Kit and method for detecting content of vitamin C in fruits and vegetables

Abstract

The invention relates to the field of fruit and vegetable detection, in particular to a kit for detecting the content of vitamin C in fruits and vegetables and a detection method, the kit comprises a kit body and reagents, the kit body comprises a sample treatment tube and a steel ball placed in the sample treatment tube, and the reagents comprise a standard substance, an extracting solution, a color developing agent and a decolorizing agent. The standard substance is vitamin C dry powder, the extracting solution is an acid solution, and the color developing agent is one of sodium bicarbonate, 2, 6-dichloroindophenol or a 2, 6-dichloroindophenol sodium salt aqueous solution. The detection method disclosed by the invention is simple to operate, rapid in detection and high in accuracy, the used kit is low in cost, and the requirement on the technical level of an operator is not high; the linear range of the detection method is wide, most fruit and vegetable samples can be detected without dilution, and the final vitamin C content is obtained; the method is low in sample consumption in detection, and is particularly suitable for fruit and vegetable samples which are low in sample quantity or difficult to obtain.

Description

Technical Field

[0001] The present invention relates to the field of fruit and vegetable detection, and specifically to a kit and a detection method for detecting the vitamin C content in fruits and vegetables. Background Art

[0002] The content of vitamin C is one of the important indicators for evaluating the quality of fruits and vegetables. According to the national food safety standard GB5009.86 - 2016, currently, there are mainly three methods for detecting the content of vitamin C. One is high - performance liquid chromatography. Although it has high sensitivity, the instrument equipment is expensive, the maintenance cost is high, it is necessary to prepare the mobile phase, equilibrate the chromatographic column, etc. The whole detection process takes a long time and requires a high technical level of the operator. Another is the fluorescence method, which includes multiple steps such as activated carbon activation and is relatively cumbersome. The other is the 2,6 - dichlorophenol indophenol titration method. Although this method is simple to operate and has low cost, it is difficult to judge the titration end point, has low sensitivity and large error. Therefore, how to simply, quickly, accurately and low - cost detect the vitamin C content in fruits and vegetables is an urgent technical problem to be solved currently. Summary of the Invention

[0003] To achieve the above object, the object of the present invention is to provide a kit for detecting the vitamin C content in fruits and vegetables, which can simply, quickly and accurately detect the vitamin C content in fruits and vegetables, and the cost of this kit is low. The present invention provides the following technical solutions: A kit for detecting the vitamin C content in fruits and vegetables includes a kit body and reagents. The kit body includes a sample treatment tube and steel beads placed in the sample treatment tube. The reagents include a standard product, an extraction solution, a color - developing agent and a decolorizing agent. The standard product is vitamin C dry powder, the extraction solution is an acid solution for extracting and stabilizing vitamin C in fruits and vegetables, and the color - developing agent is any one of a mixed solution of sodium bicarbonate and 2,6 - dichlorophenol indophenol or a mixed solution of an aqueous solution of sodium bicarbonate and 2,6 - dichlorophenol indophenol sodium.

[0004] As a further scheme of the present invention: the sample treatment tube is a 1.5 mL or 2 mL centrifuge tube.

[0005] As a further scheme of the present invention: the acid solution is an oxalic acid solution with a mass fraction of 2% or a metaphosphoric acid solution with a mass fraction of 2 - 4%, and it can also be other acidic solutions or composite acidic solutions.

[0006] As a further scheme of the present invention: the steel beads are stainless steel beads with a diameter of 2 mm - 5 mm.

[0007] As a further scheme of the present invention: the decolorizing agent is kaolin powder for removing pigments in the fruit and vegetable extraction solution to reduce interference.

[0008] As a further solution of the present invention: the concentration of sodium bicarbonate in the developer is 10.4 - 20.8 mg / L, the concentration of 2,6-dichloroindophenol or its sodium salt aqueous solution is 10 - 22 mg / L, and the concentration ratio of sodium bicarbonate to 2,6-dichloroindophenol or sodium bicarbonate to its sodium salt is 1:1 - 1.1:1.

[0009] A method for detecting the vitamin C content in fruits and vegetables, comprising the following steps: (1) Based on the system spectrophotometry, dissolve and dilute the vitamin C dry powder with the extraction solution to prepare vitamin C standard solutions with different gradients. Add the vitamin C standard solutions with different gradients into the cuvettes respectively, and then add the developer into the cuvettes so that the total volume of the reaction solution in the cuvettes is 1 mL. Immediately mix well and put it into the spectrophotometer to detect the absorbance at 515 nm. Take the vitamin C concentration as the abscissa and the absorbance as the ordinate to draw a standard curve and obtain the linear regression equation; (2) Take a small amount of fruit and vegetable samples into the sample treatment tube, add the extraction solution and the decolorizing agent, and then homogenize. After homogenization, put the sample treatment tube into the centrifuge and centrifuge at 2 - 6 °C to obtain a basically colorless supernatant as the test solution, and place it on ice for testing; (3) Measure the absorbance of the test solution by the same measurement method as in step (1), and calculate the vitamin C content in the fruits and vegetables according to the linear regression equation obtained in step (1).

[0010] As a further solution of the present invention: use a cell crusher for homogenization during homogenization.

[0011] As a further solution of the present invention: the volume of the vitamin C standard solution in step (1) and the test solution in step (3) is 50 - 70 μL, and the volume of the developer in steps (1) and (3) is 930 - 950 μL.

[0012] As a further solution of the present invention: in steps (1) and (3), start timing from the addition of the developer, and mixing and detection need to be completed within 5 s.

[0013] As a further solution of the present invention: in step (2), the mass of the fruit and vegetable sample is 0.05 g - 1 g, the mass of the decolorizing agent is 2 mg - 200 mg, the homogenization frequency is 15 - 30 times / s, the homogenization time is 0.5 - 5 min, the centrifugal force is 8000 - 10000 g, the centrifugation time is 5 - 10 min, and the ratio of the test solution to the fruit and vegetable sample is 20 mL:1 g - 1 mL:1 g.

[0014] Compared with the prior art, the beneficial effects of the present invention are: In the kit adopted by the present invention, a cell crusher is used for homogenate extraction. Compared with manual grinding, the extraction is more sufficient and faster, which can effectively reduce the oxidation of vitamin C. At the same time, the extraction of dozens of samples can be carried out, saving time and effort. The detection method of the present invention is simple to operate, fast in detection, high in accuracy, the kit used has a low cost, and the technical level requirements for operators are not high. The detection method of the present invention has a wide linear range (4 - 75 μg / mL). Except for fruits and vegetables with particularly high vitamin C content, most fruit and vegetable samples can be detected without dilution, and the final vitamin C content can be obtained. The sample consumption in the detection of the present invention is small, which is especially suitable for fruit and vegetable samples with small sample amounts or difficult to obtain. Description of the Drawings

[0015] Figure 1 It is the linear data graph of Example 1 of the present invention.

[0016] Figure 2 It is the linear data graph of Example 2 and Example 4 of the present invention.

[0017] Figure 3 It is the linear data graph of Example 3 of the present invention. Detailed Embodiments

[0018] Next, the technical solutions in the embodiments of the present invention will be clearly and completely described in conjunction with the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without making creative efforts belong to the scope of protection of the present invention.

[0019] The detection principle of the present invention is as follows: 2,6-dichloroindophenol is blue in an alkaline solution. The excessive blue alkaline dye 2,6-dichloroindophenol undergoes an oxidation-reduction reaction with vitamin C in the acidic extract. Part of the 2,6-dichloroindophenol is reduced to colorless, and the excess 2,6-dichloroindophenol is light red in an acidic medium. The light red solution has a maximum absorption at 515 nm, and the absorbance of the light red solution is linearly related to the concentration of vitamin C. Therefore, the content of vitamin C in fruit and vegetable samples can be obtained by spectrophotometry.

[0020] The following is a detailed description of the specific implementation of the present invention in conjunction with specific embodiments.

[0021] Example 1:

[0022] A kit for detecting the vitamin C content in fruits and vegetables, including a kit body and reagents; the kit body includes a sample treatment tube, and the reagents include a standard product, an extract, a color developer, and a decolorizer.

[0023] The sample treatment tube is a 1.5 m centrifuge tube and contains 2 stainless steel balls with a diameter of 4 mm.

[0024] The standard product is vitamin C dry powder and is dissolved before use.

[0025] The extraction solution is an oxalic acid solution with a mass fraction of 2%.

[0026] The color developing agent contains 20.8 mg / L sodium bicarbonate and 22 mg / L aqueous solution of sodium 2,6-dichloroindophenolate.

[0027] The decolorizing agent is kaolin powder and is used to remove pigments in the fruit and vegetable extraction solution to reduce interference.

[0028] The detection method of this embodiment includes the following steps: a) Dissolve and dilute the vitamin C standard product with the extraction solution in the reagent kit to obtain a gradient vitamin C standard solution. Respectively take 70 μL of the vitamin C standard solution and add it into a 1 mL glass colorimetric cell. Then add 930 μL of the color developing agent into the colorimetric cell so that the total volume of the reaction solution in the colorimetric cell is 1 mL. Immediately mix well within 5 s and put it into a spectrophotometer to detect the absorbance at 515 nm. Finally, take the vitamin C concentration as the abscissa and the absorbance as the ordinate to draw a standard curve and obtain a linear regression equation.

[0029] b) Take 0.1 g of fresh kiwifruit pulp into the sample treatment tube, add 1 mL of the extraction solution and 3 mg of kaolin, and then use a cell disruptor to homogenize at a frequency of 30 times / s for 1 min. After homogenization, place the sample treatment tube in a centrifuge and centrifuge at a centrifugal force of 10000 g at 4 °C for 10 min to obtain a supernatant. Take the supernatant as the test solution and place it on ice for testing.

[0030] Step c) Dilute the kiwifruit test solution 5 times with the extraction solution. Take 70 μL of the diluted solution and add it into a 1 mL glass colorimetric cell. Then add 930 μL of the color developing agent into the colorimetric cell so that the total volume of the reaction solution in the colorimetric cell is 1 mL. Immediately mix well within 5 s and put it into a spectrophotometer to detect the absorbance at 515 nm. Calculate the vitamin C content in the sample diluted solution through the linear regression equation, and then calculate the vitamin C content in the kiwifruit sample according to the dilution factor.

[0031] Example 2:

[0032] A reagent kit for detecting the vitamin C content in fruits and vegetables, including a reagent kit body and reagents; the reagent kit body includes a sample treatment tube, and the reagents include a standard product, an extraction solution, a color developing agent, and a decolorizing agent.

[0033] The sample treatment tube is a 1.5 m centrifuge tube and contains 2 stainless steel balls with a diameter of 4 mm.

[0034] The reference standard is dry vitamin C powder, which is dissolved before use.

[0035] The extraction solution is an oxalic acid solution with a mass fraction of 2%.

[0036] The color-developing agent contains 10.4 mg / L sodium bicarbonate and 11 mg / L aqueous solution of sodium 2,6-dichloroindophenol.

[0037] The decolorizing agent is kaolin powder, which is used to remove pigments in the fruit and vegetable extraction solution to reduce interference.

[0038] The detection method of this example includes the following steps: a) Dissolve and dilute the vitamin C reference standard with the extraction solution in the kit to obtain a gradient of vitamin C standard solutions. Respectively take 70 μL of the vitamin C standard solution and add it to a 1 mL glass cuvette, then add 930 μL of the color-developing agent to the cuvette so that the total volume of the reaction solution in the cuvette is 1 mL. Immediately mix well within 5 s and put it into a spectrophotometer to detect the absorbance at 515 nm. Finally, use the vitamin C concentration as the abscissa and the absorbance as the ordinate to plot the standard curve and obtain the linear regression equation.

[0039] b) Take 0.2 g of fresh calcium-rich vegetables into a sample treatment tube, add 1 mL of the extraction solution and 40 mg of kaolin, and then homogenize with a cell disruptor at a frequency of 30 times / s for 3 min. After homogenization, place the sample treatment tube in a centrifuge and centrifuge at 10000 g at 4 °C for 10 min to obtain the supernatant. Take the supernatant as the test solution and place it on ice for testing.

[0040] For step c), directly take 70 μL of the test solution and add it to a 1 mL glass cuvette, then add 930 μL of the color-developing agent to the cuvette so that the total volume of the reaction solution in the cuvette is 1 mL. Immediately mix well within 5 s and put it into a spectrophotometer to detect the absorbance at 515 nm. Calculate the vitamin C content in the sample dilution solution through the linear regression equation, and then calculate the vitamin C content in the calcium-rich vegetable sample according to the dilution factor.

[0041] Example 3:

[0042] This example is based on Example 2, the difference is that the volumes of the standard solution or the test solution and the color-developing agent in steps a) and c) are different. Specifically: The volume of the standard solution or the test solution in steps a) and c) is 50 μL; The volume of the color-developing agent in steps a) and c) is 950 μL.

[0043] Example 4:

[0044] This example is based on Example 2, with the difference being that the fruit and vegetable samples and extraction methods in step b) are different. Specifically: Take 0.3 g of ponkan pulp into a sample treatment tube, add 1 mL of extraction solution and 60 mg of kaolin, and then homogenize with a cell crusher at a frequency of 30 times / s for 2 min.

[0045] The linear data of Examples 1-4 are shown in Table 1.

[0046]

[0047] In the linear regression equation in Table 1, Y is the absorbance at 515 nm, and X is the vitamin C concentration (μg / mL).

[0048] The vitamin C content in the diluted test solution is calculated according to the linear regression equation, and finally the total vitamin C content (mg / 100 g) in the fruit and vegetable sample is calculated. The calculation formula is as follows: Total vitamin C content (mg / 100 g) = V ◊ X ◊ D / (10 ◊ M) Where: V is the volume (mL) of the added extraction solution; X is the vitamin C concentration (μg / mL) in the diluted test solution calculated according to the linear regression equation; D is the dilution factor of the test solution; M is the mass (g) of the sample. Figures 1 - 3 For the linear data of Examples 1-4, from Figures 1 - 3 It can be seen that the linear relationship of the vitamin C content measured by the kit of the present invention is good, and the linear range is 4-75 μg / mL.

[0049] The repeatability data are shown in Table 2.

[0050]

[0051] It can be seen from Table 2 that the detection method of this kit has good repeatability, and the relative standard deviations of the four examples are all less than 4%.

[0052] The spike recovery data are shown in Tables 3-5.

[0053]

[0054]

[0055]

[0056] It can be seen from Tables 3-5 that the recovery rate of the samples measured by the present invention is between 95%-105%, indicating that the detection accuracy of this method is high.

[0057] Comparative Example 1 Same as Example 1, except that the concentration of sodium bicarbonate in the developer is 104 mg / L and the concentration of sodium 2,6-dichloroindophenolate is 110 mg / L. The absorbance of the reaction solution was measured to be greater than 2. Since the absorbance is too high, normal detection cannot be carried out.

[0058] Comparative Example 2 Same as Example 2, except that the volume of the standard solution in step a) and the sample diluent in step c) is 100 μL, and the volume of the developer in steps a) and c) is 900 μL. The finally obtained linear range is 4 - 50 μg / mL, and the linear range is narrowed compared with Example 2.

[0059] Comparative Example 3 Same as Example 4, except that the concentration of sodium bicarbonate in the developer is 69.3 mg / L and the concentration of sodium 2,6-dichloroindophenolate is 73.3 mg / L. The spiked recovery rate of the sample was measured to be 66.85%. It is lower than the recovery rate obtained from the sample in Example 4, indicating that the concentrations of sodium bicarbonate and sodium 2,6-dichloroindophenolate in the developer have an important impact on the detection results.

[0060] Comparative Example 4 Same as Example 4, except that kaolin is not added for decolorization in step b). The finally measured vitamin C content in ponkan oranges is 25% higher than that in Example 4, indicating that the pigments in the sample interfere with the experimental results.

[0061] Comparative Example 5 Same as Example 1, except that in steps a) and c), after adding the developer, it is immediately mixed evenly, and the absorbance is measured after standing for 30 minutes. As a result, the color of the reaction solution becomes significantly lighter due to oxidation in the air, and the measured result is 5 times that of Example 1, indicating that the detection time has an important impact on the detection results.

[0062] It should be noted that in the present invention, unless otherwise clearly specified and limited, terms such as "fix" and "set" should be understood in a broad sense. For example, it can be a welded connection, a bolt connection, or integrated; it can be a mechanical connection or an electrical connection; it can be directly connected or indirectly connected through an intermediate medium, and it can be the internal connection of two components or the interaction relationship between two components, unless otherwise clearly limited. For those of ordinary skill in the art, the specific meanings of the above terms in the present invention can be understood according to specific circumstances.

[0063] In addition, it should be understood that although this specification is described in terms of embodiments, not every embodiment contains only an independent technical solution. This narrative style of the specification is only for clarity. Those skilled in the art should regard the specification as a whole, and the technical solutions in each embodiment can also be appropriately combined to form other embodiments that can be understood by those skilled in the art.

Claims

1. A kit for detecting the vitamin C content in fruits and vegetables, characterized in that, It includes a kit body and reagents. The kit body includes a sample treatment tube and steel beads placed in the sample treatment tube. The reagents include a standard, an extraction solution, a chromogenic agent, and a decolorizing agent. The standard is vitamin C dry powder, the extraction solution is an acid solution, and the chromogenic agent is any one of sodium bicarbonate, 2,6-dichlorophenol indophenol, or an aqueous solution of sodium 2,6-dichlorophenol indophenol.

2. The kit for detecting the vitamin C content in fruits and vegetables according to claim 1, characterized in that, The acid solution is an oxalic acid solution with a mass fraction of 2% or a metaphosphoric acid solution with a mass fraction of 2-4%.

3. The kit for detecting the vitamin C content in fruits and vegetables according to claim 1 or 2, characterized in that, The decolorizing agent is kaolin powder.

4. The kit for detecting the vitamin C content in fruits and vegetables according to claim 1, wherein In the chromogenic agent, the concentration of sodium bicarbonate is 10.4-20.8 mg / L, and the concentration of 2,6-dichlorophenol indophenol or an aqueous solution of sodium 2,6-dichlorophenol indophenol is 10-22 mg / L.

5. A method for detecting the vitamin C content in fruits and vegetables, characterized in that, Using the kit for detecting the vitamin C content in fruits and vegetables according to any one of claims 1-4, it includes the following steps: (1) Based on the system spectrophotometry, dissolve and dilute the vitamin C dry powder with the extraction solution to prepare vitamin C standard solutions with different gradients. Add the vitamin C standard solutions with different gradients into a cuvette respectively, and then add the chromogenic agent into the cuvette so that the total volume of the reaction solution in the cuvette is 1 mL. Immediately mix well and put it into a spectrophotometer to detect the absorbance at 515 nm. Take the vitamin C concentration as the abscissa and the absorbance as the ordinate to draw a standard curve and obtain a linear regression equation. (2) Take a small amount of fruit and vegetable sample into the sample treatment tube, add the extraction solution and the decolorizing agent, and then homogenize. After homogenization, put the sample treatment tube into a centrifuge and centrifuge at 2-6 °C to obtain a basically colorless supernatant as the test solution, and place it on ice for testing. (3) Use the same measurement method as in step (1) to measure the absorbance of the test solution, and calculate the vitamin C content in the fruits and vegetables according to the linear regression equation obtained in step (1).

6. The detection method for the vitamin C content in fruits and vegetables according to claim 5, wherein In step (1), the volume of the vitamin C standard solution and in step (3), the volume of the test solution is 50-70 μL, and in step (1) and step (3), the volume of the chromogenic agent is 930-950 μL.

7. The method for detecting the vitamin C content in fruits and vegetables according to claim 5 or 6, characterized in that, In step (2), the mass of the fruit and vegetable sample is 0.05 g-1 g, the mass of the decolorizing agent is 2 mg-200 mg, the homogenization frequency is 15-30 times / s, the homogenization time is 0.5-5 min, the centrifugal force is 8000-10000 g, the centrifugation time is 5-10 min, and the ratio of the test solution to the fruit and vegetable sample is 20 mL:1 g-1 mL:1 g.

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