is more than 1.38 As shown in Table 1, (fluorescence intensity evaluation) was performed only for Examples 1 and 2.

<Preparation of immersion oil for microscopes>
The components shown in Table 1 below were mixed at 25° C. for 10 minutes so that the contents (% by mass) shown in Table 1 would be obtained to prepare a microscope immersion oil. Table 1 shows the evaluation results of the obtained immersion oil for microscopes.

In Table 1 above, "LIR-30" is liquid polyisoprene (refractive index: 1.52, Abbe number: 38.9) manufactured by Kuraray Co., Ltd.
"LIR-15" is liquid polyisoprene (refractive index: 1.52, Abbe number: 40.6) manufactured by Kuraray Co., Ltd.
"LBR-352" is a liquid polybutadiene manufactured by Kuraray Co., Ltd. (refractive index: 1.51, Abbe number: 38.7).
"Phenylcyclohexane" is manufactured by Tokyo Chemical Industry Co., Ltd.
"Fineoxo call 180" is isostearyl alcohol manufactured by Nissan Chemical Industries, Ltd.
In addition, the numerical value in the component column in the said Table 1 represents the mass% of each component with respect to the total mass of the immersion oil for microscopes. The refractive index and Abbe number of each of the above components are values at a wavelength of 546 nm.

As shown in Examples 1 to 4 in Table 1, in the immersion oil for microscopes containing a predetermined component, it was confirmed that the viscosity change was suppressed and the desired effect was obtained.
In particular, when the mass ratio represented by {mass of higher alcohol/(mass of aromatic compound/mass of liquid diene polymer)} is 4.4 to 15, it was confirmed that the change in viscosity was further suppressed. Was done.
Further, as can be seen from the comparison between Examples 1 to 3 and 4, it was confirmed that when two or more liquid diene-based polymers were contained, the change in refractive index was further suppressed.
The immersion oils for microscopes of Examples 1 to 4 had a refractive index (wavelength 546 nm; 1.5170 to 1.5190) and an Abbe number (wavelength 546 nm; 39) described in "Type F" of JIS K2400 (2010). ˜47), viscosity (temperature 23° C.; 50 to 1670 mPa·s), and transmittance (wavelength 380 to 900 nm; 90% or more). The transmittance was measured using a quartz cell having an optical path length d of immersion liquid of 10 mm and a control cell containing pure water.
Further, as shown in Comparative Example 1, the immersion oil for microscopes containing the liquid diene polymer and the aromatic compound and containing no higher alcohol did not give the desired effect.

Claims (5)

A liquid immersion oil for a microscope, which contains a liquid diene polymer, an aromatic compound, and an alcohol having 10 to 25 carbon atoms . The immersion oil for microscopes according to claim 1, wherein two or more kinds of the liquid diene polymer are contained. The immersion oil for microscopes according to claim 1 or 2, wherein the liquid diene-based polymer contains liquid polyisoprene and liquid polybutadiene. The mass ratio represented by {mass of the alcohol having 10 to 25 carbon atoms /(mass of the aromatic compound/mass of the liquid diene polymer)} is 4.4 to 15. Immersion oil for microscopes as described in any one of 3 above. The aromatic compound is a compound represented by the general formula (1), the microscope immersion oil according to any one of claims 1-4.
(In General Formula (1), Ar represents an aromatic hydrocarbon ring. R ¹ represents a hydrocarbon group which may have a substituent. m represents an integer of 0 to 4. When m is 2 or more, two R ^1's may combine with each other to form a ring.)