1、 Pharmacopoeia standard of Lycium barbarum
This product is the dried and mature fruit of Lycium barbarum L., a Solanaceae plant. .
. The peel is flexible and wrinkled; The flesh is fleshy and soft. The seeds are 20 ～ 50, kidney like, flat and upturned, 1.5 ～ 1.9mm long and 1 ～ 1.7mm wide, with light yellow or brownish yellow surface. It is light and sweet.
[identification] (1) the powder of this product is yellow orange or reddish brown. The surface of epicarp epidermal cells appeared to be polygonal or long polygonal, with straight or wavy vertical walls, and parallel cuticular stripes on the surface of the peripheral wall. . The surface of seed coat stone cells is irregular and polygonal, with thick walls, wavy bends, and clear laminations.
(2) Take 0.5g of this product, add 35ml of water, heat and boil for 15 minutes, cool it, filter it, extract the filtrate with 15ml of ethyl acetate by shaking, separate the ethyl acetate solution, and concentrate it to 1ml as the test solution. Another 0.5g of Lycium barbarum reference material was prepared into the reference material solution by the same method. According to the test of thin-layer chromatography (general rule 0502), suck 5 μ l of each of the above two solutions, dot them on the same silica gel G thin-layer plate, use ethyl acetate chloroform formic acid (3:2:1) as the developing agent, develop, take out, dry, and view under the ultraviolet light (365nm). In the chromatogram of the test sample, fluorescent spots of the same color appear at the position corresponding to the chromatogram of the control medicinal material.
[inspection] the moisture content shall not exceed 13.0% (the second method of general rule 0832, the temperature is 80 ℃).
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; Cadmium shall not exceed 0.3mg/kg; Arsenic shall not exceed 2mg/kg; Mercury shall not exceed 0.2mg/kg; Copper shall not exceed 20mg/kg.
[extract] according to the hot leaching method under the determination method of water-soluble extract (general rule 2201), it shall not be less than 55.0%.
[content determination] preparation of Lycium barbarum polysaccharide reference solution take 25mg of anhydrous glucose reference solution, accurately weigh it, put it into a 250ml volumetric flask, add appropriate amount of water to dissolve it, dilute it to the scale, and shake it evenly to obtain (0.1mg of anhydrous glucose per 1ml).
Preparation of standard curve accurately measure 0.2ml, 0.4ml, 0.6ml, 0.8ml, 1.0ml of the reference solution, place them in a plugged test tube, add water to 2.0ml respectively, accurately add 1ml of 5% phenol solution each, shake well, quickly and accurately add 5ml of sulfuric acid, shake well, place for 10 minutes, place them in a 40 ℃ water bath for 15 minutes, take them out, quickly cool them to room temperature, take the corresponding reagent as blank, according to the UV visible spectrophotometry (general rule 0401), measure the absorbance at the wavelength of 490nm, and draw the standard curve with absorbance as the ordinate and concentration as the abscissa.
Determination method take about 0.5g of crude powder of this product, weigh accurately, add 100ml of ether, heat and reflux for 1 hour, stand, cool, carefully discard the ether solution, and put the residue on the water bath to volatilize the ether. Add 100ml of 80% ethanol, heat and reflux for 1 hour, filter while hot, wash the filter residue and filter with 30ml of hot 80% ethanol in several times, put the filter residue and filter paper into a flask, add 150ml of water, heat and reflux for 2 hours. Filter while it is hot, wash the filter with a small amount of hot water, combine the filtrate and washing solution, cool it, transfer it to a 250ml volumetric flask, dilute it with water to the scale, shake it well, accurately measure 1ml, place it in a stoppered test tube, add 1.0ml of water, according to the method under the preparation of the standard curve, measure the absorbance according to the law from "1ml of each 5% phenol solution accurately added", read the weight (mg) of glucose in the test solution from the standard curve, and calculate.
This product is calculated as dry product, and the content of Lycium barbarum polysaccharide is calculated as glucose (C6H12O6), which shall not be less than 1.8%.
Cut the betaine into pieces, take about 2G, weigh accurately, add 50ml of 80% methanol, heat and reflux for 1 hour, cool, filter, wash the residue and filter with 30ml of 80% methanol in several times, combine the washing solution and filtrate, concentrate to 10ml, adjust the pH value to 1 with hydrochloric acid, add 1g of activated carbon, heat and boil, cool, filter, wash with 15ml of water in several times, combine the washing solution and filtrate, add 20ml of newly prepared 2.5% ammonium chromium thiocyanate solution, mix well, and place under 10 ℃ for 3 hours. Filter with G4 vertical melting funnel, wash the sediment with a small amount of ice water, pump it dry, add acetone to dissolve the residue, transfer it to a 5ml volumetric flask, add acetone to the scale, shake well, and use it as the test solution. In addition, take an appropriate amount of betaine reference substance, weigh accurately, add hydrochloric acid methanol solution (0.5 → 100) to make a solution containing 4mg per 1ml as the reference substance solution. According to the test of thin-layer chromatography (general rule 0502), accurately suck 5 μ l of the test solution and 3 μ L and 6 μ l of the control solution, respectively cross on the same silica gel G thin-layer plate, use acetone absolute ethanol hydrochloric acid (10:6:1) as the developing agent, pre saturate for 30 minutes, develop, take out, volatilize the solvent, spray the newly prepared modified bismuth potassium iodide test solution immediately, place it for 1-3 hours until the spots are clear, scan according to the thin-layer chromatography (general rule 0502), wavelength: λ s=515nm, λ r=590nm, measure the absorbance integral value of the test sample and the absorbance integral value of the control, calculate, namely get.
The content of Betaine (C5H11NO2) shall not be less than 0.30% according to the dry product.
[nature, taste and meridian tropism] sweet and smooth. Return to liver and kidney channels.
[functions and indications] nourishing liver and kidney, benefiting essence and eyesight. It is used for asthenia and asthenia, aching waist and knees, dizziness and tinnitus, impotence and spermatorrhea, internal heat and thirst, blood deficiency and yellowing, and blurred vision.
[usage and setting] 6 ～ 12g.
[storage] store in a cool and dry place to prevent heat, moisture and moth.

2、 Chemical constituents of Fructus Lycii

The mature fruit of Lycium barbarum L. in Ningxia containsbetaine（betane)， Atropine, gyoscyamine. It also contains zeaxanthin, phycoerythrin, cryptoxanthin,Scopoletin Pavilion，Beta carotene,Alpha carotene, thiamine, riboflavin, niacin, vitamin C. seeds contain amino acids:L-aspartic acid ，L-Proline ，L-alanine，l-leucine ，L-phenylalanine，l-serine ，glycine, glutamate, cysteine,L-lysine，L-arginine，L-isoleucine，L-threonine，l-his , coumarin,L-tryptophan，L-Methionine It also contains potassium, calcium, sodium, zinc, iron, copper, chromium, strontium, lead, nickel, cadmium, cobalt, magnesium and other elements. It also contains polysaccharide with immune promoting effect, and the content is 7.09%. It also contains taurine and γ - aminobutyric acid.
The mature fruit of Lycium barbarum, a plant of the same genus, contains betaine, zeaxanthin, physalien, and amino acids:l-leucine ，L-valine ，L-Proline ，L-alanine，l-tyrosine ，L-glutamine，glycine, glutamate,L-aspartic acid ，L-asparagine，l-his ，L-tryptophanIt also contains volatile components, mainly safranal, β -, 3-hydroxy - β -ionone, 1,2-dehydrox - α -cyperene and solavetivone. It was also reported that 100g fruit contained 3.39mg carotene, 0.23mg thiamine, 0.33mg riboflavin, 1.7mg niacin, and 0.0mg vitamin C3. The seeds contain oil, and the non saponified part contains a variety of sterols: cholesterols, 7-cholesterols, which belong to 4-demethylsterols,Campesterol, 24 methylenecholesterol, isofucosterol,Stigmasterol，β - sitosterol, 24-lhnhad -5, 24 cholestanol, 24 ethyl cholestanol, cholestanol, 24 methylcholestanol, 24 ethyl-22-cholestanol; Among the 4-methylsterols, there are 31 demethylcyclocarvacrol, 31 demethylcyclocarvacrol, cycloeucalyptol, 31 demethyl-8-lanosterol, 4 α, 14 α, 24 trimethyl-8, 24 cholesterol, 31 demethyl-9 (11) - lanosterol, 4 α - methyl, - 8-cholesterol, 4-methyl-7-cholesterol, 4-methyl-24 ethyl-7-cholesterol, 4,24-dimethyl-cholesterol, graminesterol, limosterol dienol, 4 α, 24 methyl 7, 24 cholesteradienol, 4 α - methylethyl-7, 24 cholesteradienol); 4,4-dimethylsterols include 8-lanostenol, cyclocarvacrol, cyclocarvacrol, lanosterol, β - geraniol,Lupeol, 24-methylene-8-lanosteenol, 24 methylenecycloligninol, etc. In addition, it contains polysaccharide with immune promoting effect, with a content of 5.42%-8.23%[6]. It also contains taurine and γ - aminobutyric acid

3、 Pharmacological effects of Fructus Lycii

1. impact on immune function:

1.1. effects of Lycium barbarum polysaccharide on 3H TDR incorporation into mouse splenic lymphocytes and TS cells:
1.1.1. effect on 3H TDR incorporation into human splenic lymphocytes: Lycium barbarum polysaccharide (LBP) 5mg/kg and 10mg/ (kg. day) × 7 days, intraperitoneal injection can significantly increase the value of 3H TDR incorporation into splenic T cells under ConA induction, and 10mg/kg can make the incorporation index reach the peak, which is 3 times higher than the control group. . The results showed that the proliferation effect of LBP on splenic T lymphocytes was closely related to the dose. The most significant dose of LBP to increase the transformation value of splenic T lymphocytes was 10mg / (kg · day) × 7 days, and the effect on 3H TDR incorporation into cells induced by LPS, a mitotic agent for B lymphocytes, was observed. At LPS concentrations of 50.70 and 100 μ g/ml, LBP had no significant effect on the incorporation index of 3H TDR into B cells, indicating that LBP was not sensitive to B cell mitogen induced cell transformation.

1.1.2. effect on the number of spleen antibody forming cells: LBP at 5.20mg/ (kg. day) × 7 days can significantly increase the number of spleen PFC, and 5mg/kg dose can make the PFC number reach 1.4 times that of the normal immune control group. At 25.50mg/kg, the PFC number was significantly reduced, and the PFC number in the 25mg/kg group was 43% of that in the normal control group. The results showed that LBP had a suitable dose range for enhancing the number of antibody forming cells.

1.1.3. effect on the function of splenic TS cells: it was observed by using the method of inducing TS cells with an excessive amount of SRBC immunization (SOI). TS cells in splenocytes of donor mice can inhibit the PFC number of normal immunized mice by 63%, indicating that the SOI method can induce TS cells that have a significant inhibitory effect on antibody producing cells. After giving donor mice lbp5mg/ (kg. day) × 7 days, the PFC number of recipient mice decreased significantly compared with that of SOI control group, indicating that this dose can enhance the function of TS cells. When the dose was increased to 25 and 50mg/kg, the enhancing effect of LBP on the function of TS cells decreased significantly.

1.2. effect of Lycium barbarum polysaccharide on cellular immune function of S180 tumor bearing mice and its antitumor effect: 1.2.1. effect of Lycium barbarum polysaccharide on cellular immune function of normal mice: mice were injected intraperitoneally with lbp5mg/ (kg. day) for 7 consecutive days (do-6), and the spleen was taken on the seventh day to measure t lymphocyte proliferation reaction. The results showed that LBP could significantly improve the splenic T lymphocyte proliferation induced by ConA in normal mice, and its CPM value increased from 28410 ± 3110 in the control group to 64870 ± 2571 in the administration group.

1.2.2. effect of Lycium barbarum polysaccharide on cellular immune function of S180 tumor bearing mice and its antitumor effect: two days after subcutaneous inoculation of S180 tumor cells, mice were intraperitoneally injected with lbp10mg or 20mg/ (kg. day) × 7 days, and the mice were sacrificed on the 9th day, and tumor weight and spleen T lymphocyte proliferation reaction were measured. The results showed that the tumor inhibition rates of lbp10mg and 20mg/kg were 31% and 39%, respectively. At the same time, it can be seen that the CPM value of T lymphocyte proliferation reaction in lbp10mg group increased from 139 ± 62 in the non administration group to 5581 ± 783 in the administration group, and the RPI (relative proliferation index) increased from 0.3% equivalent to normal to 24.6%. Cyclophosphamide (CY) was used as a positive control in the experiment. After one subcutaneous injection of 25mg/kg, the tumor inhibition rate was 47%, and the CMP value of T lymphocyte proliferation reaction was 165 ± 31.

1.2.3. effect of Lycium barbarum polysaccharide on cellular immune function of normal mice inhibited by cyclophosphamide: mice were injected subcutaneously with cy25mg/kg and intraperitoneally with lbp5mg/ (kg. day) × 7 days on the same day, and the spleen was taken on the seventh day to measure T cell proliferation response. .

1.2.4. antitumor effect of Lycium barbarum polysaccharide combined with cyclophosphamide: mice were subcutaneously injected with cy25mg/kg 2 days after subcutaneous inoculation of S180 cells, and the tumor inhibition rate was 31%. At the same time, intraperitoneal injection of LBP 10m/ (kg. day) × 7 days, the tumor inhibition rate increased to 47%, but there was no significant difference (P > 0.05). When cy was subcutaneously injected at 12.5mg/kg, the tumor inhibition rate was 14%, and its tumor weight was not significantly different from that of the tumor control group. When combined with lbp10mg/ (kg. day) × 7 days, the tumor inhibition rate increased to 54% by intraperitoneal injection, （P<0.01）， These results indicate that LBP can enhance the tumor suppressive effect of cy.

1.3. the effect of Lycium barbarum polysaccharide and combined application of anaerobic Corynebacterium brevis vaccine on the anti-tumor proliferation activity of mouse peritoneal macrophages: the intraperitoneal injection of Lycium barbarum polysaccharide into normal mice for 40mg/ (kg. day) × 7 days can enhance the anti-tumor target cell proliferation activity of ConA treated macrophages; Lbp5, 10, 20, 40mg/ (kg. day) × 7 days and combined with low-dose (250ug/ mouse) anaerobic Corynebacterium parvum vaccine (CP) had obvious synergistic effect. When the concentration of LBP was 20mg/kg, the effect was the most significant, and the inhibition rates of P815 and P388 proliferation were 85.5% and 63.6%, respectively; . The results showed that the combined application of LBP and CP could reduce the dosage of LBP and CP, enhance the effect, and reduce the toxic and side effects of CP. LBP mainly acts as an immune enhancer of T cells and can enhance the functions of CTL and NK cells; CP is a macrophage stimulator. The experiment shows that the synergistic effect can be obtained by combining the immune enhancers acting on different links of the immune response. The experiment also showed that LBP could activate macrophages in both non-specific anti-tumor and specific anti-tumor processes.

1.4. enhancing effect of Lycium barbarum polysaccharide on interleukin-2 activity in mice: LBP was added to the culture medium (splenocytes 5 × l0/ml and cona4ug/ml) that induced interleukin-2 (IL-2) in mouse splenocytes. The prepared supernatant containing IL-2 increased the proliferation activity of adult mouse thymocytes in vitro ([3H] TDR incorporation method). The level of IL-2 promoting lymphocyte proliferation in aged mice was significantly lower than that in adult mice. LBP can significantly improve this effect of IL-2 in aged mice, reaching the level of adult mice. The experiment showed that LBP enhanced the activity of IL-2 and restored the activity of IL-2 in aged mice.

1.5. the regulatory effect of Lycium barbarum polysaccharide on the low immune function of aged mice: the number of spleen hemolytic plaque forming cells (PFC) of aged mice was significantly lower than that of normal adult mice, decreased by 51%. After intraperitoneal injection of lbp20mg/ (kg. day) for 7 days, the spleen PFC value of aged mice can be significantly increased to the normal adult level. The incorporation of 3H TDR into thymocytes induced by ConA was also significantly decreased in older mice. The incorporation value of 3H TDR into thymocytes was significantly decreased when LBP was given at 5mg/kg. . .

1.6. immunopharmacological effects of Lycium barbarum polysaccharide on T, killer T and NK cells in mice and anti immunosuppressive effects of cyclophosphamide: lbp5-10mg/kg, injected intraperitoneally, can improve the proliferation function of splenic T lymphocytes in mice, enhance the killing function of CTL (CY toxic T lymphocyte), and the specific killing rate increases from 33% to 67%. LBP5mg/kg， Intraperitoneal injection can enhance the killing function of NK cells, and the killing rate increases from 12.4% to 18%. Lbp5-10mg/kg could antagonize the immunosuppressive effect of cyclophosphamide (CY) on T, CTL and NK cells in mice; The relative proliferation index (RPI) of T lymphocytes increased from 33% to 105%, the inhibition rate of CY on CTL decreased from 51% alone to 19% and 36% combined with LBP, and the killing rate of NK cells also increased from 9.5% of CY alone to 15% and 16%. These results indicated that LBP enhanced T cell-mediated immune response and NK cell activity in normal mice and CY treated mice.
. Rats were gavaged with 2ml of Lycium barbarum bag tea per day for 2 weeks, which could significantly increase the content of immunoglobulin (especially IgM) and complement activity.

2. hypoglycemic effect: Lycium barbarum extract 6g/kg, intraperitoneal injection, can reduce the blood glucose of rabbits by about 13% within 2-3 hours, and then gradually recover. Some people believe that its hypoglycemic effect is due to its derivatives containing guanidine.

3. anti fatty liver effect: long term (75 days) oral administration of betaine in fruits in rats can increase the levels of phospholipids in blood and liver; Taking betaine in advance or at the same time could antagonize the decrease of phospholipid and total cholesterol content in rat liver caused by carbon tetrachloride, and improve it; It can improve BSP, sgp-t, alkaline phospholipase, cholinesterase, etc. .

4. effect on blood pressure: 20mg/kg of water-soluble extract of fruits intravenously can reduce blood pressure and excite respiration in anesthetized rabbits; ; It has inhibitory effect on isolated rabbit heart, and makes rabbit ear vasoconstriction. Organic solvent extracts such as methanol and acetone also have slight hypotensive effect.

5. Lycium barbarum fruit extract can promote the growth and gas production of Lactobacillus casei and Lactobacillus acidophilus.

6. effect of Lycium barbarum Decoction on hydroxyproline content and hypoxia tolerance and anti fatigue effect in mice: hydroxyproline is an amino acid in collagen fibers and proteins, which is hydroxylated from proline. In the process of aging, the lack of oxygen supply affects the hydroxylation process of proline, resulting in the low content of hydroxyproline in collagen, the shrinkage and deformation of main organs, and the weight reduction of older people. As the lung function is affected, the lung capacity decreases, the reserve capacity decreases, the resistance decreases, the muscle strength decreases, and the ability to withstand hypoxia and fatigue decreases. Therefore, hydroxyproline content is related to aging. Lycium barbarum Decoction (13.23% concentration) was given to mice by gavage of 0.3ml/20g, once a day. After 30 days, the hydroxyproline concentration increased by 15.49% compared with the control. Compared with the survival time of 27.9 ± 1.3 minutes in the control group, the survival time of Lycium barbarum decoction was 37.5 ± 2.5 minutes (p< 0.01). There was also a significant difference between the anti fatigue test and the control (p< 0.01).

7. antitumor effect: in vitro tests showed that the fruit and leaves of Lycium barbarum had obvious inhibitory effects on human gastric adenocarcinoma kato- Ⅲ cells, and the fruit stalks and leaves of Lycium barbarum had obvious inhibitory effects on human cervical cancer HeLa cells. The mechanism of action was mainly manifested in inhibiting cell DNA synthesis, interfering with cell division, and reducing the ability of cell repopulation. Wolfberry fruit lyophilized powder suspension and cyclophosphamide were combined to treat Walker carcinosarcoma 256 in rats, and it was found that it had obvious protective effect on cyclophosphamide induced leukopenia.

8. other effects: adding betaine (4-6g/kg or 8g/kg) to the feed can increase the body weight of female and male chicks and increase egg production. Mice were gavaged with a 1:10 dilution of Shenqi ointment (0.2ml/ day) for 14 days, which could significantly increase the body weight, more than twice that of the control group, and the animals had glossy fur, plump muscles, and bright red blood. In addition, Lycium barbarum and betaine have protective effects on liver damage caused by CCl4.