CN112304886B - Rapid identification method for squeezed and extracted camellia seed oil - Google Patents
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Abstract
The invention relates to a rapid identification method for squeezed and extracted camellia seed oil, and belongs to the field of detection of edible vegetable oil in food detection. At present, a simple and quick method for quickly identifying oil-tea camellia seed oil by squeezing and leaching does not exist. The method comprises the following steps: (1) weighing 2.00g of camellia seed oil sample, adding 2mL of double distilled water, oscillating, standing, taking an upper oil phase, and fixing the volume to 5mL to obtain a sample solution; (2) taking a trace starch test kit, and diluting a glucose standard solution with double distilled water in proportion to prepare a standard substance; (3) according to an anthrone colorimetric method of a kit, starch and disaccharide are decomposed into monosaccharide through acid hydrolysis; (4) respectively absorbing blank, standard substance and sample solution, adding color-developing agent, and developing in water bath; (5) measuring the absorbance A value, calculating a standard curve, and calculating the total sugar content in the sample; (6) and judging the camellia oleosa seed oil to be pressing process oil or leaching process oil according to the total sugar content in the oil sample. The invention is simple and fast.
Description
Technical Field
The invention relates to a rapid identification method for oil tea seed oil squeezing and oil tea seed oil leaching, in particular to an identification method for adopting total sugar content as oil tea seed oil squeezing and oil tea seed oil leaching, and belongs to the field of detection of edible vegetable oil in food detection.
Background
The oil tea seed oil is edible oil prepared from fruit of Camellia oleifera Abel of Camellia in Theaceae. Although there are many ways to obtain the oil-tea camellia seed oil, including squeezing, leaching, water enzyme extraction, supercritical CO2Extraction, subcritical extraction and the like, but the two processes of pressing and leaching are most commonly used.
Starch is a high-molecular carbohydrate composed of α -D-glucopyranose as a basic unit, and is the main energy storage substance of seeds. In the camellia seeds, the fat content is 30-50%, and the starch content is 10-20%. The starch exists in the form of starch granules with a particle size of about 1.2-2.0 μm in the camellia seed starch cells. Fat is present as oil bodies in the adipocytes of seeds, and generally, both are not present in the same cell. The soluble sugar in the camellia seeds mainly comprises sucrose, fructose, glucose and the like, and is slightly soluble in triglyceride, and the content of the three soluble sugars in the camellia seeds is about 10-55 mg/g.
The oil tea seed oil is processed by adopting a solvent extraction process, oil and fat are dissolved from seeds by utilizing the characteristic of polarity similarity between an organic solvent (generally a 6# solvent, or normal hexane, petroleum ether and the like) and triglyceride under the heating condition, and then the solvent is recovered under the conditions of temperature rise and pressure reduction to obtain the edible oil tea seed oil. The oil tea seed oil is processed by adopting a squeezing process, wherein the oil tea seed raw material is heated and pressed by a squeezer to destroy oil tea seed cells and oil bodies in the cells, and triglyceride is dissolved out.
The edible oil extraction solvent has high solubility to oil, and a solvent extraction method adopted in the processing of edible vegetable oil can obtain high extraction rate which can generally reach 95-98%. However, the high dissolving capacity of the No. 6 solvent, hexane, petroleum ether and other low-polarity solvents may also cause many exogenous hazards to remain in the edible vegetable oil, including solvent residues, and hazards such as polycyclic aromatic hydrocarbons, plasticizers, aflatoxins and the like, and the problems are further aggravated by the repeated use of the solvent and the high-temperature desolventizing process in the processing process. Under the existing process conditions, even if the finished product processed by the refining process is extracted with the edible oil, the solvent residue cannot be completely removed, and the limit of the edible vegetable oil hygiene standard GB2716-2018 in China on the solvent residue in the extracted finished product vegetable oil is 20 mg/kg.
In the actual production process, fresh raw materials are generally adopted for the extracted camellia seed oil, the finished oil is also sold as high-end woody edible oil, the extracted camellia seed oil generally adopts a pressed cake as the extraction raw material, the selling price of the extracted camellia seed oil is far lower than that of the extracted camellia seed oil due to the difference of the raw materials and the difference of the product quality, the price of the extracted camellia seed oil is 80-120 yuan/500 mL in 2019, and the price of the extracted camellia seed oil is 50-60 yuan/500 mL. For ordinary consumers, the two product oils are basically not identified by visual inspection, and the national standard also adopts the solvent content as the basis for identification (the solvent content required by pressed vegetable oil is 0, and the solvent content required by extracted vegetable oil is less than or equal to 20 mg/kg). The great price difference and the identification difficulty lead people to go away from danger and the extracted oil is taken as the pressed oil.
The solvent residual quantity of the vegetable oil is measured by adopting the method in the national standard GB 5009.262-2016, adopting gas chromatography, adopting headspace sample injection, requiring 5g of sample, and detecting 1 sample for more than 40min (including balance time and chromatographic separation detection time), and even if continuous sample injection is carried out, the detection time cannot be shortened.
The starch has an adsorption effect on the grease, and the specially processed porous starch can adsorb even 80% of the grease by weight of the starch. In the solvent extraction process of the oil-tea camellia seed oil, due to polarity, when the oil is extracted by using an organic solvent, the starch is not dissolved in organic solvents such as normal hexane and the like, so that the residual amount of the starch in the product is very small; similarly, sucrose, fructose and glucose are not substantially dissolved in the extraction solvent due to the difference in polarity. In the process of squeezing the oil-tea camellia seed oil, the temperature and the pressure break the cells in the oil-tea camellia seed, not only the fat-containing oil body cells are broken, but also the starch cells containing starch granules are crushed, so that when the oil in the oil body flows out under high pressure, a large amount of starch granules and soluble sugar are brought out and mixed with the oil.
Although the processes of filtration, alkali refining, winterization and the like can also remove starch and soluble sugar in a part of grease in the subsequent processing process of the camellia seed oil, the starch and the soluble sugar are not completely removed. At present, a simple and rapid method for rapidly identifying oil-tea camellia seed oil by squeezing and leaching does not exist.
Disclosure of Invention
The invention aims to overcome the defects in the prior art and provide a simple and rapid method for rapidly identifying the oil-tea camellia seed oil by squeezing and leaching.
The technical scheme adopted by the invention for solving the problems is as follows: the rapid identification method for squeezing and leaching camellia seed oil is characterized in that: the rapid identification method comprises the following steps:
(1) weighing 2.00g of camellia seed oil sample, adding 2mL of double distilled water, carrying out vortex oscillation at 1000rpm for 1min at the temperature of 45-50 ℃, standing for 5min for layering, taking the upper oil phase, repeatedly extracting for 2 times, combining the lower water phase, and fixing the volume to 5mL to obtain a sample solution;
(2) diluting glucose standard solution with double distilled water at a certain ratio to obtain standard substance with concentration of 0.00, 0.01, 0.02, 0.03, 0.04, 0.06, 0.08, 0.16 mg/mL;
(3) according to the anthrone colorimetric method of the kit, starch and disaccharide are decomposed into monosaccharide through acid hydrolysis;
(4) respectively absorbing 50 mu L of blank (double distilled water), 8 standard products prepared in the step (2) and the sample solution prepared in the step (1), and adding a color developing agent to carry out water bath color development;
(5) after the developed solution is cooled to room temperature, measuring the absorbance A value at the wavelength of 620nm, calculating a standard curve, and calculating the total sugar content in the sample according to the standard curve and the absorbance value of the sample to be detected;
(6) and judging the camellia oleosa seed oil to be squeezing process oil or leaching process oil according to the total sugar content in the oil sample.
Preferably, the formula for calculating the total sugar content in the oil sample in step (4) of the present invention is:
preferably, in the step (5), when the total sugar content in the finished camellia oleosa seed oil sample is greater than 0.7mg/g, the finished camellia oleosa seed oil is judged to be pressed; and when the total sugar content in the finished product camellia oleosa seed oil sample is less than 0.5mg/g, judging that the finished product camellia oleosa seed oil is extracted.
Preferably, the invention is suitable for the finished product of the camellia seed oil. Taste can be identified because of the heavy solvent residue of the extract in the crude oil/oil.
Preferably, the total sugar used for identification in the present invention includes starch, sucrose, glucose and fructose, excluding macromolecular carbohydrates such as cellulose.
Preferably, the method adopts the existing commercial trace starch test kit, utilizes an enzyme-labeling instrument to directly and quickly detect the content of soluble sugar in the oil-tea camellia seed oil, judges whether the source of a sample to be detected belongs to the squeezing process or the leaching process according to the content of the soluble sugar in the oil, and is quick and accurate.
Compared with the prior art, the invention has the following advantages and effects: the method judges whether the camellia oleosa seed oil is produced by a pressing process or a leaching process according to the content of total sugar (starch and soluble sugar) in the oil. The invention creatively analyzes the total sugar content in a large amount of squeezed and extracted camellia seed oil, obtains a simple and rapid method for identifying the squeezed and extracted camellia seed oil, and has the following advantages and effects:
(1) the method is simple and rapid, and more than 80 samples can be identified simultaneously within 40 minutes (96-well plate); the existing gas method can only identify 1 sample within 40 min;
(2) the cost is low, and each sample only needs about 1.0 yuan on average (calculated by the price of a micro starch test kit of 2019 Beijing Solebao science and technology Co., Ltd.);
(3) the required sample amount is small, and only 2 g of sample is needed;
(4) the operation is simple, and no complex pretreatment process is needed; the kit can be used for automatic detection or manual detection by an instrument, and after the content determination result is detected, the identification result can be obtained by contrasting the comparison data;
(5) the standard sample and the test sample are simultaneously detected, the sample is simultaneously detected while the standard curve and the fitting equation are obtained, the condition that errors are possibly caused by instrument drift due to the fact that the sample is tested after the standard sample is tested is avoided, and the detection accuracy is high.
Drawings
In order to more clearly illustrate the embodiments and/or technical solutions of the present invention, the drawings used in the description of the embodiments and/or the prior art will be briefly described below, it is obvious that the drawings in the following description are only some embodiments of the present invention, and other drawings can be obtained by those skilled in the art without creative efforts.
FIG. 1 is a schematic representation of a standard curve of total sugar content and a fitted equation in an example of the invention.
Fig. 2 is a schematic diagram of the comparison of the total sugar content in the pressed camellia oleosa seed oil and the leached camellia oleosa seed oil in the example of the invention.
Detailed Description
The present invention will be described in further detail below by way of examples with reference to the accompanying drawings, which are illustrative of the present invention and are not to be construed as limiting the present invention.
Example 1:
referring to fig. 1 to 2, the specific detection steps of the rapid identification method for pressing and leaching camellia oleosa seed oil in the embodiment are as follows:
the method is carried out by using reagents in a starch content measuring kit (in the example, a micro starch content measuring kit of Beijing Sorboard science and technology Co., Ltd., a product number BC 0700). The kit comprises 4 solvents: a standard substance, 10mg of glucose; ② reagent 1, ethanol with the concentration of 80 percent, reagent 1 can not be used in the invention; ③ reagent 2, acid hydrolysis liquid; fourthly, reagent 3, anthrone color reagent.
(1) Sample extraction: weighing 2.00g of camellia seed oil sample, adding 2mL of double distilled water, carrying out vortex oscillation at 1000rpm for 1min at the temperature of 45-50 ℃, standing for 5min for layering, taking the upper oil phase, repeatedly extracting for 2 times, combining the lower water phase, and fixing the volume to 5mL to obtain a sample solution;
(2) preparing a standard substance: taking a starch test kit, diluting a glucose standard solution with double distilled water in proportion to prepare solutions with concentrations of 0.00, 0.01, 0.02, 0.03, 0.04, 0.06, 0.08 and 0.16mg/mL for later use;
(3) preparing a working solution: adding 5.25mL of double distilled water into the reagent 3 before use, slowly adding 29.75mL of concentrated sulfuric acid, continuously stirring, fully dissolving, and standing for later use;
(4) acid hydrolysis: sucking 0.5mL of the solution obtained in the step (1), adding the solution into a 1mL test tube, adding 0.5mL of reagent 2 into the test tube, and performing oscillation extraction at normal temperature for 10min to obtain a sample solution to be detected; if the absorbance of the final test is less than 0.1 or more than 1, diluting by corresponding times and then performing the subsequent steps for determination;
(5) and (3) color development reaction: respectively absorbing 50 mu L of double distilled water (blank sample), the standard solution and the sample solution to be detected prepared in the step (4), adding the solution into an EP tube, then adding 250 mu L of the working solution prepared in the step (3), tightly covering the EP tube, placing the EP tube into a water bath at 95 ℃ for reaction for 10min, taking out the EP tube and naturally cooling the EP tube;
(6) and (3) determination: preheating an enzyme-labeling instrument (or using a spectrophotometer under the condition of less samples) for 30min, adjusting the wavelength to 620nm, and adjusting zero by using double distilled water; sucking 200 mu L of the sample developed in the step (5), adding the sample into a 96-well plate (or a cuvette), and measuring an absorbance value A at the wavelength of 620nm, wherein A is the sample absorbance value-blank absorbance value;
(7) calculating a standard curve: and (3) calculating to obtain a regression equation by taking the concentration of 8 standard samples as an abscissa and the absorbance value as an ordinate: y-5.9554 x-0.0285, R20.9981(y, absorbance value; x, sample concentration, mg/mL);
(8) calculating the total sugar content in the sample: substituting the absorbance value of the sample into the regression equation in the step (7) to calculate and obtain the glucose concentration x (mg/mL) of the sample, and calculating the total sugar content in the oil sample according to the following test:
(9) and (4) judging the camellia oleosa seed oil to be pressing process oil or leaching process oil according to the content of the total sugar in the oil sample obtained by calculation in the step (8).
Example 2:
and establishing a judgment basis.
(1) Purchasing in supermarket, sampling in enterprise and self-making 62 oil tea seed oil samples, wherein the amount of the pressed oil tea seed oil and the amount of the extracted oil tea seed oil are respectively 31.
(2) The content of total sugar in 62 oil samples obtained by detection using the rapid identification method in example 1, wherein the total sugar comprises starch, glucose, sucrose, fructose and the like, and the results are shown in the following table:
TABLE 1 test results of total sugar content in pressed and extracted finished camellia oleosa seed oil
(3) From the above data obtained, we derive: the average value of the total sugar content of the pressed oil-tea camellia seed oil finished oil is 1.1184 +/-0.1370 mg/g, and the extremely lowest value is 0.7720 mg/g; the total sugar content of the extracted oil-tea camellia seed oil finished oil is 0.1763 +/-0.1059 mg/g, and the extreme maximum value is 0.4850 mg/g; t-test analysis of two sets of data by origin showed that the two sets of data are very different (p is less than 0.01), so that the data can be used as identification basis. The invention takes the total sugar content of more than 0.7mg/g as the judgment standard of the squeezed oil and the total sugar content of less than 0.5mg/g as the judgment standard of the extracted oil.
(4) And (3) verification: and (4) tearing off the label by using a sampling sample of a food detection mechanism, marking sequential numbers, analyzing, judging by using the judgment standard in the step (3), and judging by using 11 pieces of squeezed camellia seed oil and 7 pieces of extracted camellia seed oil, wherein the misjudgment rate is zero.
Example 3:
results regarding detection accuracy:
in the sample preparation step, the oil sample may be directly tested with 50 μ L of the original camellia seed oil sample without any treatment, or the camellia seed oil sample may be mixed with double distilled water and extracted according to the step (1) in the rapid identification method in example 1. The second method adopts double distilled water extraction to reduce the interference of various complex elements such as pigments, sterols, phospholipids and the like in the oil phase, the precision and the accuracy are greatly improved, the following table shows the comparison of the results of the direct test of the same camellia seed oil sample and the test after the double distilled water extraction, and each method is repeated for 6 times:
table 2 comparison of accuracy of direct test and test results after extraction (n ═ 6)
| Method | Total sugar content results mg/g | Coefficient of variation |
| Direct testing of oil tea seed oil sample | 1.1103±0.1198 | 10.79％ |
| Oil samples were tested after double distilled water extraction | 1.0278±0.0343 | 3.33％ |
According to the results in table 2, the test results after the oil sample extraction are slightly lower than the test results without extraction, but according to the T test results of the results, the two sets of data have no significant difference, so the extraction test results can be used for representing the starch content in the oil sample. Moreover, the method of directly testing by using the oil sample has the result that the variation coefficient reaches 10.79 percent, the error is large, and the method of testing after extracting the oil sample has the variation coefficient of only 3.33 percent, and the method of testing after extracting improves the result precision by more than 3 times.
Example 4:
the rapid identification method for pressing and leaching camellia oleosa seed oil in the embodiment comprises the following steps:
(1) weighing 1 part of oil tea seed oil of unknown squeezing processing or extraction processing technology, weighing 2.05g of oil sample, adding the oil sample into a 5mL test tube, adding 2mL of double distilled water into the test tube, carrying out vortex oscillation at the temperature of 45 ℃ and the rpm of 1000 for 1min, standing for 5min for layering, taking the upper oil phase, repeatedly extracting for 2 times, combining the lower water phase, and fixing the volume to 5mL to obtain a sample solution;
(2) diluting the glucose standard solution with double distilled water to obtain solutions of 0.00, 0.01, 0.02, 0.03, 0.04, 0.06, 0.08 and 0.16mg/mL concentration for use;
(3) preparing a working solution: adding 5.25mL of double distilled water into the reagent 3 before use, slowly adding 29.75mL of concentrated sulfuric acid, continuously stirring, fully dissolving, and standing for later use;
(4) sucking 0.5mL of the sample solution obtained in the step (1), adding the sample solution into a 1mL test tube, adding a reagent 2 in a 0.5mL kit into the test tube (the kit is the same as that in the embodiment 1), and performing oscillation extraction for 10min at normal temperature to obtain a sample solution to be developed;
(5) and (3) color development reaction: respectively absorbing 50 mu L of double distilled water (blank sample), the standard solution and the sample to be detected, adding into an EP tube, then adding 250 mu L of the working solution prepared in the step (3), covering tightly the cover, placing into a water bath at 95 ℃ for reaction for 10min, taking out and naturally cooling;
(6) preheating a microplate reader for 30min, adjusting the wavelength to 620nm, and adjusting to zero by using double distilled water; sucking 200 mu L of the sample developed in the step (5), adding the sample into a 96-well plate (or a cuvette), and measuring an absorbance value A at 620nm, wherein A is the sample absorbance value-blank absorbance value; table 3 shows the test results.
Table 3 test result data
| Glucose concentration | Absorbance of the solution |
| mg/mL | A |
| 0.000 | -0.0017 |
| 0.010 | 0.0198 |
| 0.020 | 0.0795 |
| 0.030 | 0.1489 |
| 0.040 | 0.221 |
| 0.060 | 0.3187 |
| 0.080 | 0.444 |
| 0.160 | 0.935 |
| Unknown sample | 0.0227 |
(7) According to the test results in the table 2, the regression equation is calculated by taking the concentration of 8 standard samples as the abscissa and the absorbance value as the ordinate: 5.9719x-0.0279, R2＝0.998；
(8) Substituting the unknown sample with an absorbance value of 0.0227 into the regression equation in the step (7) to obtain a sample concentration of 0.0085mg/mL, and calculating the total sugar content in the oil sample to be 0.0413 mg/g;
(9)0.0413mg/g is far less than 0.5mg/g, and the camellia oleosa seed oil of the unknown process is judged to be leaching process oil.
Although the present invention has been described with reference to the above embodiments, it should be understood that the scope of the present invention is not limited thereto, and that various changes and modifications can be made by those skilled in the art without departing from the spirit and scope of the present invention.
Claims (6)
1. A rapid identification method for squeezing and leaching camellia seed oil is characterized in that: the rapid identification method comprises the following steps:
(1) weighing 2.00g of camellia seed oil sample, adding 2mL of double distilled water, carrying out vortex oscillation at 1000rpm for 1min at the temperature of 45-50 ℃, standing for 5min for layering, taking the upper oil phase, repeatedly extracting for 2 times, combining the lower water phase, and fixing the volume to 5mL to obtain a sample solution;
(2) diluting glucose standard solution with double distilled water at a certain ratio to obtain standard substance with concentration of 0.00, 0.01, 0.02, 0.03, 0.04, 0.06, 0.08, 0.16 mg/mL;
(3) according to an anthrone colorimetric method of a kit, starch and disaccharide are decomposed into monosaccharide through acid hydrolysis;
(4) respectively absorbing 50 mu L of blank, 8 standard substances prepared in the step (2) and the sample solution prepared in the step (1), and adding a color developing agent to carry out water bath color development;
(5) after the developed solution is cooled to room temperature, measuring the absorbance A value at the wavelength of 620nm, calculating a standard curve, and calculating the total sugar content in the sample according to the standard curve and the absorbance value of the sample to be detected;
(6) and judging the camellia oleosa seed oil to be squeezing process oil or leaching process oil according to the total sugar content in the oil sample.
2. The method for rapidly identifying pressed and leached camellia oleosa seed oil according to claim 1, characterized in that: the formula for calculating the total sugar content in the oil sample in the step (4) is as follows:
3. the method for rapidly identifying pressed and leached camellia oleosa seed oil according to claim 1, characterized in that: in the step (5), when the total sugar content in the finished camellia oleosa seed oil sample is more than 0.7mg/g, judging that the finished camellia oleosa seed oil is squeezed; and when the total sugar content in the finished product camellia oleosa seed oil sample is less than 0.5mg/g, judging that the finished product camellia oleosa seed oil is extracted.
4. The method for rapidly identifying pressed and leached camellia oleosa seed oil according to claim 1, characterized in that: is suitable for the finished product of camellia oleosa seed oil.
5. The method for rapidly identifying pressed and leached camellia oleosa seed oil according to claim 1, characterized in that: the total sugars of the substances used for identification included starch, sucrose, glucose and fructose, excluding cellulose.
6. The method for rapidly identifying pressed and leached camellia oleosa seed oil according to claim 1, characterized in that: the method adopts the existing commercial micro-starch test kit, utilizes an enzyme-labeling instrument to directly and rapidly detect the content of soluble sugar in the oil-tea camellia seed oil, and judges whether the source of a sample to be detected belongs to the squeezing process or the leaching process according to the content of the soluble sugar in the oil.
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