water alcohol precipitation
microwave extraction
acid extraction
base extraction
enzymatic decomposition
Ultrafiltration
ultrasound
Supercritical Fluid Extraction
There are too many methods, give you one to tell the most important one. p>There are too many methods, so I'll give you the simplest one! The data are complete
Extraction and content determination of polysaccharides from Mushrooms
Xu Yajie, Ningbo, Wang Lei, Zhang Jinyu, * Lu Jiafu
(Changchun Vocational College of Technology, Changchun 130033, Jilin)
Abstract: The extraction of polysaccharides from Mushrooms by hot water at a temperature of 70 ℃, deproteinization using sevag method, and graded sedimentation using ethanol, and the extraction by sulfuric acid and sulfuric acid using a supercritical fluid extraction method, are investigated. The method of extracting polysaccharides from the mushroom by hot water extraction at 70 ℃, removing proteins by sevag method and ethanol graded precipitation was investigated, and the content of polysaccharides was determined by sulfuric acid-anthrone method. The total yield of polysaccharide was 0.11% of the wet mass of Agaricus blazei, and the linear range of polysaccharide was 1~16 μg/mL. The RSD of polysaccharide content of Agaricus blazei was less than 2.24%, and the recoveries were 97.6%~102.8%.
Keywords: Sulfuric acid-anthracene ketone method; Polysaccharide from Flammulina Versicolor; Extraction; Content Determination
Chinese Classification Number: R284.2; O629.12 Literature Symbol Code: A
Extraction and Content Determination of Polysaccharide from
Flammulina Versicolor. p>Flammulina Velutipes
Xu Yajie, Ning Bo, Wang Lei, Zhang Jinyu, *Lu Jiafu
( Changchun Vocational Institute of Technology College, Changchun 130033, China). Changchun 130033, China)
Abstract: This paper reports the polysaccharide extraction and determination from flammunlina velutipes, the process is: extract
the polysaccharides from flammunlina velutipes by 70 ℃ water at first, then deproteinize according to the method of Sevag,
and then graft the polysaccharides on the surface of the plant. p>
and then grading precipitate by ethanol. Quantitatively determine the polysaccharides content by anthrone- sulfuric acid
method. In this method, the total yield up to 0.11%, the linearly range is 1~16 μg/mL, the RSD is not more than 2.24%,
and the range of recovery is 97.6~102.8%.
Key words: anthrone- sulfuric acid method; polysaccharides; flammunlina velutipes; extraction
0 INTRODUCTION
The golden mushroom (Flammunlina velutipes) is a member of the Stachybotrys species. Flammunlina velutipes is a genus of fungi in the order of Stramenopiles, and is a kind of fungus used for both medicine and food. Its fruiting body contains protein
31.2%, fat 5.8%, as well as VB1, VB2, VC, VPP (biotin
) and more than 10 kinds of amino acids.
At present, enoki mushrooms have been widely used in food and flavorings industry
and are only consumed after mushrooms and shiitake mushrooms in the international market. The main bioactive component of the mushroom, polysaccharide, has been widely
regarded for its tumor inhibition, anticancer, and immune-enhancing effects[1~3].
In this experiment, the crude polysaccharide of Agaricus blazei was prepared by hydrotropic alcohol precipitation [4], and the polysaccharide content was determined by phenol-sulfuric acid colorimetric method, which obtained
good results. The results showed that the method is simple, rapid,
highly sensitive and reproducible.
1 Materials and methods
1.1 Materials and instruments
1.1.1 Materials and reagents
Golden Needle Mushroom, Beijing Guanhua Agricultural Co.
Standard glucose solution (100.0 μg/mL) :
Accurately weighed 10.0 mg of glucose dried at 105 ℃ to a constant mass, added appropriate amount of distilled water to dissolve, quantitatively transferred to a volumetric flask of 100 mL
, and shaken well.
Anthrone (concentration 2 mg/mL): weigh 10.0 mg of anthrone, add appropriate amount of concentrated sulfuric acid to dissolve, quantitatively transfer to
100 mL of black paper wrapped in brown volumetric flasks, fixed to the scale,
shaking well, ready to use.
Anhydrous ethanol, nitrogen simulation, n-butanol, concentrated sulfuric acid. All the above reagents are analytically pure.
1.1.2 Instruments
UV-2401 ultraviolet-visible spectrophotometer, Shimadzu, Japan
products; 3-18K high-speed freezing centrifuge, Sigma, Germany
products; tissue masher, Changzhou Guohua Electric Appliance Co. Ltd.; Electric
Heat Blast Drying Box, Shanghai Experimental Instrument Factory; GSY-II type constant
temperature water bath box, Beijing Medical Equipment Factory.
1.2 Experimental method
Accurately weighing 100.00 g of enoki mushrooms, refluxing 300 mL of ethyl acetate at 80 ℃ for 2 h, degreasing, adding 300 mL of distilled
water, mashing with a tissue masher, and then adding 400 mL of distilled water, the sample was heated at a temperature of 70 ℃ in the
No.9, 2007
constant temperature water bath. >The supernatant was collected by centrifugation at 4000 r/min for 5 min, and then extracted in a thermostatic water bath at 70 ℃ for 4 h. The supernatant was then extracted by the sevag method [1, 2, 3]. The extract was mixed with an equal volume of 4:1 chloroform and n-butanol by the sevag method[5], and then concentrated to 100 mL under reduced pressure with a rotary evaporator, and the polysaccharide was precipitated with 95% ethanol at 1 times the volume
integral amount, and then centrifuged, and the supernatant was precipitated with 95% ethanol at 2 times the volume
integral amount, Centrifugation, the supernatant was precipitated with 4 times the volume of 95% ethanol, and the collected polysaccharide precipitates were dissolved in 1 000
mL volumetric flasks.
2 Experimental principle
The polysaccharide of mushroom can be dehydrated under the action of concentrated sulfuric acid to form glyceraldehyde derivatives
organisms, and the latter then interact with anthrone to form blue compounds[6], which can be colorimetrically quantified with the standard glucose solution of the same method
processed.
3 Results and Discussion
3.1 Preparation of glucose standard curve
The standard glucose solution of 0.05, 0.10, 0.20,
0.30, 0.40, 0.60, 0.80 mL was sucked into the stoppered cuvette, and then diluted with distilled water to 1.0 mL, and then shaken well. Then 4.0 mL of anthrone test
agent was added, and the tubes were quickly immersed in an ice-water bath and cooled by shaking, and then all tubes were immersed in a boiling-water bath for 10 min, and then cooled by cold water, and left at room temperature for 10 min. The glucose concentration C was used as the horizontal coordinate, and the absorbance value A620 was used as the vertical coordinate to obtain the glucose standard curve.
The glucose standard curve is shown in Figure 1.
The regression equation was obtained as:
A620=0.035 08 C+0.007 41, and the correlation coefficient was r=0.999 13.
The experiments showed that, at the glucose mass concentration of 1-16 μg/mL
, there is a good linear correlation between the absorbance value and the glucose mass concentration.
Experiments showed that at glucose mass concentration of 1~16 μg/mL
, the absorbance values showed a good linear relationship with glucose mass concentration.
3.2 Selection of reaction time and stability
Pipetting up 1.0 mL of the test solution, adding 4.0 mL of anthrone reagent at a mass concentration of 2 mg/mL
, and operating in the same way as in the determination of the standard curve, the absorbance was measured every 20 min. The results showed that the absorbance value of the assay solution was stable within
200 min with RSD=2.64% (n=11).
3.3 Precision test
Precision pipetting control solution 0.5 mL ***5 copies, according to the determination of the standard
curve the same method to measure its absorbance value, calculated that the RSD is less than
2.24%.
3.4 Determination of sample content
Accurately absorb 0.5 mL of sample solution, according to the same method as the standard curve
, the content of polysaccharide was calculated to be
0.11% of the wet mass of enoki mushroom.
3.5 Recovery test
Precisely sucked up 6 samples of 0.5 mL of sample solution, and added a certain amount of
control solution to make the total volume of 1.0 mL, and then measured the absorbance according to the same method of standard curve, and obtained the average recoveries of
97.6%~102.8%.
The results of the recovery experiments are shown in Table 1.
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