CN103940926B - A kind of method of antioxidant content in quick detection Radix Et Rhizoma Rhei seed - Google Patents
A kind of method of antioxidant content in quick detection Radix Et Rhizoma Rhei seed Download PDFInfo
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- CN103940926B CN103940926B CN201410170083.9A CN201410170083A CN103940926B CN 103940926 B CN103940926 B CN 103940926B CN 201410170083 A CN201410170083 A CN 201410170083A CN 103940926 B CN103940926 B CN 103940926B
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Abstract
The present invention relates to a kind of method of antioxidant content in quick detection Radix Et Rhizoma Rhei seed, the method comprises the following steps: obtain Radix Et Rhizoma Rhei seed powder after (1) being pulverized by Radix Et Rhizoma Rhei seed；(2) Radix Et Rhizoma Rhei seed powder methanol is extracted, after filtration, obtain Radix Et Rhizoma Rhei seed extracting solution；(3) Radix Et Rhizoma Rhei seed extracting solution separates through high performance liquid chromatography chromatographic column, the composition separated first flows through after first UV-detector as sample, the antioxidant that mass concentration is 10 ~ 200 mg/L 1 pumped into mutually as flowing in this sample and second pump, after 1 diphenyl 2 trinitrophenyl-hydrazine is derivative in reaction ring, enter in second UV-detector, then the chromatogram that first UV-detector and second UV-detector are recorded is compared, when the chromatogram that second UV-detector is recorded occurs the compound of negative peak, this compound is the antioxidant content in extract.The present invention is simple, quickly, it is possible to achieve antioxidant content in Radix Et Rhizoma Rhei seed is carried out the evaluation of effectiveness.
Description
Technical field
A kind of method that the present invention relates to quick detection antioxidant content, particularly relates in a kind of quickly detection Radix Et Rhizoma Rhei seed
The method of antioxidant content.
Background technology
Chinese herb rhubarb be Polygonaceae Rheum plant Rheum tanguticum (Rheum tanguticum Maxim. ex Balf.
), sorrel (Rheum p almatum Linn.) or Rheum officinale (Rh. of f icinale Baill.)
It is dried root and rhizome.Rheum tanguticum has another name called monarch's wood and pricks (Tibetan name), rheum tanguticum Tschircs, belongs to Qinghai Province genuine medicinal materials, be born in border,
Sylvan life cheuch or shrubbery, height above sea level 2300 ~ 4200m, separately also there is distribution in SOUTH OF GANSU, Eastern Tibet the north, Northwest Sichuan etc..The palm
Leaf Radix Et Rhizoma Rhei main product in Gansu, Qinghai, Tibet, the ground such as Sichuan, predominantly cultivation, yield is maximum.Rheum officinale main product in Sichuan, expensive
The ground such as state, Yunnan, Hubei, Shaanxi, cultivation or wild.
Medical material seed is the source of production of crude drugs and development, is the intrinsic factor determining Chinese crude drug quality, is development high-quality
The science premise of Producing medicinal herbs.Medicinal plant kind is various, and its seed varies owing to the reasons such as heredity, habitat there will be
Form.
Research currently for Radix Et Rhizoma Rhei seed is concentrated mainly on the quality inspections such as germination percentage, mass of 1000 kernel, cleanliness, water content
In index, Radix Et Rhizoma Rhei seed resource enriches, and wherein antioxidation aspect has no report.
1,1-diphenyl-2-trinitrophenyl-hydrazine (DPPH) free radical method has become a kind of antioxidation the most more generally acknowledged
Activity determination method, is frequently used in the antioxidation activity in vitro evaluation study method of bioactive substance.DPPH is having
Being a kind of stable free radical in machine solvent, its lone electron has strong absorption (dark purple) near 517nm, when free radical is clear
In the presence of agent, lone electron is paired, and absorption disappears or weakens, and therefore by measuring the degree that absorption weakens, can evaluate this certainly
Activity by base scavenger.But utilize DPPH method to be only capable of total antioxidant activity in extract to be analyzed, it is impossible to know antioxygen
Change active component, it is impossible to compare the size of each active substance antioxidant activity.
Summary of the invention
The technical problem to be solved is to provide one and simply, the most quickly detects in Radix Et Rhizoma Rhei seed
The method of antioxidant content.
For solving the problems referred to above, the method for antioxidant content in a kind of quick detection Radix Et Rhizoma Rhei seed of the present invention, bag
Include following steps:
(1), after Radix Et Rhizoma Rhei seed meal being broken to 40 ~ 60 mesh, obtain Radix Et Rhizoma Rhei seed powder；
By described Radix Et Rhizoma Rhei seed powder with the methanol that volumetric concentration is 50% ~ 100% of its quality 50 ~ 100 times at power
It is supersound extraction 20 ~ 50min under conditions of 200 ~ 300W, after filtration, obtains Radix Et Rhizoma Rhei seed extracting solution；
The most described Radix Et Rhizoma Rhei seed extracting solution separates through high performance liquid chromatography chromatographic column, and the composition of separation first flows through first purple
As sample after external detector, the mass concentration pumped into mutually as flowing in this sample and second pump is 10 ~ 200 mg/L's
Antioxidant 1, after 1-diphenyl-2 trinitrophenyl-hydrazine is derivative in reaction ring, enters in second UV-detector, then will
The chromatogram that described first UV-detector and described second UV-detector are recorded is compared, when described second
When the chromatogram that UV-detector is recorded occurs the compound of negative peak, the antioxygen chemical conversion that this compound is in extract
Point.
Described step (3) middle high-efficiency liquid chromatography method for detecting refers to that mobile phase A is methanol, and Mobile phase B volumetric concentration is
The acetic acid aqueous solution of 0.1%；Flow velocity is 0.6 ~ 1.2 mL/min, and flowing phase condition is: 0 min ~ 28 min, 35%A ~ 43%A,
65%B~57%B；28 min ~ 50 min, 43%A ~ 60%A, 57%B ~ 30%B.
A length of 5 ~ 25 meters of described step (3) middle reaction ring.
Described step (3) in first UV-detector detection wavelength be 254 nm；Second UV-detector detection wavelength
It is 517 nm.
The present invention compared with prior art has the advantage that
1, use antioxidant content in DPPH-HPLC method on-line determination Radix Et Rhizoma Rhei seed due to the present invention, and this method has institute
, high throughput testing, automaticity few by amount of reagent are high, the detection time is short, sensitivity and specificity relatively high, therefore,
Suitably measure antioxidant content in Radix Et Rhizoma Rhei seed, thus realize Radix Et Rhizoma Rhei seed is carried out the efficiency evaluation of pharmacological action.
2, use antioxidant content in DPPH-HPLC method on-line determination Radix Et Rhizoma Rhei seed due to the present invention, and this method only needs ratio
Relatively enter the chromatogram after first and second detector and i.e. can determine whether antioxidant content.Such as Fig. 1.Relatively each chromatograph in chromatogram
Peak, the chromatogram after entrance the second detector that wherein in chromatogram, the peak of chromatographic peak A, B, C, D, E, F, G, H is corresponding is negative peak,
Illustrate that compound A, B, C, D, E, F, G, H are antioxidant content.
3, the present invention is simple, quickly, it is easy to promote.
Accompanying drawing explanation
Below in conjunction with the accompanying drawings the detailed description of the invention of the present invention is described in further detail.
Fig. 1 is compared figure by the chromatogram that first detector of the present invention and second detector are recorded.
Detailed description of the invention
The method of antioxidant content in embodiment 1 one kinds quick detection Radix Et Rhizoma Rhei seed, comprises the following steps:
(1), after Radix Et Rhizoma Rhei seed meal being broken to 40 mesh, obtain Radix Et Rhizoma Rhei seed powder.
By Radix Et Rhizoma Rhei seed powder with the methanol that volumetric concentration is 50% of its quality 50 times under conditions of power is 200W
Supersound extraction 50min, obtains Radix Et Rhizoma Rhei seed extracting solution after filtration.
(3) Radix Et Rhizoma Rhei seed extracting solution separates through high performance liquid chromatography chromatographic column, and the composition of separation first flows through first ultraviolet inspection
As the antioxidant that mass concentration is 10 mg/L pumped into mutually as flowing in sample, this sample and second pump after surveying device
After 1,1-diphenyl-2 trinitrophenyl-hydrazine is derivative in reaction ring, enter in second UV-detector, then by first purple
The chromatogram that external detector and second UV-detector are recorded is compared, when the color that second UV-detector is recorded
When occurring the compound of negative peak in spectrogram, this compound is the antioxidant content in extract.
Wherein:
High-efficiency liquid chromatography method for detecting refers to that mobile phase A is methanol, Mobile phase B volumetric concentration be 0.1% acetic acid water-soluble
Liquid；Flow velocity is 0.6 mL/min, and flowing phase condition is: 0 min ~ 28 min, 35%A ~ 43%A, 65%B ~ 57%B；28 min ~50
Min, 43%A ~ 60%A, 57%B ~ 30%B.
A length of 5 meters of reaction ring.
First UV-detector detection wavelength is 254 nm；Second UV-detector detection wavelength is 517 nm.
The method of antioxidant content in embodiment 2 one kinds quick detection Radix Et Rhizoma Rhei seed, comprises the following steps:
(1), after Radix Et Rhizoma Rhei seed meal being broken to 60 mesh, obtain Radix Et Rhizoma Rhei seed powder.
(2) it is the condition of 300W with the methanol that volumetric concentration is 80% of its quality 100 times at power by Radix Et Rhizoma Rhei seed powder
Lower supersound extraction 30min, obtains Radix Et Rhizoma Rhei seed extracting solution after filtration.
(3) Radix Et Rhizoma Rhei seed extracting solution separates through high performance liquid chromatography chromatographic column, and the composition of separation first flows through first ultraviolet inspection
As the antioxidant that mass concentration is 100 mg/L pumped into mutually as flowing in sample, this sample and second pump after surveying device
After 1,1-diphenyl-2 trinitrophenyl-hydrazine is derivative in reaction ring, enter in second UV-detector, then by first purple
The chromatogram that external detector and second UV-detector are recorded is compared, when the color that second UV-detector is recorded
When occurring the compound of negative peak in spectrogram, this compound is the antioxidant content in extract.
Wherein:
High-efficiency liquid chromatography method for detecting refers to that mobile phase A is methanol, Mobile phase B volumetric concentration be 0.1% acetic acid water-soluble
Liquid；Flow velocity is 0.8 mL/min, and flowing phase condition is: 0 min ~ 28 min, 35%A ~ 43%A, 65%B ~ 57%B；28 min ~50
Min, 43%A ~ 60%A, 57%B ~ 30%B.
A length of 10 meters of reaction ring.
First UV-detector detection wavelength is 254 nm；Second UV-detector detection wavelength is 517 nm.
The method of antioxidant content in embodiment 3 one kinds quick detection Radix Et Rhizoma Rhei seed, comprises the following steps:
(1), after Radix Et Rhizoma Rhei seed meal being broken to 40 mesh, obtain Radix Et Rhizoma Rhei seed powder.
(2) it is the condition of 300W with the methanol that volumetric concentration is 100% of its quality 100 times at power by Radix Et Rhizoma Rhei seed powder
Lower supersound extraction 30min, obtains Radix Et Rhizoma Rhei seed extracting solution after filtration.
(3) Radix Et Rhizoma Rhei seed extracting solution separates through high performance liquid chromatography chromatographic column, and the composition of separation first flows through first ultraviolet inspection
As the antioxidant that mass concentration is 200 mg/L pumped into mutually as flowing in sample, this sample and second pump after surveying device
After 1,1-diphenyl-2 trinitrophenyl-hydrazine is derivative in reaction ring, enter in second UV-detector, then by first purple
The chromatogram that external detector and second UV-detector are recorded is compared, when the color that second UV-detector is recorded
When occurring the compound of negative peak in spectrogram, this compound is the antioxidant content in extract.
Wherein:
High-efficiency liquid chromatography method for detecting refers to that mobile phase A is methanol, Mobile phase B volumetric concentration be 0.1% acetic acid water-soluble
Liquid；Flow velocity is 1.2 mL/min, and flowing phase condition is: 0 min ~ 28 min, 35%A ~ 43%A, 65%B ~ 57%B；28 min ~50
Min, 43%A ~ 60%A, 57%B ~ 30%B.
A length of 25 meters of reaction ring.
First UV-detector detection wavelength is 254 nm；Second UV-detector detection wavelength is 517 nm.
The method of antioxidant content in embodiment 4 one kinds quick detection Radix Et Rhizoma Rhei seed, comprises the following steps:
(1), after Radix Et Rhizoma Rhei seed meal being broken to 60 mesh, obtain Radix Et Rhizoma Rhei seed powder.
By Radix Et Rhizoma Rhei seed powder with the methanol that volumetric concentration is 80% of its quality 80 times under conditions of power is 300W
Supersound extraction 20min, obtains Radix Et Rhizoma Rhei seed extracting solution after filtration.
(3) Radix Et Rhizoma Rhei seed extracting solution separates through high performance liquid chromatography chromatographic column, and the composition of separation first flows through first ultraviolet inspection
As the antioxidant that mass concentration is 100 mg/L pumped into mutually as flowing in sample, this sample and second pump after surveying device
After 1,1-diphenyl-2 trinitrophenyl-hydrazine is derivative in reaction ring, enter in second UV-detector, then by first purple
The chromatogram that external detector and second UV-detector are recorded is compared, when the color that second UV-detector is recorded
When occurring the compound of negative peak in spectrogram, this compound is the antioxidant content in extract.
Wherein:
High-efficiency liquid chromatography method for detecting refers to that mobile phase A is methanol, Mobile phase B volumetric concentration be 0.1% acetic acid water-soluble
Liquid；Flow velocity is 1.0 mL/min, and flowing phase condition is: 0 min ~ 28 min, 35%A ~ 43%A, 65%B ~ 57%B；28 min ~50
Min, 43%A ~ 60%A, 57%B ~ 30%B.
A length of 20 meters of reaction ring.
First UV-detector detection wavelength is 254 nm；Second UV-detector detection wavelength is 517 nm.
The method of antioxidant content in embodiment 5 one kinds quick detection Radix Et Rhizoma Rhei seed, comprises the following steps:
(1), after Radix Et Rhizoma Rhei seed meal being broken to 50 mesh, obtain Radix Et Rhizoma Rhei seed powder.
By Radix Et Rhizoma Rhei seed powder with the methanol that volumetric concentration is 80% of its quality 80 times under conditions of power is 250W
Supersound extraction 45min, obtains Radix Et Rhizoma Rhei seed extracting solution after filtration.
(3) Radix Et Rhizoma Rhei seed extracting solution separates through high performance liquid chromatography chromatographic column, and the composition of separation first flows through first ultraviolet inspection
As the antioxidant that mass concentration is 50 mg/L pumped into mutually as flowing in sample, this sample and second pump after surveying device
After 1,1-diphenyl-2 trinitrophenyl-hydrazine is derivative in reaction ring, enter in second UV-detector, then by first purple
The chromatogram that external detector and second UV-detector are recorded is compared, when the color that second UV-detector is recorded
When occurring the compound of negative peak in spectrogram, this compound is the antioxidant content in extract.
Wherein:
High-efficiency liquid chromatography method for detecting refers to that mobile phase A is methanol, Mobile phase B volumetric concentration be 0.1% acetic acid water-soluble
Liquid；Flow velocity is 0.8 mL/min, and flowing phase condition is: 0 min ~ 28 min, 35%A ~ 43%A, 65%B ~ 57%B；28 min ~50
Min, 43%A ~ 60%A, 57%B ~ 30%B.
A length of 10 meters of reaction ring.
First UV-detector detection wavelength is 254 nm；Second UV-detector detection wavelength is 517 nm.
Claims (1)
1. a method for antioxidant content in quick detection Radix Et Rhizoma Rhei seed, comprises the following steps:
(1), after Radix Et Rhizoma Rhei seed meal being broken to 40 ~ 60 mesh, obtain Radix Et Rhizoma Rhei seed powder；
(2) it is 200 with the methanol that volumetric concentration is 50% ~ 100% of its quality 50 ~ 100 times at power by described Radix Et Rhizoma Rhei seed powder
Supersound extraction 20 ~ 50min under conditions of ~ 300W, obtains Radix Et Rhizoma Rhei seed extracting solution after filtration；
The most described Radix Et Rhizoma Rhei seed extracting solution separates through high performance liquid chromatography chromatographic column, and the composition of separation first flows through first ultraviolet inspection
As the antioxygen that mass concentration is 10 ~ 200 mg/L pumped into mutually as flowing in sample, this sample and second pump after surveying device
Agent 1, after 1-diphenyl-2 trinitrophenyl-hydrazine is derivative in reaction ring, enters in second UV-detector, then by described
The chromatogram that first UV-detector and described second UV-detector are recorded is compared, when described second ultraviolet
When occurring the compound of negative peak in the chromatogram that detector is recorded, this compound is the antioxidant content in extract；High
Effect liquid phase chromatogram detection method refers to that mobile phase A is methanol, and Mobile phase B volumetric concentration is the acetic acid aqueous solution of 0.1%；Flow velocity is
0.6 ~ 1.2 mL/min, flowing phase condition is: 0 min ~ 28 min, 35%A ~ 43%A, 65%B ~ 57%B；28 min ~ 50 min,
43%A ~ 60%A, 57%B ~ 30%B；A length of 5 ~ 25 meters of described reaction ring；Described first UV-detector detection wavelength is
254 nm；Second UV-detector detection wavelength is 517 nm.
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| CN108918437A (en) * | 2018-05-16 | 2018-11-30 | 天津中医药大学 | A kind of the antioxidant activity detection method and application of blood-activating and stasis-removing preparation |
| CN110108808A (en) * | 2019-04-12 | 2019-08-09 | 广西壮族自治区兽医研究所 | Method that is a kind of while evaluating oxidation-resistant active ingredient and content in downy rosemyrtle root |
| CN111423487A (en) * | 2020-03-31 | 2020-07-17 | 东北林业大学 | Method for separating and purifying antioxidant active protein components guided by on-line free radical scavenging method |
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| KR20020037434A (en) * | 2000-11-14 | 2002-05-21 | 최향숙 | Determination method for radical-scavenging activities of foods using 1,1-diphenyl-2-picrylhydrazyl, and method for using of authentic compounds as antioxidant |
| CN102879499A (en) * | 2012-10-11 | 2013-01-16 | 中国科学院西北高原生物研究所 | Method for detecting antioxidant constituents in nitraria fruit juice by using 1,1-diphenyl-2-picrylhydrazyl (DPPH)-high performance liquid chromatography (HPLC) method |
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| JP4338935B2 (en) * | 2002-03-29 | 2009-10-07 | 日本ケフィア株式会社 | Antidiabetic agent using kefir as antioxidant as active ingredient, tissue repair agent having DNA repair action, and method for producing and identifying antioxidant using kefir |
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Patent Citations (2)
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|---|---|---|---|---|
| KR20020037434A (en) * | 2000-11-14 | 2002-05-21 | 최향숙 | Determination method for radical-scavenging activities of foods using 1,1-diphenyl-2-picrylhydrazyl, and method for using of authentic compounds as antioxidant |
| CN102879499A (en) * | 2012-10-11 | 2013-01-16 | 中国科学院西北高原生物研究所 | Method for detecting antioxidant constituents in nitraria fruit juice by using 1,1-diphenyl-2-picrylhydrazyl (DPPH)-high performance liquid chromatography (HPLC) method |
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| HPLC-DPPH在线法筛选黄芩提取物中抗氧化活性成分;王小淞等;《中草药》;20091231;第40卷;第225页左栏第3段、第2.1-2.3部分、图1 * |
| RP—HPLC法同时检测大黄-黄芩药中多种成分的色谱条件研究;胡律江等;《江西中医学院学报》;20101231;第22卷(第6期);第40-43页 * |
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Effective date of registration: 20220725 Address after: No.1, Tongrentang Road, Hexi street, Delingha City, Haixi Mongolian and Tibetan Autonomous Prefecture, Qinghai Province Patentee after: BEIJING TONGRENTANG HEALTH PHARMACEUTICAL (QINGHAI) Co.,Ltd. Address before: 810001 No. 59 Xiguan Street, Xining, Qinghai Patentee before: Northwest Institute of Plateau Biology, Chinese Academy of Sciences Patentee before: QINGHAI ZHONGKE PLATEAU BIOLOGICAL TECHNOLOGY DEVELOPMENT Co.,Ltd. |
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