CN105911174A - Determination method for nuciferine in lotus leaf - Google Patents
Determination method for nuciferine in lotus leaf Download PDFInfo
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- CN105911174A CN105911174A CN201610234643.1A CN201610234643A CN105911174A CN 105911174 A CN105911174 A CN 105911174A CN 201610234643 A CN201610234643 A CN 201610234643A CN 105911174 A CN105911174 A CN 105911174A
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- lotus leaf
- nuciferine
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/04—Preparation or injection of sample to be analysed
- G01N30/06—Preparation
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N2030/022—Column chromatography characterised by the kind of separation mechanism
- G01N2030/027—Liquid chromatography
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/04—Preparation or injection of sample to be analysed
- G01N30/06—Preparation
- G01N2030/062—Preparation extracting sample from raw material
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Abstract
The invention discloses a determination method for nuciferine in lotus leaf. The method comprises the steps of: 1. crushing lotus leaf, and then conducting extraction by ASE extraction method to obtain an extracted solution; 2. diluting and centrifuging the extracted solution to obtain supernatant; and 3. analyzing the supernatant by means of liquid chromatography. The determination method for nuciferine in lotus leaf provided by the invention has the advantages of simple operation and high detection precision.
Description
Technical field
The present invention relates to test in laboratory field, the assay method of Nuciferine in a kind of lotus leaf.
Background technology
Pharmacopeia in 2015 have recorded the extracting method of the Nuciferine in lotus leaf: takes this product meal about 0.5g, accurately weighed, puts tool plug
In conical flask, accurate addition methyl alcohol 50ml, weighed weight, it is heated to reflux 2.5 hours, lets cool, more weighed weight, use methyl alcohol
Supplying the weight of less loss, shake up, filter, precision measures subsequent filtrate 5ml, puts in 10ml measuring bottle, adds water to scale, shakes up,
Obtain.
Liquid chromatographic detection condition is: with octadecylsilane chemically bonded silica as filler；With acetonitrile one water one triethylamine glacial acetic acid
(27:70.6:1.6:0.78) for flowing phase；Detection wavelength is 270nm.Number of theoretical plate is calculated should be not less than by Nuciferine peak
2000。
The method operating process of pharmacopeia is complicated, and the operative skill of operating personnel is required height.
Summary of the invention
It is an object of the invention to provide the assay method of Nuciferine in a kind of lotus leaf, the method is simple to operate, and accuracy of detection is high.
The technical scheme that the present invention provides is: the assay method of Nuciferine in a kind of lotus leaf, comprises the following steps:
Step 1: use ASE extraction that extract is obtained by extraction after being pulverized by lotus leaf；
Step 2: after centrifugal for extract dilution, obtain supernatant；
Step 3: use liquid chromatography to be analyzed supernatant；
Wherein, the analysis condition of liquid chromatography is:
A) instrument: double ternary liquid phase chromatographs
B) chromatographic column C18 5μm 4.6×150mm
C) column temperature: 40 DEG C
D) flow velocity: 0.5mL/min
E) flowing phase: acetonitrile-water-triethylamine-glacial acetic acid (27:70.6:1.6:0.78)
F) detection wavelength: 270nm
In above-mentioned lotus leaf in the assay method of Nuciferine, described step 1 particularly as follows:
S11: after lotus leaf is pulverized, sieve；
S12: weigh 0.5 weight portion mangosteen powder, mixes stand-by with 0.5 weight portion diatomite；
S13: moved into by the mixture in S12 in the abstraction pool putting filter membrane in advance well, adds appropriate diatomite to reaching extraction
The Chi Kou in pond；The lid covering abstraction pool carries out static extracting；Extraction is extracted liquid after terminating；
Extraction conditions is: extractant is methyl alcohol；Extraction temperature 90 DEG C；Static extracting time 5min；Flush volume 100%；
Quiet cycle number of times 2 times.
In above-mentioned lotus leaf in the assay method of Nuciferine, the volume of described abstraction pool is 10ml.
In above-mentioned lotus leaf in the assay method of Nuciferine, the pressure of described static extracting is 1500psi.
In above-mentioned lotus leaf in the assay method of Nuciferine, the purge time of described static extracting is 60s.
In above-mentioned lotus leaf in the assay method of Nuciferine, step 2 particularly as follows: by after extract methanol dilution
Under 15000r/min, centrifugal 5min, takes supernatant.
In above-mentioned lotus leaf in the assay method of Nuciferine, the instrument used by liquid-phase chromatographic analysis is: Thermo U-3000 is double
Ternary liquid phase chromatograph.
In above-mentioned lotus leaf in the assay method of Nuciferine, the instrument that described ASE extraction is used is that ASE350 is quick
Solvent extraction instrument.
The present invention is after using technique scheme, and it has the beneficial effect that
The detection method of the present invention is simple to operate, and precision is consistent with the precision of official method, reproducible.
Accompanying drawing explanation
Fig. 1 is the rectangular plots of the chromatography Nuciferine validation verification of the present invention；
Fig. 2 is the chromatogram of the present invention.
Detailed description of the invention
Below in conjunction with detailed description of the invention, technical scheme is described in further detail, but does not constitute the present invention
Any restriction.
Embodiment 1:
Size-reduced for lotus leaf machine is pulverized, excessively No. three sieves, about 0.5g, accurately weighed, mix with 0.5g diatomite, stand-by,
It is moved into and puts filter membrane in advance wellAdding proper amount of silicon diatomaceous earth in 10ml abstraction pool, shaking is allowed to Chi Kou same gently
On one horizontal line, tighten abstraction pool upper cover.After extraction terminates, extract is shifted in 25ml volumetric flask, with methanol dilution extremely
Scale, under 15000r/min, centrifugal 5min, takes supernatant, enters LC and measures.
Extraction conditions (being shown in Table 1) is:
Table 1
Analysis method is LC liquid chromatography, liquid-phase chromatographic analysis condition:
A) the double ternary liquid phase (U-3000) of instrument: Thermo
B) chromatographic column Eclipse XDB (C185 μm 4.6 × 150mm, i.e. carbon 18 chromatographic column, diameter 4.6mm, length 150mm,
Particle diameter 5 μm)
C) column temperature: 40 DEG C
D) flow velocity: 0.5mL/min
E) flowing phase: acetonitrile-water-triethylamine-glacial acetic acid (volume ratio 27:70.6:1.6:0.78)
F) detection wavelength: 270nm
Chromatography validation verification:
Take Nuciferine reference substance appropriate, accurately weighed, add methyl alcohol and make every 1ml solution containing 16 μ g, to obtain final product.
Take Nuciferine reference substance solution (concentration: 0.1587mg/ml), respectively precision draw this solution 1 μ l, 2 μ l, 4 μ l, 5
μ l, 10 μ l enter LC and measure, and measure according to said method, the results detailed in Table 2.With absolute sample size (μ g)-average
Peak area carries out linear regression, tries to achieve regression equation: y=5.7934x-0.0315, R2=0.9997.Nuciferine is 0.1587～0.9122
In good linear relationship in μ g range, the rectangular plots of this linear relationship is shown in Fig. 1.Using of Nuciferine reference substance is above-mentioned
The chromatogram that liquid chromatography analysis obtains, refers to the collection of illustrative plates being numbered corresponding to S-1 in Fig. 2.
Table 2 Nuciferine linear relationship
The reappearance checking of the extracting process of embodiment 1:
Take sample (lot number: the 150820) 0.5g of identical lot number, totally 6 parts, accurately weighed, extract by ASE extracting method
Need testing solution, sample size is 8 μ l, with above-mentioned chromatographic condition parallel test, records the content of Nuciferine in sample and is shown in Table 2,
RSD is 3.5%, and test shows that ASE extracting method repeatability is good, the results detailed in Table 3.
Table 3
The accuracy test of the extracting process of embodiment 1
Using the ASE extracting process of embodiment 1 and the extracting process of pharmacopeia to extract lotus leaf, lotus leaf is 4 batches, and lot number divides
It is not 1512008,20150501,150820,1508144；
The ASE extracting process of embodiment 1 uses the machine that same model specification is the most different to carry out parallel to a batch of lotus leaf
Testing twice, respectively ASE1 and ASE2 represents.
In Fig. 2, the collection of illustrative plates corresponding to label 114-2-1 is the chromatogram that sample 1508144 obtains according to the method described in pharmacopeia；
Being numbered the collection of illustrative plates corresponding to orthogonal 8-1 in Fig. 2 is that sample 1508144 obtains according to the method described in embodiment 1
Chromatogram.
Concrete test result see table 4；
Table 4
Above-described be only presently preferred embodiments of the present invention, all made in the range of the spirit and principles in the present invention any amendment,
Equivalent and improvement etc., should be included within the scope of the present invention.
Claims (8)
1. the assay method of Nuciferine in a lotus leaf, it is characterised in that comprise the following steps:
Step 1: use ASE extraction that extract is obtained by extraction after being pulverized by lotus leaf；
Step 2: after centrifugal for extract dilution, obtain supernatant；
Step 3: use liquid chromatography to be analyzed supernatant；
Wherein, the analysis condition of liquid chromatography is:
A) instrument: double ternary liquid phase chromatographs
B) chromatographic column C18 5μm 4.6×150mm
C) column temperature: 40 DEG C
D) flow velocity: 0.5mL/min
E) flowing phase: acetonitrile-water-triethylamine-glacial acetic acid (27:70.6:1.6:0.78)
F) detection wavelength: 270nm.
The assay method of Nuciferine in lotus leaf the most according to claim 1, it is characterised in that described step 1 particularly as follows:
S11: after lotus leaf is pulverized, sieve；
S12: weigh 0.5 weight portion mangosteen powder, mixes stand-by with 0.5 weight portion diatomite；
S13: moved into by the mixture in S12 and put well in advance in the abstraction pool of filter membrane, adds appropriate diatomite to the Chi Kou reaching abstraction pool；The lid covering abstraction pool carries out static extracting；Extraction is extracted liquid after terminating；
Extraction conditions is: extractant is methyl alcohol；Extraction temperature 90 DEG C；Static extracting time 5min；Flush volume 100%；Quiet cycle number of times 2 times.
The assay method of Nuciferine in lotus leaf the most according to claim 2, it is characterised in that the volume of described abstraction pool is 10ml.
The assay method of Nuciferine in lotus leaf the most according to claim 2, it is characterised in that the pressure of described static extracting is 1500psi.
The assay method of Nuciferine in lotus leaf the most according to claim 2, it is characterised in that the purge time of described static extracting is 60s.
The assay method of Nuciferine in lotus leaf the most according to claim 1, it is characterised in that step 2 is particularly as follows: by being centrifuged 5min after extract methanol dilution under 15000r/min, take supernatant.
The assay method of Nuciferine in lotus leaf the most according to claim 1, it is characterised in that the instrument used by liquid-phase chromatographic analysis is: the double ternary liquid phase chromatograph of Thermo U-3000.
The assay method of Nuciferine in lotus leaf the most according to claim 1, it is characterised in that the instrument that described ASE extraction is used is ASE350 Accelerate solvent extraction instrument.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108680664A (en) * | 2018-04-28 | 2018-10-19 | 安徽粮食工程职业学院 | A kind of method of quick measurement Nuciferine purity |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102258578A (en) * | 2010-05-25 | 2011-11-30 | 天津中医药大学 | Novel lotus leaf total alkaloid extract and preparation method as well as application thereof |
| CN103494178A (en) * | 2013-09-30 | 2014-01-08 | 浙江工业大学 | Method for extracting lentinus edodes total alkaloid |
| CN103830374A (en) * | 2014-03-06 | 2014-06-04 | 天津中医药大学 | Application of three-leaf glycolipid-removal medicine in hyperuricemia |
| WO2015152577A3 (en) * | 2014-04-02 | 2015-11-26 | 연세대학교 산학협력단 | Composition for preventing and treating bone disease |
-
2016
- 2016-04-15 CN CN201610234643.1A patent/CN105911174A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102258578A (en) * | 2010-05-25 | 2011-11-30 | 天津中医药大学 | Novel lotus leaf total alkaloid extract and preparation method as well as application thereof |
| CN103494178A (en) * | 2013-09-30 | 2014-01-08 | 浙江工业大学 | Method for extracting lentinus edodes total alkaloid |
| CN103830374A (en) * | 2014-03-06 | 2014-06-04 | 天津中医药大学 | Application of three-leaf glycolipid-removal medicine in hyperuricemia |
| WO2015152577A3 (en) * | 2014-04-02 | 2015-11-26 | 연세대학교 산학협력단 | Composition for preventing and treating bone disease |
Non-Patent Citations (4)
| Title |
|---|
| FENG LIU 等: "Cloud Point Extraction and Pre-Concentration of Four Alkaloids in Nelumbo nucifera Leaves by Ultrahigh Pressure-Assisted Extraction", 《SEPARATION SCIENCE AND TECHNOLOGY》 * |
| 国家药典委员会 编: "《中华人民共和国药典 2015年版 四部》", 30 June 2015, 北京：中国医药科技出版社 * |
| 徐榕青: "《福建道地药材现代研究》", 31 October 2014, 福州：福建科学技术出版社 * |
| 李文龙 等: "加速溶剂萃取-高效液相色谱-电喷雾飞行时间质谱联用分析莲子心中生物碱", 《分析化学》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108680664A (en) * | 2018-04-28 | 2018-10-19 | 安徽粮食工程职业学院 | A kind of method of quick measurement Nuciferine purity |
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