Study On Quality Control Of Concentrated Herb Granule Of Bletilla Striata

1 ObjectiveRhizoma Bletillae is the dried tuber of Bletilla striata (Thunb.) Reichb. fand an astringent herbs. Concentrated Herb Granule (CHG), under the guidance of TCM theory, is made from a single recipe or compound recipe which goes through the modern manufacturing process of extracting, concentrating, drying, granulating and quantitative sub-packaging. Compared with the Chinese traditional medicinal herbs, CHG is convenient for carrying and administering. However, some studies reported that the chemical composition in CHG is instable and at present there is no unified national standard for CHG. Bletilla striata CHG also is lack of standard. This study focus on the technology of Bletilla striata CHG mainly include material quality, extracting process and quality standard for the quality control of Bletilla striata CHG.2 Methods2.1 Quality study of the crude drugs of Bletilla striataThe qualities of crude drugs of Bletilla striata are checked by Chinese Pharmacopoeia 2010 with moisture, ash and TLC. The methods of water soluble extract content and determination of content of cinnamic acid are established.Bletilla striata polysaccharide (BSP) is main component of crude drugs. Different BSP extraction methods were evaluated by the polysaccharide contentThrough single factor experiment and orthogonal experiment, extraction method was optimized. The crude polysaccharide purification process and alcohol precipitation process were investigated with the protein residual contents and the BSP recovery. TLC identification and content determination method of BSP were established. The DPPH free radicals and hydroxyl radicals was used to detect the antioxidant activity of polysaccharide. The antibacterial activity of Bletilla striata was evaluated by the staphylococcus aureus and bacillus subtilis.HPLC fingerprint was used to analyze samples from crude drugs of Bletilla striata. The extraction methods and the chromatograms systemic methodology were optimized for the HPLC fingerprint establishment.2.2 Bletilla striata CHG extraction processBase on the standard decoction, the parameters of Bletilla striata CHG extraction and concentration process were optimized. Bletilla striata water extraction technology of quality was evaluated by dry extract rate, TLC, HPLC fingerprint, and cinnamic acid content.2.3 Quality study of Bletilla striata CHGQuality of Bletilla striata CHG was evaluated by TLC, fingerprint, cinnamic acid content.The Bletilla striata CHG from different manufacturers were studied and compared.3 Results3.1 Quality study of the crude drugs of Bletilla striataThe 13 batches of Bletilla striata are met the Chinese Pharmacopoeia 2010 requirements in moisture, ash and TLC identification. Water-soluble extract (cold) content shall not be less than 30.0% in this study. The optimal extraction conditions of microwave assisted extraction BSP was as follows::The material liquid ratio was 1:75 (W/V, G:mL), and the extraction temperature and time was 50℃ and 15 min, extraction times was 2 times with the microwave power 700w.The polysaccharide recovery of microwave assisted extraction was 25.80%, which was better than ultrasonic extraction (13.52%) and reflux extraction (14.14%). The sequence order of factors on the effect of polysaccharide was extraction times >solid-liquid ratio> extraction time >extraction temperature. BSP TLC method studied showed that glucose and mannose had a good separation in 15℃ temperature with 32% humidity conditions. The HPLC determination method of cinnamon acid and polysaccharide were estabished. The Bletilla striata contains not less than 0.020 per cent of Cinnamic acid and 0.50 per cent of glucose and 0.70 per cent of mannose, caculated with reference to the dried drug. HPLC fingerprint has 13 common peaks and the similarity was proved to be higher than 0.902. BSP can scavenge on DPPH and hydroxyl radicals. Bletilla striata ultrasonic extraction has obvious antibacterial activity.3.2 Bletilla striata CHG extraction processBletilla striata CHG extraction and concentration process:10,8,8 times water was added to the crude drugs of Bletilla striata with decocting three times(each time with 1.5 hours), the liquid was filtrated by 300 mesh filter cloth. Under the condition of the vacuum degree-0.06～-0.09, the filtrate was concentrated by decompression concentration in 80℃. The quality study of water extract of Bletilla striata showed that the thirteen batch of the extract rate range was 10.19%～40.25%, and the cinnamic acid contain range was 0.0011%～0.0182%, and the HPLC fingerprint had 10 common peaks, the similarity of different batch was proved to be higher than 0.866.3.3 Quality study of Bletilla striata CHGThe determination method about Bletilla striata CHG were established with TLC, fingerprint, cinnamic acid content But the thin-layer chromatograms of Bletilla striata dispensing granules in 10 batches didn’t show the same spot color in the corresponding place. The cinnamic acid content of different batches of Bletilla striata CHG were different greatly. HPLC fingerprint showed the similarity between different batches of Bletilla striata CHG is low. F1 batches has the greatest similarity, while Al, A2, A3, A4 batch similarity is low, less than 0.1.4 Conclusions4.1 Quality study of the crude drugs of Bletilla striataThe 13 batches of Bletilla striata are met the Chinese Pharmacopoeia 2010 requirements. Microwave assisted method is better than conventional reflux extraction with highly efficient energy-saving advantages. The HPLC determination method of cinnamon acid and polysaccharide were estabished. BSP TLC identification was carried out and the method is simple, rapid. The establishment of Bletilla striata fingerprint determination method was suitable for quality control. Bletilla striata ultrasonic extraction liquid has obvious antibacterial activity and BSP has certain antioxidant activity.4.2 Bletilla striata CHG extraction processThe optimization process is close to the standard process, which is good for Bletilla striata CHG production. The water extract quality standard has some theoretical and practical value for the quality control of CHG.4.3 Quality study of Bletilla striata CHGThe determination method of Bletilla striata CHG was established by TLC, fingerprint, cinnamic acid content. It is simple and reproducibility for quality evaluation of Bletilla striata CHG foundation.