CN103076294A - Method for testing content of alkaloid in dendrobe - Google Patents
Method for testing content of alkaloid in dendrobe Download PDFInfo
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- CN103076294A CN103076294A CN2012105914791A CN201210591479A CN103076294A CN 103076294 A CN103076294 A CN 103076294A CN 2012105914791 A CN2012105914791 A CN 2012105914791A CN 201210591479 A CN201210591479 A CN 201210591479A CN 103076294 A CN103076294 A CN 103076294A
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- chloroform
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Abstract
The invention discloses a method for testing the content of alkaloid in dendrobe. After a dendrobe dried product is crushed, alkaloid is extracted through alkaline chloroform water circumfluence, a nonionic surfactant Tween-80 is added into test liquid, and the content of alkaloid in the dendrobe is tested through bromocresol green acid dye colorimetry. In the system for testing alkaloid in dendrobe, a proper amount of Tween-80 is added to enable the extraction efficiency of bromocresol green-chloroform to be improved, so that the sensitivity in testing alkaloid in dendrobe is improved, and quick layering, simplicity and convenience in operation, good repeatability and stable testing result are achieved.
Description
Technical field
The present invention relates to the spectrophotometric colorimetric determination and measure the content of material field, be specifically related to a kind of method of measuring stem of noble dendrobium alkaloid.
Background technology
Alkaloid is that a class is present in the nitrogenous base organic matter that has physiologically active in the biosome.Plant alkaloid is the large class secondary metabolite that plant forms for the attack of resisting animal, microorganism, virus and other plant in the long-term ecologic environment procedure of adaptation.Alkaloid is significant in growth, growth, symbiosis and the reproductive process of medicine, plant.China contained alkaloidal Chinese herbal medicine with regard to use before 2000 or three thousand years treats, and just records the multiple alkaloidal Chinese herbal medicine that contains at Shennong's Herbal, such as the top grade coptis, the stem of noble dendrobium.Stem of noble dendrobium alkaloid is one of most important active component in the stem of noble dendrobium medicinal material, also is the compound that separates the earliest and carry out structural confirmation.The alkaloidal content of the stem of noble dendrobium is mainly with the bromcresol green Wan withAcid-dye Colorimetry, and this is a kind of indirect extraction colorimetric method.Because the Dendrobium Sw alkaloid is generally lower, it is lower that the bromcresol green Wan withAcid-dye Colorimetry of commonly using at present draws rate, and measure consuming time longer.
Summary of the invention
It is low to draw rate for bromcresol green Wan withAcid-dye Colorimetry stem of noble dendrobium alkaloid commonly used at present, and measure the problem of length consuming time, the purpose of this invention is to provide a kind of modification method of measuring stem of noble dendrobium alkaloid, the sensitivity of measuring to improve stem of noble dendrobium alkaloid, and the energy fast hierarchical, shorten minute.
The present invention realizes that goal of the invention adopts following technical scheme:
A kind of method of measuring stem of noble dendrobium alkaloid take stem of noble dendrobium dry product as raw material, may further comprise the steps:
(1) stem of noble dendrobium dry product is clayed into power to get substance A;
(2) get substance A 0.2g, with 10～15 times of weight volumetric concentrations be the strong aqua of 25-28% wetting after in round-bottomed flask close plug 40 min, add chloroform 10 mL, weigh, round-bottomed flask accesses condenser pipe, puts to add hot reflux 2 h in 80 ℃ of water-baths, replenishes chloroform after the cooling to original weight, qualitative filter paper filters, and gets subsequent filtrate B;
(3) the subsequent filtrate B that step (2) is obtained adds chloroform and is settled to 10 mL, 2M Potassium Hydrogen Phthalate-sodium hydrate buffer solution 5 mL and volumetric concentration 0.04% bromcresol green solution 1 mL that add pH4.5, and Tween-80 aqueous solution 0.4 mL of interpolation volume ratio 1:60, jolting 3 min on oscillator, leave standstill 10 min, qualitative filter paper filters chloroform layer, gets subsequent filtrate C;
(4) the subsequent filtrate C that step (3) is obtained adds 0.01mol/L NaOH-ethanol solution 1 mL, shakes up, and measures absorbance in 620nm wavelength place; The alkaloid computing formula is as follows:
The present invention compared with the prior art, its beneficial effect is embodied in:
The present invention with alkaline chloroform water-bath refluxing extraction alkaloid, measures in the liquid and adds non-ionic surfactant Tween-80, bromcresol green Wan withAcid-dye Colorimetry stem of noble dendrobium alkaloid after stem of noble dendrobium dry product to be measured is pulverized.The present invention adds an amount of Tween-80 in stem of noble dendrobium alkaloid mensuration system, mainly be to utilize it to extracting solvent effect and the effective constituent generation solubilization to extracting that promotes infiltration to be arranged.Under the appropriate pH condition, the kation RH of stem of noble dendrobium alkali salt
+With bromcresol green negative ion In
-The ionic associate RH that is combined into
+In
-Quantitatively by chloroform extraction, add non-ionic surfactant Tween-80, anionic dye In
-By solubilising, changed that each material has increased bromcresol green-chloroform extraction rate in the distribution of two-phase in the system, improved the sensitivity that stem of noble dendrobium alkaloid is measured, and can fast hierarchical, shorten minute.The method is easy and simple to handle, favorable reproducibility, and measurement result is stable.
Description of drawings
Fig. 1 is assay method schematic flow sheet of the present invention.
Embodiment
Below the present invention is described further by embodiment.
Embodiment 1 gets the stem of noble dendrobium dry product of 0.2g, clay into power, wetting rear in round-bottomed flask Nei Misai 40 min with the strong aqua of 10～15 times of weight volumes, add chloroform 10 mL, weigh, round-bottomed flask access condenser pipe, put and add hot reflux 2 h in 80 ℃ of water-baths, the round-bottomed flask phegma replenishes chloroform to original weight (because adding the hot reflux chloroform evaporated) after being cooled to room temperature, and qualitative filter paper filters, and subsequent filtrate adds chloroform and is settled to 10 mL.0.2M Potassium Hydrogen Phthalate-the sodium hydrate buffer solution (being the even 0.2M of being of potassium hydrogen phthalate and naoh concentration) 5 mL and volumetric concentration 0.04% bromine cresols solutions of chlorine 1 mL that add pH4.5 in the filtrate, and add the Tween-80 aqueous solution (Tween-80 of 1 volume: 60 volume water) of 0.4 mL volume ratio 1:60, jolting 3 min on oscillator, leave standstill 10 min, qualitative filter paper filters chloroform layer, subsequent filtrate adds massfraction 0.01mol/L NaOH-ethanol solution 1 mL, shake up, do blank with the same operation of chloroform 10 mL, measure absorbance in 620nm wavelength place.Whole assay method step as shown in Figure 1.Test findings is as shown in table 1.
Table 1 tween is on the impact of stem of noble dendrobium protocorms alkaloid content determination
As shown in Table 1, the absorbance that adds Tween-80 in the alkaloid content determination system obviously increases.
Embodiment 2, and method of operating is with embodiment 1, and difference is for measuring the tween addition to the impact of stem of noble dendrobium protocorms alkaloid content determination by the Tween-80 that adds different volumes, and the result is as shown in table 2.
Table 2 tween addition is on the impact of stem of noble dendrobium protocorms alkaloid content determination
As shown in Table 2, the 1:60 Tween-80 aqueous solution of Different adding amount is to tween alkaloid content determination system
Have a significant effect, wherein the addition of 0.4 mL is the strongest to the effect of enhanced sensitivity of stem of noble dendrobium alkaloid content determination system.
Take the alkaloids extract volume as horizontal ordinate, the absorbance that 620nm wavelength place measures is ordinate, and it is as follows to get regression equation:
y
1 = 0.0685x + 0.0063 R
2 = 0.9996
y
2 = 0.0813x + 0.024 R
2 = 0.9905
y
1, y
2Expression does not add the regression equation that tween is measured system and added tween mensuration system respectively, and x represents the alkaloids extract volume in the following formula, the absorbance that the y representative is measured, R
2Represent related coefficient.y
2The regression equation slope is than y
1Increase nearly 1/5.
The alkaloid computing formula is as follows:
Wherein alkaloid mark product are dendrobine, and alkaloid mark product weigh 20 μ g, and alkaloid mark product absorbance is 0.174, the heavy 0.2g of sample.
Claims (1)
1. a method of measuring stem of noble dendrobium alkaloid is characterized in that, take stem of noble dendrobium dry product as raw material, may further comprise the steps:
(1) stem of noble dendrobium dry product is clayed into power to get substance A;
(2) get substance A 0.2g, with 10～15 times of weight volumetric concentrations be the strong aqua of 25-28% wetting after in round-bottomed flask close plug 40 min, add chloroform 10 mL, weigh, round-bottomed flask accesses condenser pipe, puts to add hot reflux 2 h in 80 ℃ of water-baths, replenishes chloroform after the cooling to original weight, qualitative filter paper filters, and gets subsequent filtrate B;
(3) the subsequent filtrate B that step (2) is obtained adds chloroform and is settled to 10 mL, 2M Potassium Hydrogen Phthalate-sodium hydrate buffer solution 5 mL and volumetric concentration 0.04% bromcresol green solution 1 mL that add pH4.5, and Tween-80 aqueous solution 0.4 mL of interpolation volume ratio 1:60, jolting 3 min on oscillator, leave standstill 10 min, filter chloroform layer, get subsequent filtrate C;
(4) the subsequent filtrate C that step (3) is obtained adds 0.01mol/L NaOH-ethanol solution 1 mL, shakes up, and measures absorbance in 620nm wavelength place; The alkaloid computing formula is as follows:
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103604877A (en) * | 2013-10-29 | 2014-02-26 | 安徽农业大学 | Construction method and applications of high performance liquid chromatography (HPLC) fingerprint of alkaloid compounds of dendrobe |
| CN106501437A (en) * | 2016-12-27 | 2017-03-15 | 安徽农业大学 | Based on the method that UPLC/LTQ Orbitrap Velos MS detect stem of noble dendrobium alkaloid component |
| CN107764756A (en) * | 2017-09-11 | 2018-03-06 | 云南中烟工业有限责任公司 | A kind of method of dispersive liquid-liquid microextraction combination photometry measure nicotine |
| CN108645802A (en) * | 2018-05-16 | 2018-10-12 | 安徽农业大学 | A kind of detection method of dendrobium candidum total alkaloid content |
| CN109991213A (en) * | 2019-01-11 | 2019-07-09 | 贵州大学 | A kind of uncaria alkaloid detection method based on Histochemical localization and chemical composition analysis |
| CN113332383A (en) * | 2021-03-05 | 2021-09-03 | 杭州蚕宝生物技术有限公司 | Method for efficiently extracting alkaloid from dendrobium officinale |
-
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103604877A (en) * | 2013-10-29 | 2014-02-26 | 安徽农业大学 | Construction method and applications of high performance liquid chromatography (HPLC) fingerprint of alkaloid compounds of dendrobe |
| CN103604877B (en) * | 2013-10-29 | 2015-09-30 | 安徽农业大学 | A kind of efficient liquid-phase chromatograph finger print atlas construction method of stem of noble dendrobium alkaloids composition and application |
| CN106501437A (en) * | 2016-12-27 | 2017-03-15 | 安徽农业大学 | Based on the method that UPLC/LTQ Orbitrap Velos MS detect stem of noble dendrobium alkaloid component |
| CN107764756A (en) * | 2017-09-11 | 2018-03-06 | 云南中烟工业有限责任公司 | A kind of method of dispersive liquid-liquid microextraction combination photometry measure nicotine |
| CN108645802A (en) * | 2018-05-16 | 2018-10-12 | 安徽农业大学 | A kind of detection method of dendrobium candidum total alkaloid content |
| CN109991213A (en) * | 2019-01-11 | 2019-07-09 | 贵州大学 | A kind of uncaria alkaloid detection method based on Histochemical localization and chemical composition analysis |
| CN113332383A (en) * | 2021-03-05 | 2021-09-03 | 杭州蚕宝生物技术有限公司 | Method for efficiently extracting alkaloid from dendrobium officinale |
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