CN110790736A - Method for extracting and purifying carminic acid in soft sweets - Google Patents
Method for extracting and purifying carminic acid in soft sweets Download PDFInfo
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- CN110790736A CN110790736A CN201911116811.7A CN201911116811A CN110790736A CN 110790736 A CN110790736 A CN 110790736A CN 201911116811 A CN201911116811 A CN 201911116811A CN 110790736 A CN110790736 A CN 110790736A
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
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- C07D309/00—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only ring hetero atom, not condensed with other rings
- C07D309/02—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only ring hetero atom, not condensed with other rings having no double bonds between ring members or between ring members and non-ring members
- C07D309/08—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only ring hetero atom, not condensed with other rings having no double bonds between ring members or between ring members and non-ring members with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
- B01D15/08—Selective adsorption, e.g. chromatography
- B01D15/10—Selective adsorption, e.g. chromatography characterised by constructional or operational features
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- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
- B01D15/08—Selective adsorption, e.g. chromatography
- B01D15/42—Selective adsorption, e.g. chromatography characterised by the development mode, e.g. by displacement or by elution
- B01D15/424—Elution mode
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- G—PHYSICS
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- G01N30/02—Column chromatography
- G01N30/26—Conditioning of the fluid carrier; Flow patterns
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Abstract
The invention discloses a method for extracting and purifying carminic acid in soft sweets, which comprises the following steps: (1) adding water into a sample, heating at 70-80 ℃ to dissolve the sample, sequentially adding a saturated sodium sulfate solution and a dilute acid solution, and uniformly mixing to obtain a sample solution; (2) centrifuging the sample solution, and adding the centrifugate into a chromatographic column filled with polyamide; (3) eluting the chromatographic column with a washing solution, eluting the chromatographic column with water until the eluent is neutral, and discarding the eluent; (4) eluting the chromatographic column with resolving liquid for multiple times, collecting eluate, and adding phosphoric acid solution. Aiming at the soft sugar products, the carminic acid is adsorbed by polyamide powder to remove impurities in an acidic environment, and after the adsorption and the elution are carried out by using the analytic liquid, the complex matrixes such as the sugar and the like can be effectively separated from the carminic acid to obtain the carminic acid extracting solution with high purity, so that the problem of inaccurate detection caused by low content of the carminic acid in the soft sweets is solved, and the accurate detection of the content of the carminic acid in the soft sweets is realized.
Description
Technical Field
The invention belongs to the technical field of health-care products and foods, and particularly relates to a method for extracting and purifying carminic acid in soft sweets.
Background
Cochineal red is a natural pigment extracted from cochineal bodies, is pink to purple, is an anthraquinone natural pigment, is different from other natural pigments, has stable physicochemical properties, is regarded as the safest natural pigment, and can be used as a colorant for drinks such as wine, fruit pulp and cold drinks, candies, cakes, meats, sausages and the like. GB 2760 supplement 2014 food safety national standard food additive use standard specifies the addition amount of cochineal red, and the addition amount needs to be calculated by carminic acid.
The currently reported detection methods of carminic acid mainly include spectrophotometry, thin layer chromatography, high performance liquid chromatography and the like. In the detection process of the spectrophotometry, the total absorbance at a single wavelength is required, so that the condition that non-carminic acid also absorbs at the wavelength cannot be eliminated, the accuracy of the detection result is influenced, and the analysis and the determination are time-consuming. And the high performance liquid chromatography is easily interfered by complex matrixes in food and is limited in application. The conventional high performance liquid chromatography only simply processes a sample, and the sample facing complex matrixes such as saccharides and the like in actual detection cannot effectively separate carminic acid, is difficult to characterize and easily blocks a chromatographic column, so that the service life of the chromatographic column is greatly reduced. Meanwhile, the content of carminic acid in the soft candy sample is very low, and the carminic acid cannot be effectively detected only by simple treatment, so that the aim of controlling the carminic acid is difficult to achieve.
Disclosure of Invention
In order to overcome the defects of the prior art, the invention aims to provide a method for extracting and purifying carminic acid from soft sweets, which can effectively separate complex matrixes such as saccharides and the like from carminic acid and improve the accuracy of detecting the content of carminic acid.
The invention is realized by the following technical scheme:
a method for extracting and purifying carminic acid in soft sweets comprises the following steps:
(1) adding water into a soft sweet sample, heating at 70-80 ℃ to dissolve the sample, sequentially adding a saturated sodium sulfate solution and a dilute acid solution, and uniformly mixing to obtain a sample solution; the dilute acid is citric acid or formic acid;
(2) centrifuging the sample solution, and adding the centrifugate into a chromatographic column filled with polyamide;
(3) eluting the chromatographic column with a washing solution, eluting the chromatographic column with water until the eluent is neutral, and discarding the eluent; the washing liquid contains (0-4) by volume: (1-3): (18-30) a mixed aqueous solution of a saturated sodium sulfate solution, a citric acid solution and water;
(4) eluting the chromatographic column for multiple times by using the resolving liquid, collecting the eluent, and adding a phosphoric acid solution to obtain a carminic acid extracting solution; the analysis solution contains (0.8-1.2) by volume: (8-12): (36.8-41.2) a mixed aqueous solution of ammonia water, ethanol and water.
Preferably, in the step (1), the ratio of the soft candy sample, water, saturated sodium sulfate solution and dilute acid solution is (0.1-5) g: (8-30) ml: (0-4) ml: (0.4-3) ml.
Preferably, in the step (1) and the step (2), the mass concentration of the citric acid is 0.8-1.2%.
Preferably, in the step (2), the rotation speed of the centrifugation is 6000 to 8000r/min, and the centrifugation time is 3 to 5 min.
Preferably, in the step (1), after two solvents, namely a saturated sodium sulfate solution and a dilute acid solution, are sequentially added, the sample solution needs to be ensured not to generate solidification and gel substances, and after centrifugation, the centrifugate is clear and has good fluidity and can smoothly pass through a chromatographic column filled with polyamide powder (the flow rate is more than 0.5 drop/S); if the requirement cannot be met, in the step (2), the sample solution is dissolved by absolute ethyl alcohol and then centrifuged to obtain centrifugate and residue, and the centrifugate is added into a chromatographic column filled with polyamide for elution.
Further preferably, the residue is washed for 1-2 times by using ethanol with the volume concentration of 60-70%, the centrifugate and the ethanol washing liquid are combined and concentrated under reduced pressure at the temperature of 60-80 ℃ to obtain a concentrated solution, and the concentrated solution is transferred to a chromatographic column filled with polyamide by using a citric acid solution for elution.
Preferably, in the step (2), the particle size of the polyamide is 100-200 meshes, the specification of the chromatographic column is 10mm multiplied by 250mm, a cock is arranged, and the polyamide powder is dispersed by a small amount of 1% citric acid solution and transferred to the chromatographic column.
Preferably, in the step (4), the volume concentration of the phosphoric acid solution is 3-4%.
Compared with the prior art, the invention has the following beneficial effects:
the invention provides a method for extracting and purifying carminic acid in soft sweets aiming at soft sugar products, wherein the carminic acid is adsorbed by polyamide powder to remove impurities in an acidic environment, and after being eluted by analytic liquid, complex matrixes such as sugar and the like can be effectively separated from the carminic acid to obtain a high-purity carminic acid extracting solution, so that the problem of inaccurate detection caused by low content of the carminic acid in the soft sweets is solved, and the accurate detection of the content of the carminic acid in the soft sweets is realized.
Detailed Description
The present invention is further illustrated by the following specific embodiments, which are not intended to limit the scope of the invention.
The raw materials of the invention are all commercially available.
Example 1: extraction and purification of carminic acid in acerola cherry vitamin C soft sweets
(1) Adding 3g of acerola cherry vitamin C soft candy sample into 20mL of water, heating the mixture in a water bath at 70 ℃ to dissolve the sample, sequentially adding 4mL of saturated sodium sulfate solution and 2mL of 1% citric acid solution, and uniformly mixing to obtain a sample solution;
(2) centrifuging the sample solution at 8000r/min for 5min, adding centrifugate into chromatographic column filled with polyamide (weighing polyamide powder 0.5g, dispersing with small amount of 1% citric acid solution, transferring into chromatographic column), eluting the chromatographic column with 10ml of washing solution, eluting the chromatographic column with water until the eluate is neutral, and discarding the eluate; the washing solution is a mixed aqueous solution of 4mL of saturated sodium sulfate solution and 1mL of 1% citric acid solution, and water is added to the mixed aqueous solution to 25 mL;
(3) eluting the chromatographic column for multiple times by using 9mL of resolution solution, collecting eluent, and adding 0.5mL of 3% phosphoric acid solution to obtain a carminic acid extracting solution; the analysis solution is a mixed aqueous solution of 1mL of ammonia water and 10mL of ethanol, and water is added to 50 mL.
Example 2: extraction and purification of carminic acid in acerola cherry vitamin C soft sweets
(1) Adding 10mL of water into 3g of acerola cherry vitamin C soft candy sample, heating the mixture in a water bath at 70 ℃ to dissolve the sample, sequentially adding 1mL of saturated sodium sulfate solution and 0.5mL of 1% formic acid solution, and uniformly mixing to obtain a sample solution;
(2) adding absolute ethyl alcohol into the sample solution to 50mL, quickly and uniformly mixing, and centrifuging at 8000r/min for 5min to obtain a centrifugal liquid and residues; washing residues with ethanol with the volume concentration of 70% for 2 times, 5mL each time, combining the centrifugate and the ethanol washing solution, and concentrating under reduced pressure at 60-80 ℃ to about 5mL to obtain a concentrated solution; transferring the concentrated solution to a chromatographic column filled with polyamide (0.5 g of polyamide powder is weighed, dispersed by a small amount of 1% citric acid solution and transferred to the chromatographic column) by 15mL of 1% citric acid solution in several times, eluting with 1% citric acid solution until the effluent is colorless, then washing with water until the effluent is neutral, and discarding the eluate;
(3) eluting the chromatographic column for multiple times by using 9mL of resolution solution, collecting eluent, and adding 0.4mL of 3% phosphoric acid solution to obtain a carminic acid extracting solution; the analysis solution was a mixed aqueous solution of 1.2mL of ammonia water and 8mL of ethanol, and water was added to 50 mL.
Comparative example 1: (see CN201510545167.0 method)
Taking 3g of acerola cherry vitamin C soft sweet sample, adding 10mL of water, heating in water bath at 70 ℃ to dissolve the sample, adding water to a constant volume, filtering with a microporous filter membrane, and measuring.
Comparative example 2:
taking 3g of acerola cherry vitamin C soft candy sample, adding 30 mL of mixed liquor of ethanol/ammonia water/water (7: 2: 1), carrying out ultrasonic extraction for 30min, adding 20mL of mixed liquor for the second time, carrying out ultrasonic extraction for 30min, combining the extracting solutions, drying by distillation in a water bath, carrying out constant volume with an ammonia water solution with the pH =9.0, filtering, and detecting.
The test solutions prepared in examples 1-2 and comparative examples 1-2 were measured by high performance liquid chromatography, wherein the chromatographic conditions were: a chromatographic column: an LC Column C18 chromatographic Column (250 mm × 4.6mm, 4 μm); mobile phase: mobile phase A: taking 6.80g of monopotassium phosphate, adding 1000mL of water for dissolving, and adjusting the pH value to 3.0 by using phosphoric acid; mobile phase B: acetonitrile, flow rate: 1.0mL/min, column temperature: 35 ℃, wavelength: 494 nm; the gradient elution was performed according to the following table 1, and the detection results are shown in table 2:
TABLE 1 gradient elution Table
| Time (min) | Mobile phase A (%) | Mobile phase B (%) |
| 0 | 95 | 5 |
| 25 | 75 | 25 |
| 25.01 | 95 | 5 |
| 30 | 95 | 5 |
TABLE 2 test results
| Theoretical content | Detecting the content | |
| Example 1 | 2.6mg/kg | 2.7mg/kg |
| Example 2 | 2.6mg/kg | 2.5mg/kg |
| Comparative example 1 | 2.6mg/kg | Is unable to detect |
| Comparative example 2 | 2.6mg/kg | Not detected out |
As can be seen from the results in Table 2, the sample in comparative example 1 can not pass through the membrane, can seriously block the chromatographic column and can not be detected; comparative example 2 the fondant was not soluble in the ethanol/ammonia/water (7: 2: 1) mixture and carminic acid was not extracted and was not detected; according to the method, the carminic acid is adsorbed by polyamide powder to remove impurities in an acidic environment, and after the carminic acid is eluted by the analytic solution, complex matrixes such as saccharides and the like can be effectively separated from the carminic acid, and the detected carminic acid is closer to a theoretical value, so that the method disclosed by the invention can be used for accurately detecting the content of the carminic acid in the soft candy.
Claims (8)
1. A method for extracting and purifying carminic acid in soft sweets is characterized by comprising the following steps:
(1) adding water into a soft sweet sample, heating at 70-80 ℃ to dissolve the sample, sequentially adding a saturated sodium sulfate solution and a dilute acid solution, and uniformly mixing to obtain a sample solution; the dilute acid is citric acid or formic acid;
(2) centrifuging the sample solution, and adding the centrifugate into a chromatographic column filled with polyamide;
(3) eluting the chromatographic column with a washing solution, eluting the chromatographic column with water until the eluent is neutral, and discarding the eluent; the washing liquid contains (0-4) by volume: (1-3): (18-30) a mixed aqueous solution of a saturated sodium sulfate solution, a citric acid solution and water;
(4) eluting the chromatographic column for multiple times by using the resolving liquid, collecting the eluent, and adding a phosphoric acid solution to obtain a carminic acid extracting solution; the analysis solution contains (0.8-1.2) by volume: (8-12): (36.8-41.2) a mixed aqueous solution of ammonia water, ethanol and water.
2. The method for extracting and purifying carminic acid from soft sweets according to claim 1, wherein in the step (1), the ratio of the soft sweets sample, the water saturated sodium sulfate solution and the dilute acid solution is (0.1-5) g: (8-30) ml: (0-4) ml: (0.4-3) ml.
3. The method for extracting and purifying carminic acid from soft sweets according to claim 1, wherein in the step (1) and the step (2), the mass concentration of the citric acid is 0.8% -1.2%.
4. The method for extracting and purifying carminic acid from soft sweets according to claim 1, wherein in the step (2), the rotation speed of the centrifugation is 6000-8000 r/min, and the centrifugation time is 3-5 min.
5. The method for extracting and purifying carminic acid from soft sweets according to claim 1, wherein in the step (2), the sample solution is dissolved by absolute ethyl alcohol and then centrifuged to obtain a centrifugate and a residue, and the centrifugate is added into a chromatographic column filled with polyamide for elution.
6. The method for extracting and purifying carminic acid from soft sweets according to claim 5, wherein the residues are washed with ethanol with a volume concentration of 60-70%, the centrifugate and the ethanol washing solution are combined, the mixture is concentrated under reduced pressure at 60-80 ℃ to obtain a concentrated solution, and the concentrated solution is transferred to a chromatographic column filled with polyamide by using a citric acid solution for elution.
7. The method for extracting and purifying carminic acid from soft sweets as claimed in claim 1, wherein in step (2), the particle size of the polyamide is 100-200 mesh.
8. The method for extracting and purifying carminic acid from soft sweets according to claim 1, wherein in the step (4), the volume concentration of the phosphoric acid solution is 3% to 4%.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| DE3319756A1 (en) * | 1982-06-02 | 1983-12-08 | Vysoká škola chemicko-technologická Praha, Praha | Process for isolating carminic acid |
| CN107037152A (en) * | 2017-04-14 | 2017-08-11 | 汤臣倍健股份有限公司 | The detection method of carminic acid content in a kind of gelatine capsule |
| CN108287092A (en) * | 2017-12-29 | 2018-07-17 | 东莞理工学院 | For detecting synthetic dyestuff standard sample and preparation method thereof in hard candy |
-
2019
- 2019-11-15 CN CN201911116811.7A patent/CN110790736A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| DE3319756A1 (en) * | 1982-06-02 | 1983-12-08 | Vysoká škola chemicko-technologická Praha, Praha | Process for isolating carminic acid |
| CN107037152A (en) * | 2017-04-14 | 2017-08-11 | 汤臣倍健股份有限公司 | The detection method of carminic acid content in a kind of gelatine capsule |
| CN108287092A (en) * | 2017-12-29 | 2018-07-17 | 东莞理工学院 | For detecting synthetic dyestuff standard sample and preparation method thereof in hard candy |
Non-Patent Citations (3)
| Title |
|---|
| 付忠源: "高效液相色谱法测定猪肉脯中胭脂虫红的含量", 《理化检验(化学分册)》 * |
| 戚穗坚，杨丽主编: "《普通高等教育"十五"国家级规划教材 食品分析实验指导》", 31 August 2018, 中国轻工业出版社 * |
| 顾正健等: "高效液相色谱法同时测定冷饮中禁限用色素胭脂虫红酸、喹啉黄和碱性蓝26", 《化学试剂》 * |
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