Studies On Preparation And Quality Control Of Multi-Components In Astragalus Membranaceus (Fisch.) Bge. And Salvia Miltiorrhiza Bge.

As commonly used herbs, Astragalus membranaceus (Fisch.) Bge. and Salvia miltiorrhiza Bge., have a long history in Traditional Chinese Medicine (TCM). Modern medical studies indicate that Radix astragali has a function of rejecting viral infection in cell culture, animals and anthropometric, and can markedly stimulate human body to release ITF. Several constituents, including triterpenoids saponins, flavonoids, polysaccharides, amino acids and microelements, etc. were isolated from it. The saponin is the active constituent, which has apparent activation in anti-inflammatory, relieving pain, anti-aging, anti-tumoring and immune enhancement. Salvia miltiorrhiza Bge. have good effects on promoting blood flow, regulating menstruation, and relieving pain, clearing hot and calming the nerves. It has been widely used in curing the disease of cardiovascular system clinically. It has more than thirty active components including the liposoluble constituents, such as the salvioland and hydrosoluble constituents such as the tanshinol and the salvianolic acid. Its hydrosoluble constituents including tanshinol, protocatechualdehyde, salvianolicacid B, etc. have functions of anti-thrombusing, improving blood circulation, anti-oxidation etc. Its liposoluble constituents have active effects on anti-inflammatory, restraining the oxidation of low density lipoprotein, etc. The different chemical constituents have different pharmacological actions, therefore it accords with the mechanism of pharmacological action of chemical composition in Chinese crude drugs and the synergetical principle of traditional Chinese medical rational. We have to achieve the aim of controlling the quality of Chinese herbs. This is the only way to establish and consummate the quality criteria of Chinese crude drugs.But the reference standards of multitude index constituents are hard to obtain and there are not well-rounded assaying of simultaneously determining the multitude active constituents in a medicinal substance. Because of that, we extracted and separated calycosin-7-O-β-D-glucopyranoside, astragaloside Ⅰ , astragaloside Ⅱ, astragaloside IV and acetylastragaloside from Astragalus membranaceus (Fisch.) Bge. and malol,salvianolic acid, rosmarnci acid, tanshinone II A, clyptotanshinon and tanshinone I from Salvia miltiorrhiza Bge. and identified their structures, and studied the craft of the reference standards of Astragalus membranaceus (Fisch.) Bge. and Salvia miltiorrhiza Bge. Basing on the control articles of high purity, we established the quality control methods of multitude constituents of Astragalus membranaceus (Fisch.) Bge. by HPLC/MSD and Salvia miltiorrhiza Bge with HPLC/DAD. In this part of the separation identification of several active constituents and the craft of some reference standards, we primarily analyzed several active constituents in Astragalus membranaceus (Fisch.) Bge. and Salvia miltiorrhiza Bge., separated eleven chemical compounds by modern separation techniques and identified their structures with the chemical identification and modern spectroscopic assaying. They are calycosin-7-O-p-D-glucopyranoside, astragaloside I, astragaloside II, astragaloside IV, and acetylastragaloside from Astragalus membranaceus (Fisch.) Bge. and malol, salvianolic acid, rosmarinci acid, tanshinone II A, clyptotanshinon and tanshinone I from Salvia miltiorrhiza Bge. Based on accurate identification to the substances separated from the two herbs, we studied the craft of the two reference standards in the two herbal medicines. The yield of astragaloside IV, salvianolic acid B, rosmarinci acids are 0.15%, 0.17%, 0.08%, respectively, and their purity is over 99.0%. The results indicated that the craft accord with the regulations and the referace standards can be prepared easily. The groundwork is established for the multitude index constituents assaying of the crude drugs.The second part was studies on the quality control of multiple index constituents in Astragalus membranaceus (Fisch.) Bge. and Salvia miltiorrhiza Bge. This part includes the quality control methods of multiple constituents of Astragalus membranaceus (Fisch.) Bge. by HPLC/MSD and Salvia miltiorrhiza Bge. by HPLC/DAD. Firstly, the HPLC/MSD method was established to determine the contents of astragaloside I , astragaloside II, astragaloside IV and acetylastragaloside in Astragalus membranaceus (Fisch.) Bge. The HPLC was carried out on a C18 column (Agilent Zorbax SB C18, 4.6mmx250mm, 5^im) with a sample injection volume of lOjxl. The mobile phase was acetonitrile and water with gradient elutionat a flow rate of l.Oml/min. For operation in MS mode, a mass spectrometer was used for all analyses. Ionization was achieved using electrospray in the negative ionization mode. The following parameters were optimized for analysis: the splitting voltage of 100V for negative ionization mode, desolvation gas (N2) was heated to 350°C;nerbuliserand desolvation gas flow was 7.0 1-min"1. The capillary tubing voltage was 3500V. The linear ranges of astragaloside I, astragaloside II, astragaloside IV, and acetylastragaloside were 0.08960-14.34 ngml"1 (r=0.9999), 0.08240-3.296 ng-ml1 (r=0.9997), 0.1052~4.208jig-ml"1 (r=0.9998), 0.02050-3.280 Hg-ml"1 (r=0.9998), respectively. The accuracy, stability, precision, reproducibility, limit of detection and recovery are all within the requirement. Secondly, The HPLC method was established to determine the contents of salvianolic acid B, rosmarinci acid, tanshinone II A, clyptotanshinon in Salvia miltiorrhiza Bge. The separation of HPLC was achieved using Agilent Zorbax SB C18 (4.6mmx250mm, 5 urn) with a sample injection volume of 20^,1 and the wavelength was 280nm. The mobile phase was comprised of water-methanol (2% acetic acid) with gradient elution. The results indicated that the linear ranges of salvianolic acid B, rosmarinci acid, tanshinone IIA and clyptotanshinon were 3.750-120.2 fig-ml'1 (r=0.9999), 34.20-1095 ng-ml"1 (r=0.9999), 0.6300-20.30 ngml"1 (r=0.9999), 1.020-32.68 (xg-ml1 (r=0.9996) respectively. The stability, precision, reproducibility and the recovery were within the requirement. The scientific foundation was provided to evaluate the quality of medicinal materials more accurately and generally.