CN116626008A - A Qualitative and Quantitative Detection Method of Lithium Isobutyrate-L-Proline Salt Based on Fluorescent Properties - Google Patents

Abstract

The invention discloses a qualitative and quantitative detection method of lithium isobutyrate-L-proline salt based on fluorescence characteristics, and the method is used in the production and synthesis process of lithium isobutyrate-L-proline salt to detect synthetic products In the qualitative detection of lithium isobutyrate and L-proline by solvent method, the obtained white solid is added into a non-fluorescent solvent to prepare a solution, and the solution is prepared by one of the following methods or A variety of detection: ① irradiate under the 365nm dark box type ultraviolet analyzer to observe whether there is fluorescence phenomenon; ② use ultraviolet-visible absorption spectrum detection to observe its absorption peak; ③ use fluorescence spectrum detection to observe its fluorescence peak; ④ use fluorescence spectrometer Detect and observe the characteristic peaks in the two-dimensional spectrum; use the standard curve method to quantify them. The solution of lithium isobutyrate-L-proline salt in non-fluorescent solvent has unique fluorescence characteristics. Based on these unique characteristics, the qualitative and quantitative detection of lithium isobutyrate-L-proline salt is carried out.

Description

A Qualitative and Quantitative Detection of Lithium Isobutyrate-L-Proline Salt Based on Fluorescent Properties method

technical field

The invention belongs to the technical field of fluorescence detection, and in particular relates to a qualitative and quantitative detection method of lithium isobutyrate-L-proline salt based on fluorescence characteristics.

Background technique

Lithium salt can be used in the field of mental illness, and has good curative effect and preventive effect on recurrent episodes of mania and depression in bipolar disorder, and has a unique effect of preventing suicide risk. Recent studies have also found that lithium salts can exert neuroprotective effects by acting on GSK-3, WNT, AKT and neurotransmitters. Lithium salts have selective activity on GSK-3β. There are two main modes of action for lithium salts to inhibit GSK-3β. : (1) directly inhibit the activity of GSK-3β as a Mg ²⁺ competitive inhibitor; (2) indirectly inhibit the activity of GSK-3β by increasing the expression of phosphorylated AKT, phosphorylated GSK-3β (ser9) and MCL-1 . This protective effect has potential applications in the prevention and treatment of neurodegenerative diseases, including Alzheimer's disease. Lithium salts currently commonly used in clinical practice are lithium carbonate, lithium citrate, lithium acetate and long-acting lithium salts, among which lithium carbonate is the most commonly used.

Due to the potential kidney damage and thyroid function damage in the long-term medication of inorganic acid lithium in the treatment of mental diseases, it will also lead to the disorder of blood pH value, cause metabolic acidosis, and increase the burden on the kidneys. During the research and development, it was found that a variety of small molecular organic acids including butyric acid, isobutyric acid, valproic acid and folic acid have important effects on the central nervous system, can effectively relieve anxiety and depression and other emotional abnormalities, and can Delays degeneration of the central nervous system. Therefore, the development of organic acid salts is expected to change the in vivo distribution defects of existing inorganic lithium salts.

The company has developed a new type of lithium organic acid amino acid salt, Chinese patent CN114081881, an organic lithium amino acid salt, crystal form, composition and application; lithium organic acid (lithium isobutyrate) and amino acid (L-proline ) is added into an appropriate amount of solvent and prepared by a single solvent method or a mixed solvent method, and the obtained white solid has better safety while strengthening the curative effect on central nervous system diseases. In the research and development, we also found that lithium isobutyrate-L-proline salt has unique fluorescence characteristics. Therefore, we provide an organic acid lithium amino acid salt (lithium isobutyrate-L-proline) Qualitative and quantitative detection methods for fluorescence properties.

Contents of the invention

The object of the present invention is to provide a kind of qualitative and quantitative detection method of lithium isobutyrate-L-proline salt based on fluorescence characteristic.

In order to achieve the above object, the present invention adopts following technical means:

The first aspect of the present invention provides a qualitative detection method based on fluorescence characteristics of lithium isobutyrate-L-proline salt, which is used in the production and synthesis process of lithium isobutyrate-L-proline salt to synthesize Qualitative detection of the product, using lithium isobutyrate and L-proline to prepare a white solid during the synthesis of lithium isobutyrate-L-proline salt, the white solid is added into a non-fluorescent solvent to prepare a solution , the solution is detected by one or more of the following methods:

① Place the solution in a box-type ultraviolet analyzer, irradiate it with 365nm ultraviolet light, and observe its fluorescence phenomenon. If fluorescence phenomenon is observed, there is lithium isobutyrate-L-proline salt in the prepared white solid ;

②The solution is detected by UV-visible absorption spectrum with a UV-visible absorption spectrometer, and its absorption peaks are observed; if there are absorption peaks at 260nm and 320nm, and the 320nm absorption peak is higher than the 260nm absorption peak, then the prepared white solid has isophthalic acid Lithium butyrate-L-proline salt;

③The solution is detected by fluorescence spectrum with a fluorescence spectrometer, and its fluorescence peak is observed; if there is a fluorescence peak at 370nm, then there is lithium isobutyrate-L-proline salt in the prepared white solid;

④ Use a fluorescence spectrometer to detect the fluorescence spectrum of the solution, and observe its two-dimensional spectrum; if there are two characteristic peaks at 280/380nm and 380nm/427nm excitation/emission in the two-dimensional spectrum, and the excitation/emission characteristic peak at 380nm/427nm is strong Excite/emit characteristic peaks at 280/380nm, and there is lithium isobutyrate-L-proline salt in the prepared white solid.

Further, there are two characteristic peak bands in the two-dimensional spectrum, characteristic peak band 1: excitation wavelength 260-300nm, corresponding to emission wavelength 350-400nm; characteristic peak band 2: excitation wavelength 360-400nm, corresponding to chromogenic wavelength 410-450nm; the characteristic peaks at the excitation/emission wavelengths of 280/380nm and 380nm/427nm in the two characteristic peak bands have remarkable characteristics.

The second aspect of the present invention provides a method for quantitative detection of lithium isobutyrate-L-proline salt based on fluorescence characteristics, comprising the following steps:

① Weigh lithium isobutyrate-L-proline salt powder, set the milligram-level concentration, and use the doubling dilution method to prepare standard lithium isobutyrate-L-proline salt solution samples with different dilutions. Lithium butyrate-L-proline salt solution sample is detected by fluorescence spectrum with excitation/emission wavelength of 380/427nm, the corresponding fluorescence value is obtained, and lithium isobutyrate-L-proline salt is obtained by fitting the fluorescence value and concentration The standard curve of the solution;

② Fluorescence spectrum analysis is carried out on the lithium isobutyrate-L-proline salt solution to be tested using 380/427nm excitation/emission wavelength to obtain the fluorescence value;

③Using the fluorescence value of the lithium isobutyrate-L-proline salt solution to be tested obtained in step ②, the concentration of the lithium isobutyrate-L-proline salt solution to be tested can be obtained by using the standard curve.

Further, if the concentration of the obtained lithium isobutyrate-L-proline salt solution to be tested is at the microgram level, the following method is used for further detection:

① Weigh lithium isobutyrate-L-proline salt powder, set the concentration in micrograms, and prepare standard lithium isobutyrate-L-proline salt solution samples with different dilutions by doubling dilution method. Lithium butyrate-L-proline salt solution sample is detected by fluorescence spectrum with 280/380nm excitation/emission wavelength, and the corresponding fluorescence value is obtained. Lithium isobutyrate-L-proline salt is obtained by fitting the fluorescence value and concentration The standard curve of the solution;

② Fluorescence spectrum analysis is carried out on the lithium isobutyrate-L-proline salt solution to be tested using 280/380nm excitation/emission wavelength to obtain the fluorescence value;

③Using the fluorescence value of the lithium isobutyrate-L-proline salt solution to be tested obtained in step ②, the concentration of the lithium isobutyrate-L-proline salt solution to be tested can be obtained by using the standard curve.

The third aspect of the present invention provides a kind of lithium isobutyrate-L-proline salt quantitative detection method based on fluorescence characteristic, can also adopt following steps:

①Weigh the lithium isobutyrate-L-proline salt powder, set the concentration gradient and prepare standard lithium isobutyrate-L-proline salt solution samples with different dilutions. Amino acid salt solution samples are detected by ultraviolet-visible absorption spectrum at a wavelength of 260nm or 320nm to obtain light absorption intensity, and the standard curve of lithium isobutyrate-L-proline salt solution is obtained by light absorption intensity and concentration fitting;

② The lithium isobutyrate-L-proline salt solution to be tested is detected by ultraviolet-visible absorption spectrum at a wavelength of 260nm or 320nm to obtain the light absorption intensity;

③ According to the light absorption intensity of the lithium isobutyrate-L-proline salt solution to be tested obtained in step ②, the concentration of the lithium isobutyrate-L-proline salt solution to be tested can be obtained by using the standard curve.

Furthermore, the standard lithium isobutyrate-L-proline salt solution sample is prepared by dissolving lithium isobutyrate-L-proline salt powder in a non-fluorescent solvent.

Beneficial effects of the present invention

The present invention has the following beneficial effects: the aqueous solution of lithium isobutyrate-L-proline salt irradiates and emits fluorescence under a 365nm dark box type ultraviolet analyzer, and has obvious absorption peaks at 260nm and 320nm of the ultraviolet-visible absorption spectrum, and The intensity of the absorption peak shows a positive correlation with the concentration. The lithium isobutyrate-L-proline salt is detected by fluorescence spectrum, and there is an obvious fluorescence peak at 370nm, and the fluorescence value shows a positive correlation with the concentration of the solution. In the two-dimensional graph of the fluorescence spectrometer, there are two prominent characteristic peaks at 280/380nm and 380nm/427nm excitation/emission, and at the excitation/emission wavelength of 380/427nm, the fluorescence value is positively correlated with the solution concentration, based on these Unique characteristics, during the production and synthesis process, qualitative detection of lithium isobutyrate-L-proline salt synthesis products; based on the positive correlation between the absorption peak and the concentration and the positive correlation between the fluorescence value and the concentration Concentrations were quantified.

Description of drawings

Fig. 1 shows the detection result of lithium isobutyrate-L-proline salt solution and contrast compound solution in the dark box type ultraviolet analyzer detection;

Fig. 2 shows the lithium isobutyrate-L-proline salt solution of gradient concentration under the detection result of ultraviolet-visible absorption spectrum;

Figure 3 shows the detection results of different compound aqueous solutions under the ultraviolet-visible absorption spectrum;

Fig. 4 shows the fluorescence spectrum detection result of the lithium isobutyrate-L-proline salt of gradient concentration in different solvents;

Fig. 5 shows the detection result of fluorescence spectrum of lithium isobutyrate-L-proline salt under different solvents;

Figure 6 shows a two-dimensional contour map of lithium isobutyrate-L-proline salt solution under fluorescence spectroscopy;

Figure 7 shows a three-dimensional map of lithium isobutyrate-L-proline salt solution under the fluorescence spectrum;

Fig. 8 shows lithium isobutyrate-L-proline salt solution in the detection of ultraviolet-visible absorption spectrum, the linear relationship between absorption peak and concentration at 320nm place;

Fig. 9 shows the linear relationship between the concentration y and the fluorescence value x of the high-concentration lithium isobutyrate-L-proline salt solution under the excitation/emission wavelength of 380/427nm;

Figure 10 shows the linear relationship between the concentration y and the fluorescence value x of the low-concentration lithium isobutyrate-L-proline salt solution at the excitation/emission wavelength of 280/380nm;

Among them, in Figure 1, from left to right, there are purified water, lithium diisobutyrate, lithium bisproline, lithium isobutyrate + proline equivalent mixture, lithium isobutyrate-L-proline Salt.

Detailed ways

Unless otherwise stated, implied from the context, or customary in the art, all parts and percentages in this application are by weight and the testing and characterization methods used are current as of the filing date of this application. Where applicable, the contents of any patent, patent application, or publication referred to in this application are hereby incorporated by reference, and equivalent patent families are also incorporated by reference, especially those disclosed by these documents with respect to the state of the art. Synthetic techniques, product and process design, definitions of polymers, comonomers, initiators or catalysts, etc. If the definition of a specific term disclosed in the prior art is inconsistent with any definition provided in the present application, the definition of the term provided in the present application shall prevail.

Numerical ranges in this application are approximations and therefore may include values outside the range unless otherwise indicated. Numerical ranges include all values from the lower value to the upper value in increments of 1 unit provided that there is a separation of at least 2 units between any lower value and any higher value. For example, if a composition, physical or other property (such as molecular weight, melt index, etc.) is stated as 100 to 1000, it means that all individual values are explicitly recited, such as 100, 101, 102, etc., and all subranges, such as 100 to 166, 155 to 170, 198 to 200, etc. For ranges containing numerical values less than one or containing fractional numbers greater than one (eg, 1.1, 1.5 etc.), one unit is considered to be 0.0001, 0.001, 0.01 or 0.1, as appropriate. For ranges containing single digit numbers less than ten (eg, 1 to 5), 1 unit is typically considered to be 0.1. These are only specific examples of what is intended to be expressed, and all possible combinations of numerical values between the lowest value and the highest value enumerated are considered to be expressly recited in this application.

When used with reference to a chemical compound, unless expressly stated otherwise, the singular includes all isomeric forms and vice versa (eg, "hexane" includes all isomers of hexane, individually or collectively). In addition, references to "a", "an" or "the" include plural forms unless expressly stated otherwise.

The terms "comprising", "comprising", "having" and their derivatives do not exclude the existence of any other components, steps or processes, and have nothing to do with whether these other components, steps or processes are disclosed in the present application. To remove any doubt, all compositions in this application using the terms "comprising", "comprising", or "having" may contain any additional additives, excipients or compounds, unless expressly stated otherwise. Conversely, the term "consisting essentially of" excludes from the scope of any hereinafter recited terms any other component, step or process, except those necessary for operational performance. The term "consisting of" does not include any component, step or process not specifically described or listed. Unless expressly stated otherwise, the term "or" refers to the listed members individually or to any combination thereof.

In order to make the technical problems, technical solutions and beneficial effects solved by the present invention clearer, the present invention will be further described in detail below in conjunction with the embodiments.

Example

The following examples are used herein to demonstrate preferred embodiments of the invention. It should be appreciated by those of skill in the art that the techniques disclosed in the examples which follow represent techniques discovered by the inventors to be employed in the practice of the invention, and thus can be considered preferred modes for its practice. However, those skilled in the art should understand from this specification that many modifications can be made to the specific embodiments disclosed herein, and the same or similar results can still be obtained without departing from the spirit or scope of the present invention.

Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which this invention belongs, and the disclosures cited herein and their cited materials are all incorporated by reference .

Those skilled in the art will recognize, or be able to ascertain through routine experimentation, many equivalents to the specific embodiments of the invention described herein. These equivalents are to be covered by the claims.

The preparation synthetic method of lithium isobutyrate-L-proline salt:

Single solvent method: add an appropriate amount of good solvent n-butanol to the equivalent chemical equivalent of lithium isobutyrate and L-proline, dissolve it just under heating and reflux, stir for about 3 hours, filter while it is hot, and naturally cool and crystallize , filtered the solid, and dried in vacuo to constant weight to obtain a white solid.

Mixed solvent crystallization method: add an appropriate amount of good solvent-ethanol to the equivalent chemical equivalent of lithium isobutyrate and L-proline, heat to reflux to make it just dissolve, and then add an appropriate amount of poor solvent-tetrahydrofuran to just precipitate solid, add a good solvent to make it re-dissolved, reflux for 3h, naturally cool and crystallize, filter the solid, vacuum dry to constant weight to obtain a white solid.

1. Qualitative detection of fluorescence characteristics of lithium isobutyrate-L-proline salt

(1) Dark box UV analyzer detection

Example 1

Take the lithium isobutyrate-L-proline salt sample and dissolve it in purified water, prepare a sample solution with a concentration of 100g/ml and put it into the sample tube.

The sample tube is placed in a dark box-type ultraviolet analyzer, and irradiated with 365nm ultraviolet light for detection, and the fluorescence phenomenon is observed.

Comparative example 1

Take the purified water and put it into the sample tube, put the sample tube in the dark box type ultraviolet analyzer, use 365nm ultraviolet light to detect, and observe its fluorescence phenomenon.

Comparative example 2

Take lithium bisisobutyrate sample and dissolve it in purified water, prepare a sample solution with a concentration of 100g/ml and put it into a sample tube.

The sample tube is placed in a dark box-type ultraviolet analyzer, and irradiated with 365nm ultraviolet light for detection, and the fluorescence phenomenon is observed.

Comparative example 3

Dissolve the lithium bisproline sample in purified water, prepare a sample solution with a concentration of 100g/ml and put it into the sample tube.

The sample tube is placed in a dark box-type ultraviolet analyzer, and irradiated with 365nm ultraviolet light for detection, and the fluorescence phenomenon is observed.

Comparative example 4

Lithium isobutyrate and proline were mixed in equivalent equivalents and the sample was dissolved in purified water to prepare a sample solution with a concentration of 100mg/ml and put it into a sample tube.

The sample tube is placed in a dark box-type ultraviolet analyzer, and irradiated with 365nm ultraviolet light for detection, and the fluorescence phenomenon is observed.

The results showed that: as shown in Figure 1, the equivalent mixtures of purified water, lithium diisobutyrate, lithium bisproline, lithium isobutyrate and proline did not show fluorescence phenomenon, only lithium isobutyrate-L - The sample tube of the proline salt solution has obvious fluorescence phenomenon and exhibits unique fluorescence characteristics.

(2) Ultraviolet visible absorption spectrum detection

Example 2

Preparation of stock solution: Weigh 0.3g lithium isobutyrate-L-proline salt powder sample and dissolve it in 9ml purified water solvent, the concentration is 33.33g/L.

Dilute the original solution by 2 times, 3 times, 4 times, 5 times, 6 times, 10 times, 20 times, 40 times, 60 times, 80 times, 100 times, 150 times, 200 times and set different concentration gradients.

The lithium isobutyrate-L-proline salt solution was detected by ultraviolet-visible absorption spectrum.

The results show: as shown in Figure 2, the lithium isobutyrate-L-proline salt solution has obvious absorption peaks at 260nm and 320nm, and the intensity of the absorption peaks is positively correlated with the concentration;

Example 3

Preparation of stock solution: use purified water to prepare a certain concentration of proline, lithium proline, lithium isobutyrate, isobutyric acid, lithium isobutyrate and proline equivalent mixture, lithium isobutyrate-L-proline Salt aqueous solution, the concentration of proline is 0.28986mol/L, the concentration of lithium proline is 0.27548mol/L, the concentration of lithium isobutyrate is 0.3546mol/L, the concentration of isobutyric acid is 2.69886mol/L, and the concentration of lithium isobutyrate -L-proline salt concentration is 0.03987mol/L. The aqueous solutions of different compounds were detected by UV-Vis absorption spectroscopy.

The results show that: as shown in Figure 3, the UV-visible absorption spectrum scans of different compounds in aqueous solution have obvious differences in absorption peaks, and the lithium isobutyrate-L-proline salt solution has obvious absorption peaks at 260nm and 320nm , is specific.

Comparative example 5

To prepare a solution, weigh 0.3g of lithium bisisobutyrate powder sample and dissolve it in 9ml of purified water solvent, with a concentration of 33.33g/L. It was detected by UV-Vis absorption spectroscopy.

The results show that: the lithium diisobutyrate solution has no obvious characteristic peak absorption peaks at 260nm and 320nm.

Comparative example 6

To prepare a solution, weigh 0.3 g of lithium bisproline powder sample and dissolve it in 9 ml of purified water, with a concentration of 33.33 g/L. It was detected by UV-Vis absorption spectroscopy.

The results show that: the lithium diisobutyrate solution has no obvious characteristic peak absorption peaks at 260nm and 320nm.

(3) Fluorescence spectrum detection

Example 4

Preparation of stock solution: Weigh 0.3g lithium isobutyrate-L-proline salt powder sample and dissolve it in 9ml purified water solvent, the concentration is 33.33g/L.

The stock solution was diluted 2 times, 3 times, 4 times, 5 times, 6 times, 10 times and 20 times to set different concentration gradients.

The lithium isobutyrate-L-proline salt solution was detected by fluorescence spectrum using a Hitachi fluorescence spectrometer.

Fluorescence spectrum detection results show that, as shown in Figure 4, the lithium isobutyrate-L-proline salt solution has an obvious fluorescence peak at 370nm, and the fluorescence value and concentration are positively correlated.

Comparative example 7

Preparation of stock solution: Weigh 0.3g of lithium isobutyrate-L-proline salt powder sample and dissolve it in 9ml of methanol solvent, with a concentration of 33.33g/L.

The lithium isobutyrate-L-proline salt solution was detected by fluorescence spectrum using a Hitachi fluorescence spectrometer.

Comparative example 8

Preparation of stock solution: Weigh 0.3g of lithium isobutyrate-L-proline salt powder sample and dissolve it in 9ml of methanol and water mixed at a ratio of 1:1 to form a solvent with a concentration of 33.33g/L.

The lithium isobutyrate-L-proline salt solution was detected by fluorescence spectrum using a Hitachi fluorescence spectrometer.

The results show that, as shown in Figure 5, in different solvents, the lithium isobutyrate-L-proline salt solution has fluorescence characteristics, and in different solvents, the fluorescence wavelength and the corresponding fluorescence value are different. The detection of the fluorescence spectrum of the -L-proline salt solution has a certain impact, which will cause a certain shift in the fluorescence emission wavelength, and at the same time, the solvent can be used to shift the fluorescence wavelength of the lithium isobutyrate-L-proline salt solution To optimize the method of fluorescence spectrum detection, so that it has a better effect in specific detection.

Comparative example 9

Weigh 0.3g of isobutyric acid sample and dissolve it in 9ml of purified water solvent, the concentration is 33.33g/L; the isobutyric acid solution is detected by fluorescence spectrum with Hitachi fluorescence spectrometer.

As a result of fluorescence spectroscopy, the isobutyric acid solution has no obvious fluorescence characteristics at the emission wavelength of 370nm.

Comparative example 10

Weigh 0.3g of lithium isobutyrate sample and dissolve it in 9ml of purified water solvent, the concentration is 33.33g/L; the lithium isobutyrate solution is detected by fluorescence spectrum with Hitachi fluorescence spectrometer.

As a result of fluorescence spectroscopy, the lithium isobutyrate solution has no fluorescence characteristics at the emission wavelength of 370nm.

Comparative example 11

Weigh 0.3g proline sample and dissolve it in 9ml purified water solvent, the concentration is 33.33g/L; the proline solution is detected by fluorescence spectrum with Hitachi fluorescence spectrometer.

As a result of fluorescence spectroscopy, the proline solution has no fluorescence characteristics at the emission wavelength of 370nm.

Comparative example 12

Weigh 0.3g lithium proline sample and dissolve it in 9ml purified water solvent, the concentration is 33.33g/L; the lithium proline solution is detected by fluorescence spectrum with Hitachi fluorescence spectrometer.

As a result of fluorescence spectroscopy, the lithium proline solution has no fluorescence characteristics at the emission wavelength of 370nm.

Comparative example 13

The isobutyric acid solution, the lithium isobutyrate solution, the proline solution, and the lithium proline solution are formed into a mixed solution, and the mixed solution is detected by fluorescence spectrum with a Hitachi fluorescence spectrometer.

As a result of fluorescence spectroscopy, the mixed solution has no fluorescence characteristics at the emission wavelength of 370nm.

The results showed that only the lithium isobutyrate-L-proline salt solution had a fluorescence peak at 370nm, which was its unique fluorescence characteristic.

(4) Fluorescence spectrometer two-dimensional contour map and three-dimensional map detection

Example 5

The lithium isobutyrate-L-proline salt solution in Example 4 is detected by a F97PRO fluorescence spectrometer, the vertical coordinate is the excitation wavelength in the range of 200nm-900nm, the abscissa is the emission wavelength in the range of 200nm-900nm, and Two-dimensional map;

The lithium isobutyrate-L-proline salt solution in Example 4 is detected by a F98 fluorescence spectrometer, the vertical coordinate is the excitation wavelength, and the range is 250nm-500nm, and the abscissa is the emission wavelength, and the range is 250nm-600nm, to obtain The three-dimensional map is shown in Figure 7.

The result shows 1: As shown in Figure 6, lithium isobutyrate-L-proline salt has two characteristic peaks of 280/380nm and 380nm/427nm excitation/emission; among them, the yellow high and low fields are fluorescence values, and the red bar Lines represent excitation wavelength scattering peaks; may represent Raman scattering. As shown in Figure 7, lithium isobutyrate-L-proline salt has 280/380nm, and there is an excitation/emission wavelength peak near 380nm/427nm, the closer to the excitation/emission wavelength position of this band, the corresponding fluorescence The higher the response value.

The result shows 2: Figure 6 and Figure 7 belong to the two-dimensional contour map and three-dimensional map, and the two mutually confirmed that lithium isobutyrate-L-proline salt has two fluorescence characteristic peaks, and the 280/380nm characteristic peak is far away from the red Raman scattering peaks, so the characteristic peaks here are not easily affected by Raman scattering peaks, and are more suitable for qualitative and quantitative analysis of samples that are prone to ultra-low concentrations. Since the characteristic peak at 380nm/427nm is close to the Raman scattering peak, the characteristic peak here is easily interfered by the scattering peak, which is suitable for qualitative and quantitative analysis of high-concentration sample solutions. It can be seen from the three-dimensional spectrum that under the same concentration conditions, the characteristic peak at 380nm/427nm has a higher fluorescence response value than the characteristic peak at 280/380nm, and the characteristic peak shape is more obvious.

The result shows 3: It can be seen from Figure 6 and Figure 7 that lithium isobutyrate-L-proline salt has characteristic peak range 1: excitation wavelength 260-300nm, corresponding emission wavelength 350-400nm; characteristic peak range 2 has : The excitation wavelength is 360-400nm, and the corresponding chromogenic wavelength is 410-450nm; there are fluorescence response values in these two corresponding bands, and the excitation/emission wavelengths of 280/380nm and 380nm/427nm belong to the characteristic peaks of the peak.

Due to the lithium isobutyrate-L-proline salt solution under the irradiation of 365nm ultraviolet light in the box-type ultraviolet analyzer, the fluorescence phenomenon is observed; when adopting the ultraviolet-visible absorption spectrometer to carry out the ultraviolet-visible absorption spectrum detection, it has 260nm and 320nm Absorption peak, and the absorption peak at 320nm is higher than the absorption peak at 260nm; when using a fluorescence spectrometer for fluorescence spectrum detection, it has a fluorescence peak at 370nm; there are two characteristic peaks at 280/380nm and 380nm/427nm excitation/emission in the fluorescence spectrum two-dimensional map , and the 380nm/427nm excitation/emission characteristic peaks are stronger than the 280/380nm excitation/emission characteristic peaks. And these fluorescence properties are unique to lithium isobutyrate-L-proline salt, so they can be used as qualitative detection during the preparation and synthesis of lithium isobutyrate-L-proline salt.

2. Quantitative detection of fluorescence characteristics of lithium isobutyrate-L-proline salt

Example 6

With purified water, the stock solution of lithium isobutyrate-L-proline salt solution in Example 2 and the different concentration gradient solutions diluted 2 times, 3 times, 4 times, 5 times, 6 times, and 10 times were adopted UV-visible Absorption spectroscopy detection.

The results show that there are miscellaneous peaks interference in the absorption peak of lithium isobutyrate-L-proline salt solution at 260nm, and the characteristic peak at 260nm is not as clear as the characteristic peak at 320nm, and the characteristic peak at 320nm is preferred for quantitative detection.

As shown in Figure 8, the absorption peak at 320nm has a linear relationship with the concentration, the linear equation is: y=0.0052+0.01382x, and the correlation coefficient is 0.9997.

Example 7

Prepare the lithium isobutyrate-L-proline salt solution with purified water to a high concentration of milligram level, adopt the doubling dilution method, set the concentration 0, 1mg/ml, 2mg/ml, 4mg/ml, 8mg/ml, use The F97PRO fluorescence spectrometer detects at the excitation/emission wavelength of 380/427nm, and the fluorescence values are: 0.01, 25.6, 53.64, 99.76, 195.2.

The results show: as shown in Figure 9, under the fixed excitation/emission wavelength of 380/427nm, the concentration y and the fluorescence value x show a good linear relationship, the linear equation is: y=-0.081426+0.041172x, the correlation coefficient is 0.9997 , with a good linear correlation. It shows that at high concentrations of the sample solution, the characteristic peaks here show a good linear relationship.

Example 8

Use purified water to prepare the lithium isobutyrate-L-proline salt solution to a low concentration of micrograms, and use the doubling dilution method to set the concentration of 0, 0.31ug/ml, 0.61ug/ml, 1.22ug/ml, 2.44ug /ml, 4.88ug/ml, 9.77ug/ml, 19.53ug/ml, 39.06ug/ml, 78.13ug/ml, 156.25ug/ml, using F98 fluorescence spectrometer to detect at the excitation/emission wavelength of 280/380nm, The fluorescence values were -0.19, 17.6, 22.21, 27.98, 40.84, 67.57, 120, 227.9, 433.5, 833.7, 1579, respectively.

The results show that: as shown in Figure 10, at a fixed excitation/emission wavelength of 280/380nm, the concentration y and the fluorescence value x exhibit a linear relationship; within the range of 0.31ug/ml to 156.25ug/ml, the linear relationship is relatively Well, the linear equation is: y=-2.2366+0.0992x, and the correlation coefficient is 0.9997. It shows that at a low concentration of the sample solution, the characteristic peaks here show a good linear relationship, and the linear range is large, and the interference at 280/380nm wavelength is small, and the accuracy and precision are high.

In summary, the results of Examples 1 to 8 show that lithium isobutyrate-L-proline salts belong to organometallic coordination salts and have specific fluorescence phenomena, indicating that there is a metal coordination bond in the organometallic compound molecule, and It is an organic bimolecular metal coordination salt, resulting in fluorescence phenomenon.

The lithium isobutyrate-L-proline coordination salt can be qualitatively and quantitatively detected through ultraviolet analyzer, ultraviolet-visible absorption spectrum detection and fluorescence spectrometer and other spectral instruments; and it can be qualitatively and quantitatively detected at high and low concentrations .

All documents mentioned in this application are incorporated by reference in this application as if each were individually incorporated by reference. In addition, it should be understood that after reading the above teaching content of the present invention, those skilled in the art can make various changes or modifications to the present invention, and these equivalent forms also fall within the scope defined by the appended claims of the present application.

Claims (6)

1. The qualitative detection method of the lithium isobutyrate-L-proline salt based on fluorescence characteristics is used for qualitative detection of synthetic products in the production and synthesis process of the lithium isobutyrate-L-proline salt, and is characterized in that: the method comprises the steps of obtaining a white solid in the process of preparing and synthesizing lithium isobutyrate-L-proline salt by using lithium isobutyrate and L-proline, adding the white solid into a non-fluorescent solvent to prepare a solution, and detecting the solution by adopting one or more of the following methods:

(1) placing the solution in a box-type ultraviolet analyzer, irradiating with 365nm ultraviolet light, observing fluorescence, and if fluorescence is observed, obtaining white solid with lithium isobutyrate-L-proline salt;

(2) carrying out ultraviolet-visible absorption spectrum detection on the solution by adopting an ultraviolet-visible absorption spectrometer, and observing an absorption peak; if the absorption peaks at 260nm and 320nm exist and the absorption peak at 320nm is higher than the absorption peak at 260nm, the prepared white solid contains lithium isobutyrate-L-proline salt;

(3) carrying out fluorescence spectrum detection on the solution by adopting a fluorescence spectrometer, and observing a fluorescence peak; if the fluorescence peak exists at 370nm, the prepared white solid contains lithium isobutyrate-L-proline salt;

(4) carrying out fluorescence spectrum detection on the solution by adopting a fluorescence spectrometer, and observing a two-dimensional map of the solution; if two significant characteristic peaks of 280/380nm and 380nm/427nm excitation/emission exist in the two-dimensional spectrum, and the characteristic peak of 380nm/427nm excitation/emission is stronger than the characteristic peak of 280/380nm excitation/emission, lithium isobutyrate-L-proline salt exists in the prepared white solid.

2. The qualitative detection method for lithium isobutyrate-L-proline salt based on fluorescence characteristics according to claim 1, wherein the method comprises the following steps: two characteristic peak wave bands exist in the two-dimensional map, and the characteristic peak wave band 1 is: excitation wavelength is 260-300 nm, and the corresponding emission wavelength is 350-400 nm; characteristic peak band 2: excitation wavelength is 360-400 nm, and the corresponding color development wavelength is 410-450 nm; the characteristic peaks at the excitation/emission wavelengths 280/380nm,380nm/427nm in the two characteristic peak bands have significant characteristics.

3. A quantitative detection method of lithium isobutyrate-L-proline salt based on fluorescence characteristics is characterized in that: the method comprises the following steps:

(1) weighing lithium isobutyrate-L-proline salt powder, setting milligram-grade concentration, preparing standard lithium isobutyrate-L-proline salt solution samples with different dilutions by adopting a double-ratio dilution method, performing fluorescence spectrum detection on the standard lithium isobutyrate-L-proline salt solution samples by adopting 380/427nm excitation/emission wavelength to obtain corresponding fluorescence values, and fitting the fluorescence values with the concentration to obtain a standard curve of the lithium isobutyrate-L-proline salt solution;

(2) performing fluorescence spectrum analysis on the lithium isobutyrate-L-proline salt solution to be detected by adopting 380/427nm excitation/emission wavelength to obtain a fluorescence value;

(3) and (3) obtaining the fluorescence value of the lithium isobutyrate-L-proline salt solution to be detected obtained in the step (2) by adopting a standard curve.

4. The method for quantitatively detecting the lithium isobutyrate-L-proline salt based on the fluorescence characteristics according to claim 3, wherein the method comprises the following steps: if the concentration of the obtained lithium isobutyrate-L-proline salt solution to be detected is in microgram level, the method is adopted for further detection:

(1) weighing lithium isobutyrate-L-proline salt powder, setting microgram-level concentration, preparing standard lithium isobutyrate-L-proline salt solution samples with different dilutions by adopting a double-ratio dilution method, performing fluorescence spectrum detection on the standard lithium isobutyrate-L-proline salt solution samples by adopting 280/380nm excitation/emission wavelength to obtain corresponding fluorescence values, and fitting the fluorescence values with the concentration to obtain a standard curve of the lithium isobutyrate-L-proline salt solution;

(2) performing fluorescence spectrum analysis on the lithium isobutyrate-L-proline salt solution to be detected by adopting 280/380nm excitation/emission wavelength to obtain a fluorescence value;

(3) and (3) obtaining the fluorescence value of the lithium isobutyrate-L-proline salt solution to be detected obtained in the step (2) by adopting a standard curve.

5. The method for quantitatively detecting the lithium isobutyrate-L-proline salt based on the fluorescence characteristic according to claim 2, wherein the method comprises the following steps: the following steps may also be employed:

(1) weighing lithium isobutyrate-L-proline salt powder, setting concentration gradients, preparing standard lithium isobutyrate-L-proline salt solution samples with different dilutions, performing ultraviolet-visible absorption spectrum detection on the standard lithium isobutyrate-L-proline salt solution samples by adopting wavelength of 260nm or 320nm to obtain light absorption intensity, and fitting the light absorption intensity and the concentration to obtain a standard curve of the lithium isobutyrate-L-proline salt solution;

(2) carrying out ultraviolet-visible absorption spectrum detection on the lithium isobutyrate-L-proline salt solution to be detected by adopting the wavelength of 260nm or 320nm to obtain light absorption intensity;

(3) and (3) obtaining the concentration of the lithium isobutyrate-L-proline salt solution to be detected by adopting a standard curve according to the light absorption intensity of the lithium isobutyrate-L-proline salt solution to be detected obtained in the step (2).

6. The method for quantitatively detecting the lithium isobutyrate-L-proline salt based on the fluorescence characteristics according to any one of claims 3 to 5, wherein the method comprises the following steps: the standard lithium isobutyrate-L-proline salt solution sample is prepared by dissolving lithium isobutyrate-L-proline salt powder in a non-fluorescent solvent.