Fruit juice authenticity identification model and adulteration proportion determination method thereof

Abstract

The invention relates to the field of food detection, and provides a fruit juice true and false identification model and an adulteration proportion determination method thereof.The fruit juice with pulp and fruit juice without pulp are purchased to serve as samples to be pretreated for use, the uniform samples are placed in a plastic centrifugal tube, a proper amount of water is added for ultrasonic extraction and centrifugation, and an adulteration ratio is obtained; the method comprises the following steps: firstly, performing solid-phase extraction on a sample to obtain a supernatant, fixing the volume of the supernatant to obtain a sample extracting solution, performing pre-leaching and activating treatment on a solid-phase extraction small column, adding the sample extracting solution into the small column, collecting filtrate after abandoning part of initial filtrate, and finally placing the collected filtrate under a specific high-performance liquid chromatography condition to finish the analysis and determination of the sample. The contents of L-malic acid and D-malic acid in the sample are calculated, and the authenticity and adulteration proportion of the product can be determined according to the formula [1-(1-CL / CT) * 2] * 100. The fruit juice identification technology with high sensitivity, short detection period and wide adaptability is realized through an accurate detection process and a scientific calculation method, and the method is simple, high in practicability and high in accuracy.

Description

Technical Field

[0001] The invention relates to the field of food detection, in particular to a fruit juice authenticity identification model and an adulteration ratio determination method thereof. Background Art

[0002] With the improvement of people's living standards and the change of demand, various fruit juice drinks have emerged and are increasing in the market. However, at the same time, people have gradually discovered that some "fruit juice drinks" that are not in line with their names or even fake are flooding the market. Among them, some drinks have very low juice content, which seriously damages the interests of consumers.

[0003] Malic acid, also known as 2-hydroxysuccinic acid, has two enantiomers, D- and L-malic acid. L-malic acid is widely found in fruits (especially apples, pears, bananas, citrus fruits, etc.) and their products. The human body only has L-malate dehydrogenase, so the human body can only use L-malic acid. D-malic acid is not only unusable by the human body, but also increases the metabolic burden of the organism. Chemically synthesized malic acid is an equal mixture of L- and D-types, so only half can be used by the human body. In the production of fruit products such as fruit juice and fruit juice drinks, adulteration occurs due to profit-driven reasons, such as adding artificially synthesized malic acid to apple juice to impersonate natural juice.

[0004] At present, the mainstream method for distinguishing real and fake fruit juices at home and abroad is to measure the δ13C total carbon of the stable carbon isotope of the juice and compare it with the known true δ13C total carbon range. After years of data accumulation and research, the European Fruit and Vegetable Juice Association has given the δ13C total carbon determination range for more than 20 kinds of fruit and vegetable juices, including the δ13C total carbon of apple juice, pear juice, grape juice, etc.

[0005] However, in actual applications, misjudgment may occur due to differences in origin and variety, especially the addition of carbon-3 plant syrup, which cannot be identified. Therefore, it is urgent and important to establish an identification technology with high sensitivity, short detection cycle and wide adaptability. Summary of the invention

[0006] In order to solve the above technical problems, the present invention provides a fruit juice authenticity identification model and an adulteration ratio determination method thereof. By determining the contents of two enantiomers of D- and L-malic acid in the fruit juice, the authenticity of the fruit juice and the adulteration ratio can be accurately determined to solve the problems in the prior art.

[0007] A fruit juice authenticity identification model and a method for determining the adulteration ratio thereof, comprising the following steps:

[0008] S1, sample preparation, two samples of juice with pulp and juice without pulp are used as test specimens after pre-treatment;

[0009] S2, analysis and determination, placing a uniform sample in a plastic centrifuge tube, adding an appropriate amount of water for ultrasonic extraction and centrifugation, obtaining the supernatant and fixing the volume to obtain the sample extract, then pre-rinsing and activating the solid phase extraction column, adding the sample extract to the column, discarding part of the initial filtrate and collecting the filtrate, and finally placing the collected filtrate under specific high performance liquid chromatography conditions to complete the analysis and determination of the sample;

[0010] S3, result calculation, calculating the content of L-malic acid and D-malic acid in the sample;

[0011] S4, authenticity identification and adulteration ratio determination, based on the fact that apple, pear and orange juice all contain L-malic acid but no D-malic acid, the authenticity identification and adulteration ratio determination method of apple, pear and orange juice is as follows: determine the content of L-malic acid and D-malic acid in the juice, and determine the authenticity and adulteration ratio of the product according to the formula [1-(1-CL / CT)x2]x100.

[0012] Preferably, in S1, the specific preparation process of the two samples of juice with pulp and juice without pulp is as follows:

[0013] Juice with pulp: Use a homogenizer to make a homogenate, put it into a clean container, seal it, and store it in a freezer below -18℃;

[0014] Juice without pulp: Shake well, put into clean container, seal, and store in freezer below -18℃.

[0015] Preferably, in S2, the specific process of the analysis and determination is as follows:

[0016] Weigh 5 g of uniform sample into a 50 mL plastic centrifuge tube, add 20 mL of water, shake well, perform ultrasonic extraction at room temperature for 20 min at 40 KHz frequency, centrifuge at 9000 r / min for 15 min, and transfer the supernatant to a 50 mL volumetric flask;

[0017] The residue was extracted again with 10 mL of water according to the above steps, the two extracts were combined, the volume was made up to the mark with water, and the sample extract was obtained by shaking well;

[0018] Pre-rinse and activate the SPE cartridge with 5 mL of methanol and 5 mL of water in sequence;

[0019] 5 mL of sample extract was added to the activated solid phase extraction column, 2 mL of initial filtrate was discarded, and the filtrate was collected for analysis by high performance liquid chromatography.

[0020] Preferably, in the liquid chromatograph analysis, the reference conditions in the liquid chromatograph include:

[0021] Chromatographic column: C18 column, 250mm×4.6mm×5μm, or equivalent column;

[0022] Column temperature: 30°C; flow rate: 1 mL / min; injection volume: 10 μL; detection wavelength: 302 nm;

[0023] Mobile phase: 16 mmol / L L-valine and 8 mmol / L copper acetate aqueous solution;

[0024] Mobile phase elution: isocratic elution for 30 min;

[0025] Among them, the mobile phase refers to the liquid that carries the sample through the chromatographic column during the chromatography process, and the mobile phase elution refers to the process of flushing the chromatographic column with the mobile phase so that different components in the sample can be separated in the chromatographic column based on the differences in their interactions with the stationary phase and the mobile phase;

[0026] Isocratic elution means that the composition of the mobile phase remains unchanged throughout the analysis process, and the chromatographic column is continuously flushed at a constant flow rate, so that the various components in the sample are distributed and separated between the stationary phase and the mobile phase, and thus detected at different times.

[0027] Preferably, in S3, the contents of L-malic acid and D-malic acid in the sample are calculated according to the following formula:

[0028]

[0029] Wherein, ω means the content of L-malic acid or D-malic acid in the sample, in grams per kilogram, C means the concentration of L-malic acid or D-malic acid in the sample solution, in milligrams per milliliter; C0 means the concentration of L-malic acid or D-malic acid in the blank solution, in milligrams per milliliter; V means the volume of the extract, in milliliters; f means the dilution factor of the sample solution; and m means the mass of the sample, in grams.

[0030] In S4, the authenticity and adulteration ratio of the product are determined based on the CL / CT value. The specific ratio results are as follows:

[0031] Assuming CL / CT = 1, it is pure natural juice, without adding artificial malic acid;

[0032] CL / CT = 0.95, containing 90% natural juice and 10% artificial malic acid;

[0033] CL / CT = 0.85, containing 70% natural juice and 30% artificial malic acid;

[0034] CL / CT = 0.75, containing 50% natural juice and 50% artificial malic acid;

[0035] CL / CT=0.5, contains 0% natural fruit juice and 100% artificial malic acid is added.

[0036] Preferably, in S2, the related devices used in the analysis and determination include the equipment used in the above analysis: 50mL plastic centrifuge tube, ultrasonic instrument, centrifuge, 50mL volumetric flask, solid phase extraction column and high performance liquid chromatography.

[0037] Compared with the prior art, the present invention has the following beneficial effects:

[0038] 1. The present invention realizes the precise determination of L-malic acid and D-malic acid in juice through perfect juice sample preparation and analysis and determination method, and accurately identifies the authenticity and adulteration ratio of juice.

[0039] 2. The present invention overcomes the problem of misjudgment caused by different origins and varieties and the inability to identify the addition of carbon-3 plant syrup through precise detection procedures and scientific calculation methods, and realizes an apple juice identification technology with high sensitivity, short detection cycle and wide adaptability. The method is simple, practical and accurate. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] Figure 1 It is a schematic flow chart of the method of the present invention;

[0041] Figure 2 It is a schematic flow chart of the analysis and determination steps of the present invention. DETAILED DESCRIPTION

[0042] The following embodiments of the present invention are described in further detail in conjunction with the accompanying drawings and examples. The following examples are used to illustrate the present invention, but are not intended to limit the scope of the present invention.

[0043] The present invention provides a fruit juice authenticity identification model and a method for determining the adulteration ratio thereof, comprising the following steps:

[0044] S1, sample preparation, two samples of juice with pulp and juice without pulp are used as test specimens after pre-treatment;

[0045] S2, analysis and determination, placing a uniform sample in a plastic centrifuge tube, adding an appropriate amount of water for ultrasonic extraction and centrifugation, obtaining the supernatant and fixing the volume to obtain the sample extract, then pre-rinsing and activating the solid phase extraction column, adding the sample extract to the column, discarding part of the initial filtrate and collecting the filtrate, and finally placing the collected filtrate under specific high performance liquid chromatography conditions to complete the analysis and determination of the sample;

[0046] S3, result calculation, calculating the content of L-malic acid and D-malic acid in the sample;

[0047] S4, authenticity identification and adulteration ratio determination, based on the fact that apple, pear and orange juice all contain L-malic acid but no D-malic acid, the authenticity identification and adulteration ratio determination method of apple, pear and orange juice is as follows: determine the content of L-malic acid and D-malic acid in the juice, and determine the authenticity and adulteration ratio of the product according to the formula [1-(1-CL / CT)x2]x100.

[0048] Example:

[0049] like Figure 1-2 As shown, in this embodiment, apple juice is used as an example to identify the authenticity and the proportion of adulteration;

[0050] First, sample collection was carried out. Multiple apple juice samples were collected from different channels such as supermarkets and direct purchase from orchards, including products from different brands and origins, to ensure the representativeness and universality of the experiment.

[0051] The collected samples shall be numbered and recorded, indicating the source, production date and other information.

[0052] Instrument testing and preparation:

[0053] Check and calibrate the ultrasonic instrument to ensure that its frequency is stable at 40KHz. Check the centrifuge, set the speed to 9000r / min, and conduct a test run to ensure it works properly.

[0054] Prepare 50mL plastic centrifuge tubes, 50mL volumetric flasks, solid phase extraction columns and other experimental equipment, and make sure they are clean and dry.

[0055] The mobile phase required for the high performance liquid chromatography was prepared by accurately weighing a certain amount of L-valine and cupric acetate, dissolving them in water, and preparing a 16 mmol / L L-valine and 8 mmol / L cupric acetate aqueous solution.

[0056] The prepared mobile phase was filtered through a 0.45 μm microporous filter membrane and degassed to remove bubbles therein.

[0057] Debug and calibrate the HPLC, set the chromatographic column, C18 column, 250mm×4.6mm×5μm or equivalent column; the temperature is 30℃, the flow rate is 1mL / min, the injection volume is 10μL, the detection wavelength is 302nm, and perform a blank run to ensure that the instrument baseline is stable and there are no interfering peaks;

[0058] S1, Sample preparation:

[0059] Juice with pulp: Use a homogenizer to make a homogenate, put it into a clean container, seal it, and store it in a freezer below -18℃;

[0060] Juice without pulp: Shake well, put into clean container, seal, and store in freezer below -18℃.

[0061] S2, analysis and determination:

[0062] Weigh 5 g of uniform sample into a 50 mL plastic centrifuge tube, add 20 mL of water, shake well, and perform ultrasonic extraction at 40 kHz for 20 min at room temperature. Then centrifuge at 9000 r / min for 15 min, and transfer the supernatant to a 50 mL volumetric flask.

[0063] The residue was extracted once again with 10 mL of water according to the above ultrasonic extraction and centrifugation steps, the two extracts were combined, the volume was made up to 50 mL with water, and shaken to obtain the sample extract.

[0064] Pre-rinse and activate the solid phase extraction column with 5 mL of methanol and 5 mL of water in turn. Pipette 5 mL of sample extract and add it to the activated solid phase extraction column, discard 2 mL of initial filtrate, and collect the filtrate. Inject the collected filtrate into a high performance liquid chromatograph for analysis and determination. The analysis and determination conditions of the high performance liquid chromatograph are as follows: use a C18 chromatographic column, 250 mm × 4.6 mm × 5 μm or an equivalent chromatographic column, set the column temperature to 30 ° C, use 16 mmol / L L-valine and 8 mmol / L copper acetate aqueous solution as the mobile phase, the flow rate is 1 mL / min, the injection volume is 10 μ L, the detection wavelength is 302 nm, and the mobile phase elution mode is isocratic elution for 30 min.

[0065] S4. Result calculation:

[0066] Calculation of fruit juice with pulp:

[0067] The concentration of L-malic acid in the pulp juice sample solution was determined by high performance liquid chromatography to be 120 mg / mL, the concentration of L-malic acid in the blank solution was 5 mg / mL, the volume of the extract was 50 mL, the dilution factor of the sample solution was 2, and the mass of the sample was 5 g.

[0068] According to the formula:

[0069]

[0070] ω(L-malic acid) = [(120-5) × 50 × 2] / 5000 = 2.3 g / kg

[0071] Similarly, the concentration of D-malic acid in the pulp juice sample solution was determined to be 0 mg / mL, and no D-malic acid was detected, so the total malic acid content in the pulp juice CT1 = 2.3 g / kg.

[0072] Calculation of results for juice without pulp:

[0073] The concentration of L-malic acid in the commercially available apple juice sample solution without pulp was determined by high performance liquid chromatography to be 80 mg / mL, the concentration of L-malic acid in the blank solution was 3 mg / mL, the volume of the extract was 50 mL, the dilution factor of the sample solution was 2, and the mass of the sample was 5 g.

[0074] According to the formula:

[0075]

[0076] ω(L-malic acid) = [(80-3) × 50 × 2] / 5000 = 1.54 g / kg

[0077] The concentration of D-malic acid in the commercially available apple juice sample solution without pulp was determined to be 10 mg / mL. According to the above formula, it can be calculated as follows:

[0078] ω(D-malic acid) = [(10-0) × 50 × 2] / 5000 = 0.2 g / kg

[0079] The total malic acid content in commercially available apple juice without pulp is CT2 = 1.54 g / kg + 0.2 g / kg = 1.74 g / kg.

[0080] Through perfect apple juice sample preparation and analytical determination methods, the precise determination of L-malic acid and D-malic acid in apple juice is achieved, and the authenticity and adulteration ratio of apple juice are accurately identified.

[0081] S5. Authenticity identification and adulteration ratio determination:

[0082] Identification of authenticity of fruit juice with pulp:

[0083] The CL / CT value of the pulped juice was calculated, where CL1 was the L-malic acid content in the pulped juice and CT was the total malic acid content in the pulped juice, i.e., the sum of the L- and D-malic acid contents.

[0084] CL=2.3g / kg,CT=2.3g / kg

[0085] CL / CT=2.3 / 2.3=1

[0086] Substituting CL / CT=1 into the formula [1-(1-CL / CT)x2]x100, the result is 100%. It can be judged that the pulp juice is pure natural juice without adding artificial malic acid.

[0087] How to identify the authenticity of fruit juice without pulp:

[0088] The CL / CT value of commercially available pulpless apple juice was calculated, where CL is the L-malic acid content in the pulpless juice and CT is the total malic acid content in the pulpless juice, i.e., the sum of the L- and D-malic acid contents.

[0089] CL=1.54g / kg, CT2=1.74g / kg

[0090] CL / CT2=1.54 / 1.74≈0.885

[0091] The purity of juice is (in %): [1-(1-CL / CT)x2]x100. If CL / CT2=0.885, the result is [1-(1-0.885)x2]x100=77%;

[0092] From this, it can be judged that the commercially available apple juice without pulp contains about 77% natural juice.

[0093] Through precise detection procedures and scientific calculation methods, we have overcome the problems of misjudgment due to different origins and varieties and the inability to identify the addition of carbon-3 plant syrup, and achieved a high-sensitivity, short detection cycle, and wide-adaptability apple juice identification technology. The method is simple, practical, and accurate.

[0094] The embodiments of the present invention are provided for the purpose of illustration and description. Although the embodiments of the present invention have been shown and described above, it can be understood that the above embodiments are exemplary and cannot be understood as limitations of the present invention. Ordinary technicians in this field can change, modify, replace and modify the above embodiments within the scope of the present invention.

Claims

1. A fruit juice authenticity identification model and a method for determining the adulteration ratio thereof, characterized in that: include: S1, sample preparation, two samples of juice with pulp and juice without pulp were selected as test samples for pre-treatment and standby use; S2, analysis and determination, placing a uniform sample in a plastic centrifuge tube, adding an appropriate amount of water for ultrasonic extraction and centrifugation, obtaining the supernatant and fixing the volume to obtain the sample extract, then pre-rinsing and activating the solid phase extraction column, adding the sample extract to the column, discarding part of the initial filtrate and collecting the filtrate, and finally placing the collected filtrate under specific high performance liquid chromatography conditions to complete the analysis and determination of the sample; S3, result calculation, calculating the content of L-malic acid and D-malic acid in the sample; S4, authenticity identification and adulteration ratio determination. Through the determination of fresh apples, pears and oranges of different varieties and origins, it was found that fresh apples, pears and oranges all contain L-malic acid but no D-malic acid. On this basis, the authenticity identification and adulteration ratio determination method of apple, pear and orange juice is obtained as follows: determine the content of L-malic acid and D-malic acid in the juice, and determine the authenticity and adulteration ratio of the product according to the formula [1-(1-CL / CT)x2]x100, where CT=CL+CD.

2. A fruit juice authenticity identification model and method for determining the adulteration ratio thereof as claimed in claim 1, characterized in that: In S1, the specific preparation process of the two samples of juice with pulp and juice without pulp is as follows: Juice with pulp: Use a homogenizer to make a homogenate, put it into a clean container, seal it, and store it in a freezer below -18℃; Juice without pulp: Shake well, put into clean container, seal, and store in freezer below -18℃.

3. A fruit juice authenticity identification model and method for determining the adulteration ratio thereof as claimed in claim 1, characterized in that: In S2, the specific process of the analysis and determination is as follows: Weigh 5 g of uniform sample into a 50 mL plastic centrifuge tube, add 20 mL of water, shake well, perform ultrasonic extraction at room temperature for 20 min at 40 KHz frequency, centrifuge at 9000 r / min for 15 min, and transfer the supernatant to a 50 mL volumetric flask; The residue was extracted again with 10 mL of water according to the above steps, the two extracts were combined, the volume was made up to the mark with water, and the sample extract was obtained by shaking well; Pre-rinse and activate the SPE cartridge with 5 mL of methanol and 5 mL of water in sequence; 5 mL of sample extract was added to the activated solid phase extraction column, 2 mL of initial filtrate was discarded, and the filtrate was collected for liquid chromatography analysis.

4. A fruit juice authenticity identification model and method for determining the adulteration ratio thereof as claimed in claim 3, characterized in that: In the liquid chromatograph analysis and determination, the reference conditions in the liquid chromatograph include: Chromatographic column: C18 column, 250mm×4.6mm×5μm, or equivalent column; Column temperature: 30°C; flow rate: 1 mL / min; injection volume: 10 μL; detection wavelength: 302 nm; Mobile phase: 16mmol / L L-valine and 8mmol / L copper acetate aqueous solution; Mobile phase elution: isocratic elution for 30 min; Among them, the mobile phase refers to the liquid that carries the sample through the chromatographic column during the chromatography process, and the mobile phase elution refers to the process of flushing the chromatographic column with the mobile phase so that different components in the sample can be separated in the chromatographic column based on the differences in their interactions with the stationary phase and the mobile phase; Isocratic elution means that the composition of the mobile phase remains unchanged throughout the analysis process, and the chromatographic column is continuously flushed at a constant flow rate, so that the various components in the sample are distributed and separated between the stationary phase and the mobile phase, and thus detected at different times.

5. A fruit juice authenticity identification model and method for determining the adulteration ratio thereof as claimed in claim 1, characterized in that: In S3, the contents of L-malic acid and D-malic acid in the sample are calculated according to the following formula: Wherein, ω means the content of L-malic acid or D-malic acid in the sample, in grams per kilogram, C means the concentration of L-malic acid or D-malic acid in the sample solution, in milligrams per milliliter; C0 means the concentration of L-malic acid or D-malic acid in the blank solution, in milligrams per milliliter; V means the volume of the extract, in milliliters; f means the dilution factor of the sample solution; and m means the mass of the sample, in grams.

6. A fruit juice authenticity identification model and method for determining the adulteration ratio thereof as claimed in claim 1, characterized in that: In S2, the related devices used in the analysis and determination include: 50mL plastic centrifuge tube, ultrasonic instrument, centrifuge, 50mL volumetric flask, solid phase extraction column and high performance liquid chromatography.

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