阅读说明：本技术 一种清香型白酒的指纹图谱检测方法 (Fingerprint spectrum detection method for fen-flavor liquor ) 是由 杜思邈 王申 卜睿臻 赵宇 曲明 于 2019-12-10 设计创作，主要内容包括：本发明涉及一种清香型白酒的指纹图谱检测方法,包括以乳酸乙酯、乙酸乙酯、正丙醇、乙酸丁酯、异戊醇、2,3,5-三甲基吡嗪中至少一种为有效成分,并用所述有效成分进行气相色谱含量测定和气相色谱指纹图谱构建中的至少一个过程。本发明对清香型白酒指纹图谱进行了研究,建立了气相色谱指纹图谱测定条件,并进行了方法学考察,根据多批大生产样品,制订了清香型白酒气相色谱指纹图谱标准,在生产过程中可有效的规范生产操作,确保了清香型白酒类最终产品的质量稳定,也是其下游产品控制质量的保证。(The invention relates to a fingerprint detection method of fen-flavor liquor, which comprises the steps of taking at least one of ethyl lactate, ethyl acetate, n-propanol, butyl acetate, isoamylol and 2,3,5-trimethylpyrazine as an effective component, and carrying out at least one of gas chromatography content determination and gas chromatography fingerprint construction by using the effective component. The method provided by the invention is used for researching the faint scent type white spirit fingerprint, establishing the gas chromatography fingerprint determination condition, carrying out methodology investigation, making the faint scent type white spirit gas chromatography fingerprint standard according to a plurality of large-scale production samples, effectively standardizing production operation in the production process, and ensuring the stable quality of the final product of the faint scent type white spirit and the control quality of the downstream product.)

1. A fingerprint detection method of fen-flavor liquor is characterized by comprising the steps of taking at least one of ethyl lactate, ethyl acetate, n-propanol, butyl acetate, isoamylol and 2,3,5-trimethylpyrazine as an effective component, and carrying out at least one of gas chromatography content determination and gas chromatography fingerprint construction by using the effective component.

2. The method for detecting the fingerprint of the fen-flavor liquor according to claim 1, wherein the gas chromatography conditions are as follows: using LZP-950 capillary chromatography columns; carrier gas: nitrogen with flow rate of 0.5-2.0 ml/min; vaporization chamber temperature: 150 ℃ and 250 ℃; the sample amount is 0.1-0.5 mul; tail blowing: 40-60 ml/min; split-flow method sample injection, the split-flow ratio (12-18) is 1; a detector: a hydrogen flame ionization detector with the temperature of 210 ℃ and 250 ℃; the hydrogen flow rate is 30-50ml/min, and the air flow rate is 300-500 ml/min; temperature programming: the initial temperature is 50-80 deg.C, the temperature is kept for 5-10min, the temperature is programmed to 150 deg.C at 3 deg.C/min, the temperature is kept for 0.01-1min, the temperature is continuously programmed to 200 deg.C at 4 deg.C/min, and the temperature is continuously kept for 5-15 min.

3. The method for detecting the fingerprint of the fen-flavor liquor according to claim 1 or 2, wherein the gas chromatography conditions are as follows: using LZP-950 capillary chromatography columns; carrier gas: nitrogen gas with flow rate of 1.0 ml/min; vaporization chamber temperature: 200 ℃; the sample amount is 0.3 mul; tail blowing: 30 ml/min; injecting sample by a split flow method, wherein the split flow ratio is 15: 1; a detector: detector Temperature (TD)230 ℃; a hydrogen Flame Ionization Detector (FID); the hydrogen flow rate is 40ml/min, and the air flow rate is 400 ml/min; temperature programming: the initial temperature is 65 ℃, the temperature is kept for 8min, the temperature is programmed to 150 ℃ at 3 ℃/min, the temperature is kept for 0.1min, the temperature is continuously programmed to 200 ℃ at 4 ℃/min, and the temperature is continuously kept for 10 min.

4. The fingerprint detection method of fen-flavor liquor as claimed in claim 1, wherein the relative retention time of ethyl acetate to ethyl lactate is 0.315-0.321, the relative retention time of n-propanol to ethyl lactate is 0.500-0.506, the relative retention time of butyl acetate to ethyl lactate is 0.584-0.590, the relative retention time of isoamyl alcohol to ethyl lactate is 0.775-0.781, and the relative retention time of 2,3,5-trimethylpyrazine to ethyl lactate is 1.130-1.136.

5. The fingerprint detection method for fen-flavor liquor as claimed in claim 1 or 4, wherein the relative retention time of ethyl acetate to ethyl lactate is 0.318, the relative retention time of n-propanol to ethyl lactate is 0.503, the relative retention time of butyl acetate to ethyl lactate is 0.587, and the relative retention time of isoamyl alcohol to ethyl lactate is 0.778, and the relative retention time of 2,3,5-trimethylpyrazine to ethyl lactate is 1.133.

6. The fingerprint detection method of fen-flavor liquor as claimed in claim 1, wherein the relative peak area of ethyl acetate to ethyl lactate is 0.818-0.837, the relative peak area of n-propanol to ethyl lactate is 0.095-0.105, the relative peak area of butyl acetate to ethyl lactate is 0.081-0.091, the relative peak area of isoamyl alcohol to ethyl lactate is 0.223-0.233, and the relative peak area of 2,3,5-trimethylpyrazine to ethyl lactate is 0.127-0.137.

7. The fingerprint detection method of fen-flavor liquor as claimed in claim 1 or 6, wherein the relative peak area of ethyl acetate to ethyl lactate is 0.828, the relative peak area of n-propanol to ethyl lactate is 0.100, the relative peak area of butyl acetate to ethyl lactate is 0.086, the relative peak area of isoamyl alcohol to ethyl lactate is 0.228, and the relative peak area of 2,3,5-trimethylpyrazine to ethyl lactate is 0.132.

Technical Field

The invention belongs to the field of detection of fen-flavor liquor, and particularly relates to a fingerprint spectrum detection method of fen-flavor liquor.

Background

China is the origin of wine, is one of the earliest countries in the world for brewing wine, and the earliest legend is Dukang wine brewing. The wine comprises Chinese liquor, yellow wine, beer, wine, rice wine, medicated liquor, etc. Wherein the liquor is a special distilled liquor of China, takes yeast and yeast as saccharification leaven, takes starch as raw material, and is brewed by stewing, saccharification, fermentation, distillation, ageing and blending; the wine is colorless (or yellowish) and transparent, has aromatic and pure smell, is sweet and clean in mouth, has high alcohol content, and has compound fragrance with esters as main body after being stored and aged. The differences of the climate, raw materials, process and the like in the wide-range living things of China form the characteristics of various varieties and complex components of the wine. At present, the identification and evaluation of wine at home and abroad mainly comprise: sensory evaluation (mainly comprising color, aroma, taste and style, which is one of the main methods for identifying fake wine), physical and chemical index inspection (comprising alcoholic strength, total acid, total ester and solid), sanitation index (comprising methanol, fusel oil, cyanide, lead, manganese and food additive) and the like. In recent years, the technical content of the national famous white spirit and the local famous special white spirit is higher and higher, the quality of the white spirit is evaluated only by sense, so that the quality of the white spirit cannot be distinguished, and the development of new wine products and the supervision and control standard of the quality are limited. Therefore, the identification and analysis of the quality and components of the liquor become the work focus of liquor workers, and the quality stability of the liquor is also an urgent problem to be solved in the liquor industry.

The fingerprint refers to a spectrogram or an image of a chromatogram or a spectrum which can mark the characteristics of a sample obtained by properly processing the sample and adopting a certain analysis means such as the spectrum or the chromatogram. These maps or images are unique and representative, just like a human fingerprint, and are therefore visually referred to as fingerprints. The fingerprint spectrum can be used for measuring various chemical components of the sample without quantification, provides the outflow curve characteristic of the fragrance component of each white spirit, can also find out the related trace components of the white spirit and the quantity ratio relationship rule characteristic thereof, and can effectively embody the integrity and the comprehensive action of the product, so the fingerprint spectrum technology can be applied to the analysis and the quality identification of the white spirit, and provides a reference basis for identifying counterfeit and shoddy white spirits. At present, the fingerprint spectrum technology has certain research in the aspects of analysis of traditional Chinese medicines, tobacco and wines. In contrast, the traditional Chinese medicine fingerprint spectrum technology is mature, but the methods are similar in the spectrogram analysis process, so that the information in the spectrum is sufficiently explored by referring to each other, and the fingerprint spectrum of the wine is more scientifically established.

The application of fingerprint spectrum in liquor blending process, identification of true and false, quality control and classification of unknown liquor is very wide. These can be directly compared by fingerprint spectra, and qualitative identification can be carried out on the spectra according to the number of the peaks and the size of the peak shapes. In addition, the classification of unknown white spirit, the quality control of white spirit and other aspects often involve the similarity calculation of fingerprint spectrums, and the conventional technologies for establishing the fingerprint spectrums by carrying out quantitative identification on the fingerprint spectrums along with the development of scientific technologies include a spectrum technology and a chromatographic technology, such as an ultraviolet fingerprint spectrum, an infrared fingerprint spectrum, an atomic absorption fingerprint spectrum, an atomic fluorescence fingerprint spectrum and the like. The chromatographic techniques mainly comprise gas chromatography (combination) and high performance liquid chromatography (combination). Most of the white spirit is volatile substances, so the fingerprint spectrum of the white spirit is constructed by adopting a gas chromatography technology.

Disclosure of Invention

The invention aims to provide a fingerprint spectrum detection method of fen-flavor liquor, which can effectively standardize production operation in the production process, ensure the stable quality of the final product of the fen-flavor liquor and ensure the control quality of the downstream products.

A fingerprint detection method for fen-flavor liquor comprises the steps of taking at least one of ethyl lactate, ethyl acetate, n-propanol, butyl acetate, isoamylol and 2,3,5-trimethylpyrazine as an effective component, and carrying out at least one of gas chromatography content determination and gas chromatography fingerprint construction by using the effective component.

Further, the gas chromatography conditions were: using LZP-950 capillary chromatography columns; carrier gas: nitrogen with flow rate of 0.5-2.0 ml/min; vaporization chamber temperature: 150 ℃ and 250 ℃; the sample amount is 0.1-0.5 mul; tail blowing: 40-60 ml/min; split-flow method sample injection, the split-flow ratio (12-18) is 1; a detector: a hydrogen flame ionization detector with the temperature of 210 ℃ and 250 ℃; the hydrogen flow rate is 30-50ml/min, and the air flow rate is 300-500 ml/min; temperature programming: the initial temperature is 50-80 deg.C, the temperature is kept for 5-10min, the temperature is programmed to 150 deg.C at 3 deg.C/min, the temperature is kept for 0.01-1min, the temperature is continuously programmed to 200 deg.C at 4 deg.C/min, and the temperature is continuously kept for 5-15 min.

Further, the gas chromatography conditions were: using LZP-950 capillary chromatography columns; carrier gas: nitrogen gas with flow rate of 1.0 ml/min; vaporization chamber temperature: 200 ℃; the sample amount is 0.3 mul; tail blowing: 30 ml/min; injecting sample by a split flow method, wherein the split flow ratio is 15: 1; a detector: detector Temperature (TD)230 ℃; a hydrogen Flame Ionization Detector (FID); the hydrogen flow rate is 40ml/min, and the air flow rate is 400 ml/min; temperature programming: the initial temperature is 65 ℃, the temperature is kept for 8min, the temperature is programmed to 150 ℃ at 3 ℃/min, the temperature is kept for 0.1min, the temperature is continuously programmed to 200 ℃ at 4 ℃/min, and the temperature is continuously kept for 10 min.

Further, the relative retention time of ethyl acetate to ethyl lactate is 0.315-0.321, the relative retention time of n-propanol to ethyl lactate is 0.500-0.506, the relative retention time of butyl acetate to ethyl lactate is 0.584-0.590, the relative retention time of isoamyl alcohol to ethyl lactate is 0.775-0.781, and the relative retention time of 2,3,5-trimethylpyrazine to ethyl lactate is 1.130-1.136.

Further, the relative retention time of ethyl acetate to ethyl lactate was 0.318, the relative retention time of n-propanol to ethyl lactate was 0.503, the relative retention time of butyl acetate to ethyl lactate was 0.587, the relative retention time of isoamyl alcohol to ethyl lactate was 0.778, and the relative retention time of 2,3,5-trimethylpyrazine to ethyl lactate was 1.133.

Furthermore, the relative peak area of ethyl acetate relative to ethyl lactate is 0.818-0.837, the relative peak area of n-propanol relative to ethyl lactate is 0.095-0.105, the relative peak area of butyl acetate relative to ethyl lactate is 0.081-0.091, the relative peak area of isoamyl alcohol relative to ethyl lactate is 0.223-0.233, and the relative peak area of 2,3,5-trimethylpyrazine relative to ethyl lactate is 0.127-0.137.

Further, the relative peak area of ethyl acetate to ethyl lactate was 0.828, the relative peak area of n-propanol to ethyl lactate was 0.100, the relative peak area of butyl acetate to ethyl lactate was 0.086, the relative peak area of isoamyl alcohol to ethyl lactate was 0.228, and the relative peak area of 2,3,5-trimethylpyrazine to ethyl lactate was 0.132.

The invention has the beneficial effects that:

the method provided by the invention is used for researching the faint scent type white spirit fingerprint, establishing the gas chromatography fingerprint determination condition, carrying out methodology investigation, making the faint scent type white spirit gas chromatography fingerprint standard according to a plurality of large-scale production samples, effectively standardizing production operation in the production process, and ensuring the stable quality of the final product of the faint scent type white spirit and the control quality of the downstream product. In addition, the identification of the measured fingerprint adopts a traditional Chinese medicine chromatogram fingerprint similarity evaluation system provided by the State pharmacopoeia Committee, the operation is convenient and fast, the obtained similarity result is used for evaluating the fingerprint of the fen-flavor liquor product, and the conclusion is objective and accurate.

Drawings

FIG. 1 is a fingerprint spectrum of 10 batches of fen-flavor liquor; wherein, S1-S10 respectively correspond to 10 batches of fen-flavor liquor samples;

FIG. 2 is a standard fingerprint spectrum of fen-flavor liquor.

Detailed Description

It should be noted that the embodiments and features of the embodiments may be combined with each other without conflict. The raw materials used in the invention are all commercial products, and the specific implementation process is as follows:

1. name (R)

Name: fen-flavor liquor.

2. The source of the test sample is as follows: tianjin fishery wine industry, Limited liability company.

3. Preparation of test solution

Sucking a certain amount of Chinese liquor sample, filtering with 0.45 μm filter membrane, and collecting filtrate.

4. Preparation of reference solutions

The main effective components of the fen-flavor liquor are Ethyl acetate (Ethyl acetate), N-propanol (N-propanol), Butyl acetate (Butyl acetate), Isoamyl alcohol (Isoamyl alcohol), Ethyl lactate (Ethyl lactate), 2,3,5-trimethyl pyrazine (2,3,5-trimethyl pyrazine) and Triethyl phosphate (Triethyl phosphate). According to experimental study, ethyl lactate with stable peak time and moderate peak area is selected as a reference substance, and the reference substance is provided by Beijing Tanzhong ink quality testing technology Co., Ltd (batch No. 0431908 for content measurement).

5. Establishment of detection method

5.1 instruments, reagents and chromatographic conditions

The instrument comprises the following steps: agilent 7890A gas chromatography; agilent GC chromatography workstation.

Reagent: acetonitrile and methanol are used as chromatographic purity (Spectrum), water is used as ultrapure water, and other reagents are analytical purity.

A chromatographic column: capillary chromatography columns (LZP-950, Productivity series No. 0824803, 50 m.times.0.32 mm.times.1.0. mu.m, maximum service temperature 230 ℃).

Carrier gas: nitrogen and flow rate to be inspected; vaporization chamber temperature: 200 ℃; the sample amount is 1 mul; tail blowing: 30 ml/min;

a detector: detector Temperature (TD)230 ℃; a hydrogen Flame Ionization Detector (FID); the hydrogen flow rate was 40ml/min and the air flow rate was 400 ml/min.

5.2 selection of the flow Rate of the Carrier gas

The nitrogen flow rates are respectively 0.5, 1.0 and 2.0ml/min, and the influence of the three flow rates on the peak appearance time, the peak area, the peak type, the analysis time and the like of the base wine fingerprint is respectively considered, and the result shows that when the nitrogen flow rate is 1.0ml/min, the peak appearance time and the peak separation degree are also good, the integral analysis time is more reasonable within 60min, and therefore the nitrogen flow rate is finally determined to be 1.0 ml/min.

5.3 optimization of temperature Programming

The following temperature programming method is respectively adopted to investigate the fingerprint spectrum of the base wine.

① the initial temperature is 60 deg.C, the temperature is kept for 3min, the temperature is programmed to 180 deg.C at 3.5 deg.C/min, and the temperature is kept for 10 min.

② the initial temperature is 65 deg.C, the temperature is kept for 8min, the temperature is programmed to 150 deg.C at 3 deg.C/min, the temperature is kept for 0.1min, the temperature is continuously programmed to 200 deg.C at 4 deg.C/min, and the temperature is kept for 10 min.

As a result, the time for peak appearance and the separation degree of peaks are better when the temperature is programmed by ② method, and the number of peaks is larger in the analysis time, so that the method for finally determining the temperature programming is that the initial temperature is 65 ℃, the temperature is kept for 8min, the temperature is programmed to 150 ℃ by 3 ℃/min, the temperature is kept for 0.1min, the temperature is continuously programmed to 200 ℃ by 4 ℃/min, and the temperature is continuously kept for 10 min.

5.4 Split ratio selection

Selecting a split flow method for sample injection, setting split flow ratios as 15:1, 30:1 and 60:1, and respectively inspecting the peak emergence time, the peak area, the peak type and the analysis time of the base wine fingerprint, wherein the result shows that the peak emergence time and the peak separation degree are better when the split flow ratio is 15:1, the number of the peaks is more and the response value is proper, so that the split flow ratio is finally determined as 15: 1.

5.5 identification of main chromatographic peak in gas chromatography fingerprint of fen-flavor liquor

The main chromatographic peaks in the finished chromatogram can be assigned according to the retention time with the standard: s is ethyl lactate, No. 1 is acetaldehyde, No. 2 is ethyl acetate, No. 3 is an unknown peak, No. 4 is n-propanol, No. 5 is butyl acetate, No. 6 is isoamyl alcohol, No. 7 is 2,3,5-trimethyl pyrazine, No. 8 is diethyl pimelate. According to the physicochemical requirements in GB/T10781.2-2006 fen-flavor liquor, acetaldehyde No. 1 and diethyl pimelate No. 8 are non-index components, and No. 3 is an unknown peak, so that the three components are not subjected to content measurement. Under the fingerprint spectrum measuring condition, the measured fingerprint spectrum of the fen-flavor liquor can reflect the main flavor components of the fen-flavor liquor.

6. Verification of detection method

6.1 method repeatability test

6 parts of test solution is prepared in parallel according to the step 3 by taking the fen-flavor liquor with the batch number of 2019052901, and the results are shown in tables 1 and 2 according to the method.

TABLE 1 repeatability test results of the gas phase fingerprint spectrum of fen-flavor liquor (relative retention time of main peak)

TABLE 2 repeatability test results of the gas phase fingerprint spectrum of fen-flavor liquor (relative peak area of main peak)

The results show that the test method has good repeatability.

6.2 sample stability test

The fen-flavor liquor with the batch number of 2019052901 is taken and prepared in parallel according to the step 3, the stability is respectively checked for 0 hour, 3 hours, 6 hours, 9 hours, 12 hours, 24 hours, 36 hours and 48 hours, the test results are shown in tables 3 and 4.

TABLE 3 stability test results of the gas phase fingerprint spectrum of fen-flavor Chinese liquor (relative retention time of main peak)

TABLE 4 stability test results of the gas phase fingerprint spectrum of fen-flavor Chinese liquor (relative peak area of main peak)

The result shows that the retention time of all the common peaks in the test solution is basically consistent with the peak area of the main peak (accounting for more than 5 percent of the total peak area) (RSD is less than 1 percent), the similarity of the fingerprints obtained by 7 times of sample injection is calculated by using the fingerprint obtained by the 1 st sample injection as the reference and using the traditional Chinese medicine chromatogram fingerprint similarity evaluation system (2012.130723 version, published by the national pharmacopoeia committee), the result similarity accords with the technical requirements of the fingerprints, and the test solution can be stably measured within 48 hours.

The methodology investigation result shows that the method for determining the fingerprint of the fen-flavor liquor has good sample stability and method repeatability, and can accurately determine the fingerprint of the product.

7. Acquisition of standard fingerprint spectrum of fen-flavor liquor and determination of similarity limit

7.120 measurement of batch production finished product and acquisition of standard fingerprint

20 batches of finished products are all produced by Tianjin fishery wine industry, Limited liability company, and the batch numbers are respectively: 2019052901, 2019052902, 2019052903, 2019052904, 2019052905, 2019052906, 2019052907, 2019052908, 2019052909, 20190529010, 20190529011, 20190529012, 20190529013, 20190529014, 20190529015, 20190529016, 20190529017, 20190529018, 20190529019, 20190529020.

Preparing a test sample solution according to the step 3, and determining the liquid phase fingerprint spectrum superposition spectrum of 20 batches of finished products according to the method; obtaining standard fingerprints based on 20 batches of finished product fingerprints by using a traditional Chinese medicine chromatogram fingerprint similarity evaluation system; and manually calculating to obtain a standard fingerprint.

7.2 determination of similarity limit of fen-flavor liquor

And taking the standard fingerprints of the fen-flavor liquor generated by the similarity calculation software as reference, and calculating the similarity of the fingerprints of each batch of finished products by using a traditional Chinese medicine chromatographic fingerprint similarity evaluation system, wherein the result similarity is more than 0.90 (see table 5). According to the actual production, in order to effectively and comprehensively control the product quality, the similarity between the fingerprint of the fen-flavor liquor and the standard fingerprint is calculated by similarity software, and the similarity is more than 0.90.

Table 520 batches of similarity investigation results of fingerprint spectrums of fen-flavor liquor

7.3 determination of relative retention time of each characteristic peak and fluctuation range of relative peak area in fingerprint spectrum of fen-flavor liquor

And (3) importing the obtained detection data of 20 batches of finished product fingerprints into EXCEL, calculating the relative retention time and the relative peak area of each characteristic peak and a reference peak, and obtaining the detection results shown in tables 6 and 7.

Table 620 batch faint scent type white spirit fingerprint spectrum relative retention time detection result

Note: s is a reference peak

Table 720 batch faint scent type white spirit fingerprint relative peak area detection result

Note: s is a reference peak

The average value of the relative retention time and the relative peak area of each characteristic peak in 20 batches of finished fingerprint is respectively used as the relative retention time and the relative peak area value of each characteristic peak of the standard fingerprint, and the fluctuation range of the relative retention time and the relative peak area of each characteristic peak is limited according to the actual production in order to effectively and comprehensively control the product quality, which is shown in tables 8 and 9.

TABLE 8 Standard fingerprint data of fen-flavor Chinese liquor

Note: s is a reference peak

TABLE 9 relative retention time of standard fingerprint and fluctuation range of relative peak area of fen-flavor liquor

Note: s is a reference peak

8. Primary examination of stability of fingerprint spectrum of fen-flavor liquor

The standard fingerprints are used as reference, a traditional Chinese medicine chromatogram fingerprint similarity evaluation system is used for inspecting the fingerprint similarity change conditions of 10 batches of fen-flavor liquor so as to verify the stability, and the results are shown in a table 10.

Table 1010 batches of fen-flavor liquor room temperature sample retention fingerprint similarity stability investigation results

Test results show that the changes of the fingerprint spectra of the fen-flavor liquor are small within 12 months of sample retention at room temperature, the similarity is over 0.99, the product quality is proved to be stable, and the fingerprint spectrum standard can effectively control the quality of finished products.

While one embodiment of the present invention has been described in detail, the description is only a preferred embodiment of the present invention and should not be taken as limiting the scope of the invention. All equivalent changes and modifications made within the scope of the present invention shall fall within the scope of the present invention.