Achyranthes bidentata
1、 Pharmacopoeia standard of Achyranthes bidentata       This product is the dried root of Achyranthes bidentata BL. When the stems and leaves wither in winter, dig them, remove the fibrous roots and sediment, bundle them into small bundles, dry them until they are wrinkled, cut the top flush, and dry them in the sun.
      [properties] the product is slender cylindrical, straight or slightly curved, 15-70cm long and 0.4-1cm in diameter. The surface is grayish yellow or light brown, with slightly twisted fine longitudinal wrinkles, sparse lateral root marks and transverse lenticel like protrusions. It is hard and brittle, easy to break, becomes soft after moisture, has a flat section, light brown, slightly horny like and oily, and the xylem of the central vascular bundle is large, yellowish white, with many yellowish white punctate vascular bundles scattered around it, which are intermittently arranged in 2-4 rounds. It is slightly sweet and slightly bitter.
      [identification] (1) cross section of this product: the cork layer is a series of flat cells extending tangentially. The inner layer of the bolt is narrow. The outer tough type of the heterotypic vascular bundle is intermittently arranged in 2-4 rounds. The vascular bundle of the outermost round is smaller, and some only have 1-several ducts. The cambium between bundles is connected into a ring, and the inward vascular bundle is larger; The xylem is mainly composed of vessels and small wood fibers, and the xylem in the root center integrates 2-3 groups. Parenchyma cells contain calcium oxalate sand crystals.
      (2) Take 4G of this product powder, add 50ml of 80% methanol, heat and reflux for 3 hours, filter, evaporate the filtrate, add 15ml of water to the residue, dissolve it with slight heat, add it to D101 macroporous adsorption resin column (inner diameter is 1.5cm, column height is 15cm), elute with 100ml of water, discard the water, elute with 100ml of 20% ethanol, discard the eluent, elute with 100ml of 80% ethanol, collect the eluent, evaporate to dryness, add 1ml of 80% methanol to the residue to dissolve it as the test solution. Another 4G of Achyranthes bidentata reference medicinal material was prepared into the reference medicinal material solution by the same method. . . In the chromatogram of the test sample, spots with the same color appear at the corresponding positions of the chromatogram of the control medicinal material and the chromatogram of the control sample.
      [check] moisture   .
      Total ash   Not more than 9.0% (general rule 2302).
      Sulfur dichloride residue   According to the determination method of sulfur dioxide residue (general rule 2331), it shall not exceed 400mg/kg.
      [extract] according to the hot leaching method under the determination of alcohol soluble extract (general rule 2201), saturated n-butanol with water as solvent shall not be less than 6.5%.
      [content determination] determine according to HPLC (general rule 0512).
      ; Octadecylsilane bonded silica gel was used as filler; The mobile phase was acetonitrile water formic acid (16:84:0.1); The detection wavelength was 250nm. The number of theoretical plates should not be less than 4000 according to the peak of β - ecdysterone.
      Preparation of reference solution   Take an appropriate amount of β - ecdysterone reference substance, accurately weigh it, add methanol to make a solution containing 0.1 mg per 1ml.
      Preparation of test solution   Take about 1g of this product powder (passing through No. 3 screen), accurately weigh it, place it in a corked conical flask, add water to saturate 30ml of n-butanol, tightly plug it, soak it overnight, sonicate (power 300W, frequency 40KHz) for 30 minutes, filter it, wash the container and residue with 10ml of methanol for several times, combine the filtrate and lotion, evaporate it to dryness, add methanol to dissolve the residue, transfer it to a 5ml volumetric flask, add methanol to the scale, and shake it evenly.
      Assay   Precisely suck 10 μ l of the reference solution and 10 μ l of the test solution respectively, inject them into the liquid chromatograph, and determine.
      Calculated as a dry product, the content of β - ecdysterone (c27h44o7) shall not be less than 0.030%.
      Decoction pieces
      [processing] Achyranthes bidentata   Remove impurities, wash, moisten thoroughly, remove residual reed heads, cut into sections, and dry.
      This product is a cylindrical segment. The outer skin is grayish yellow or light brown, with fine longitudinal wrinkles and transverse lenticels. It is hard and brittle, easy to break, and becomes soft with moisture. The cut surface is flat, light brown or brown, slightly horny and oily, the wood of the central vascular bundle is large, yellowish white, and there are many yellowish white punctate vascular bundles scattered outside, which are intermittently arranged in 2-4 rounds. It is slightly sweet and slightly bitter
      [extract] the same as that of medicinal materials, not less than 5.0%.
      [identification], [inspection], [content determination] are the same as those of medicinal materials.
      Wine Achyranthes bidentata   Remove the Achyranthes bidentata and stir fry it according to the wine roasting method (general rule 0213).
      This product is shaped like a segment of Achyranthes bidentata, with a slightly dark surface color and occasional focal spots. It has a slight wine aroma.
      [extract] the same as that of medicinal materials, not less than 4.0%.
      [identification], [inspection], [content determination] are the same as those of medicinal materials.
      [nature, taste and meridian tropism] bitter, sweet, sour, flat. Return to liver and kidney channels.
      [functions and indications] expel blood stasis and dredge meridians, tonify liver and kidney, strengthen muscles and bones, diuresis and gonorrhea, and lead blood downward. It is used for amenorrhea, dysmenorrhea, waist and knee soreness, muscle and bone weakness, gonorrhea, edema, headache, dizziness, toothache, aphthous ulcer, hematemesis, bleeding.
      [usage and dosage] 5 ～ 12g.
      [note] pregnant women should use it with caution.
      [storage] store in a cool and dry place and keep away from moisture.


2、 Chemical constituents of Achyranthes bidentata
. It also contains a variety of polysaccharides: one is a polysaccharide with anti-tumor activity precipitated from the aqueous extract of the root with acetone [3]; One is a water-soluble oligosaccharide ABS composed of 6 glucose residues and 3 mannose residues, which has significant activity of enhancing immune function [4]; The other is the peptide polysaccharide ABAB with immune activity, which is composed of glucuronic acid, galactose, galacturonic acid, arabinose and rhamnose in a 12:2:1:1 molar ratio. The peptide content is 24.7%, mainly composed ofglycine（glycine）、 GlutamicidL-aspartic acid (L-Aspartic acid) andl-serine . Also containsβ - ecdysterone（ecdysterone），25s Achyranthes bidentata sterone（25S-Inokosterone），25R Achyranthes bidentata sterone（25R-Inokosterone）[6]，Amaranth sterone(rubro sterone) [7] andL-arginine（L-arginine），glycine，l-serine ，L-aspartic acid , glutamate,L-threonine（threonine），L-Proline （proline），l-tyrosine （tyrosine），L-tryptophan（TCMLIByptophan），L-valine （valine），L-phenylalanine（L-phenylalanine），l-leucine .

3、 Pharmacological action of Achyranthes bidentata
1. ecdysterone contained in Achyranthes bidentata has strong protein synthesis promoting effect on protein assimilation. Two hours after intragastric or intraperitoneal injection of ecdysterone, the incorporation of amino acid precursors in the nuclei, mitochondria and microsomes of mouse liver increased, and the effect was stronger after 4 hours. At the same time, enhanced protein synthesis was also seen in the kidney, but returned to the original level after 4 hours. Sucrose density sample centrifugation analysis showed that the administered blood RNA had twice the casting activity compared with the control blood, suggesting that the protein synthesis effect of ecdysterone was at least accompanied by the role of promoting m-RNA synthesis. After 60 days of continuous administration of ecdysterone, the weight of weaned male mice increased rapidly, the protein synthesis level of liver and kidney also increased, and the liver tissue examination showed the hyperfunction of hepatocytes.
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2.2. effect on formaldehyde induced rat plantar swelling: Twenty healthy rats were randomly divided into two groups. The left and right plantar were subcutaneously injected with 2.5% formaldehyde 0.1ml to cause inflammation, and the left and right feet were used as self-control. After intraperitoneal injection of Achyranthes bidentata 16.6ml/ (kg · day), there was significant difference between the experimental group and the control group (p< 0.05).
2.3. effect on acetic acid thermophilic reaction mice oral administration of Achyranthes bidentata decoction has a very significant analgesic effect.
3. effect on cardiovascular system: 3.1. effect on heart: the extract of Achyranthes bidentata alcohol has certain inhibitory effect on isolated frog heart and anesthetized cat, and the effect of water decoction on anesthetized dog myocardium is more obvious.
3.2. effect on animal blood pressure: Achyranthes bidentata decoction has a transient hypotensive effect on anesthetized cats and dogs. When blood pressure drops, it is accompanied by respiratory excitation. The hypotensive effect also has no rapid tolerance phenomenon. It is presumed that its mechanism of action may be related to the release of histamine, cardiac inhibition and expansion of peripheral blood vessels.
4. effect on intestinal tract: rabbits and both sexes were selected, and duodenum, jejunum and ileum were selected respectively, with each segment of 3-4cm, according to the routine experimental method of smooth muscle. In 16 rabbits, 56 segments of intestine were given alcohol extract, which could excite the above intestinal segments, improve the tension, strengthen the contractility, and recover soon after changing the fluid.
5. effect on uterus: 5.1. effect on isolated rat uterine smooth muscle: 28 virgin rats (without sexual cycle smear examination) were randomly selected, and the body was 230 ± sdi7.3g. The isolated uterine specimens were prepared according to the conventional method. The uterine muscle segment was 1.5cm long, placed in a constant temperature Luo trough (20ml) containing dejaoln solution I, adjusted the baseline tension of 1.5g, and continuously injected with 95%o2+5%co2 (32 ± 0.5 ℃, ph=7.4-7.6) to balance for 30-60 minutes, and then started the experiment. During the experiment, the myogenic transducer was connected, and the uterine contraction activity was recorded by a desktop automatic balance recorder. In order to calculate the area under the uterine contraction curve (referred to as uterine contraction area, UCA) as an index. . The total glucosides of Achyranthes bidentata can obviously excite the uterine smooth muscle of rats. After administration, the uterine contraction amplitude increased, the frequency accelerated, and the tension also increased. Mainly regular rhythmic contraction, rarely spastic contraction. UCA was significantly increased compared with that before administration (p< 0.001). . The peak of UCA in each group also gradually moved forward, and the latency of uterine contraction was correspondingly shortened. Ten specimens were randomly selected from each dose group of the above experiment. The light was administered once, and after 3 minutes of action, the solution was changed and rinsed three times. After the specimen was recovered, the drug was administered once again to observe the effect of repeated administration on uterine smooth muscle. The results showed that the effect of Achyranthes bidentata saponins on the excitation of uterine smooth muscle was significantly weakened with the increase of the dose. When the dose increased to more than 0.5mg/ml, the re administration did not cause significant excitation of uterine smooth muscle.
. Before the experiment, the immature rats were taken vaginal smears, and the cycle was determined. According to the turnercd method, the sexual cycle was divided, and the estrous and diestrus rats were selected. Pregnant rats were divided into three pregnancy states according to the gestational period: early pregnancy within 7 days, mid pregnancy from 8 to 14 days, and late pregnancy above 15 days. Another 4 rats were subcutaneously injected with?? Diethylstilbestrol 0.lmg/kg, a total of 2 days, to create an artificial estrus period for the experiment. The experimental method was the same as before. 8-10 specimens were used in each group to compare the effects of 0.5mg/ml total glucosides of Achyranthes bidentata on uterine smooth muscle under different physiological conditions. The results are shown in Table 5. The UCA pregnant group was significantly higher than the non pregnant group within 10 minutes before administration (p< 0.05). . The youngest group was the lowest, and the late pregnancy group was the highest.
. The total saponins of Achyranthes bidentata were given 4.5mg/ml, and the effects of the drugs on the cervix and uterine horn were compared. The results are shown in Table 6. Achyranthes bidentata saponins 0.5mg/ml had no obvious excitatory effect on rat cervix. There was no significant change in contraction amplitude and frequency except that the tension of rat cervix increased to a certain extent after administration; It has an obvious effect on the uterine horn of rats, increasing the tension, contraction amplitude and frequency.
5.4. effect of rabbit uterus in vivo: three rabbits weighing 2-2.51g who were not pregnant and three rabbits who were pregnant in the second trimester (10-20 days of gestation) were selected for surgery under intravenous anesthesia with sodium pentobarbital 25mg/kg, and uterine contraction was recorded with the method of uterine suspension in situ. Tie the lower end of the uterine cannula firmly with the incision of the abdominal wall, and immerse the uterus in the low calcium ledeburite solution (1/4 normal calcium) heated to 38 ℃. After a static load of 3-5g and stable for 30-60 minutes, the experiment was started when the uterine activity described was constant. Local administration was adopted, that is, the liquid in the cannula was sucked out first, and then 0.5mg/ml of total saponin of Achyranthes bidentata (prepared with low calcium Ledeb solution) was added, and the observation and record were made for 30 minutes. After the first administration of drugs to 4 animals (half of non pregnant and half of the second trimester), the fluid was changed and rinsed for several times, and the drug was administered again once.
. It shows that the tension increases, the contraction amplitude increases, the frequency accelerates, and the excitatory effect generally lasts for about 25 minutes and gradually weakens. Repeated administration still had poor excitatory effect, but the effect was weakened compared with the first administration. Zhu He et al analyzed the mechanism of the excitatory effect of total saponins of Achyranthes bidentata (ABS) on rat uterus in vitro by using rat uterine contraction area (UCA) as an index and indomethacin and chlorpromazine as blockers. The experimental results showed that the excitatory effect of ABS on rat uterus ex vivo could be significantly attenuated by the administration of indomethacin (75mg/ rat gavage or 20ug/ml indomethacin in bath) before the experiment. Prostaglandin e2200mg/ml could significantly enhance the uterine function of ABS excited rats. Chlorpromazine 0.5mg/ml could also significantly attenuate the excitation effect of ABS on the uterus of pregnant and nonpregnant rats, while atropine 10ug/ml had no obvious effect on the uterine excitation induced by ABS. . It is suggested that ABS may stimulate the release of prostaglandins and 5-HT in rat uterus.
6. anti fertility effect: 6.1. anti fertility and abortion effect of total saponins of Achyranthes bidentata (ABS) on rats: 36 female virgin rats weighing 200-250g and 18 mature male rats weighing more than 250g were selected and housed in cages for more than 2 weeks. According to the literature method, the female rats were randomly divided into three groups (12 in each group) for experiments. After stopping the drug, the female rats were sacrificed, the uterus was dissected, and the pregnancy percentage, average number of live births, no births and implantation were calculated in each group. There was no significant difference in each index between the administration group and the control group. In abortion test, healthy female rats (230-260g) and male rats (> 750g) were selected and mated in 2:1 cages. . Dl4-dl9 were given ABS aqueous solution, all animals were sacrificed on the 20th day of pregnancy, and the animals were examined by laparotomy. Results compared with the control group, the rate of fetal loss was not significantly affected in the administration group, but the average body weight of the fetal was significantly lower than that of the control group (p< 0.01), and there was no production.
6.2. Anti fertility and anti implantation effects of ABS on mice: healthy adult mice were selected. The female rats weighed 26-35g and the male rats weighed 30-45g. They were mated in cages at a ratio of 2:1. The pregnant mice were randomly divided into the administration group and the control group after vaginal plug examination. Dl-d10 in the treatment group and Yin suppository was given by gavage once a day, while distilled water was given to the control group. When DL3 appeared in the vaginal suppository, the animals were sacrificed and the uterus was dissected. The inspection index was the same as that of the anti fertility experiment in rats. The results showed that s250mg/kg gavage for 10 days showed a certain anti fertility effect. The average number of stillbirths, live births and implantation points in this group were significantly different from those in the control group (p< 0.05. When the dose was increased by 500mg/kg or more, the pregnant rat rate was zero, showing 100% anti fertility effect. According to the literature method, the half effective amount of anti fertility effect of ABS was 218 ± 48mg/kg (p=0.95). The anti implantation test was performed at DL-5 when the vaginal plug appeared, and the drug was administered by gavage once a day. The animals were sacrificed 6 days after the drug was stopped, and the fetuses were dissected. Results the number of pregnant mice in the administration group was significantly less than that in the control group after s500mg/kg gavage (p< 0.05).
7. other effects: ecdysterone can improve liver function and reduce plasma cholesterol. Achyranthes bidentata decoction or alcohol extract can slightly diuresis and shorten the age of silkworm.
In vivo process of ecdysterone: the mouse experiment with 3H ecdysterone showed that after intraperitoneal injection for 10 minutes, the radioactivity intensity reached the highest level in various organs, and then rapidly decreased. Oral administration is slower, and the amount of liver, blood, kidney and other organs is lower than that of intraperitoneal injection. No matter by gavage or intraperitoneal injection, the liver was the highest in all organs. When injected intraperitoneally, the radioactive activity can appear rapidly in feces or urine, but it appears slowly when taken orally. It can be seen that there is more ecdysterone in feces and urine when administered by both routes, indicating that the absorbed ecdysterone is mainly excreted into the intestine through hepatobiliary secretion.