White China Root Quality Evaluation Method Research

Rhizoma Heterosmilacis Japonicae is the dried of Heterosmilax japonica Kunth.,Heterosmilax yunnanensis. or Heterosmilax chinensis Wang.. Rhizoma Heterosmilacis Japonica is used in clinic as an important Traditional Chinese Medicine (TCM).14 samples of Rhizoma Heterosmilacis Japonica were purchased from TCM shops or straightly collected from the plant habitats in different places of China. High performance liquid chromatographic fingerprint chromatogram was established and quality assessment and identification was researched for Rhizoma Heterosmilacis Japonica. The chemical compotions of fatty acid from Rhizoma Heterosmilacis Japonica have been studied by GC-MS and known substance reference methods. And the method of quantitative determination of 2 kinds of fatty acids from Rhizoma Heterosmilacis Japonica was established. The monosaccharides in the Rhizoma Heterosmilacis Japonica polysaccharide was determined by precolumn derivation HPLC method. The methods above provided reliable basis to control the quantity of Rhizoma Heterosmilacis Japonica and as well as to use it reasonably.The method of HPLC fingerprint chromatogram was established for Rhizoma Heterosmilacis Japonica. The separation was performed on a KromasilTM C18 analytical column (250 mm×4.6 mm i.d.,5μm) with the mobile phase consisting of acetonitrile and water with gradient elution mode at the flow rate of 1.0 mL·min-1. Elution procedure:0-80 min,95% B-65% B; 80-90 min, 65% B. The detection was set at 215nm.14 batches of Rhizoma Heterosmilax japonica was determined by HPLC fingerprint chromatogram. The Hierarchical cluster was applied to the fingerprints of 14 batches of Rhizoma Heterosmilax japonica. And the 14 batches of Rhizoma Heterosmilax japonica was divided into four grades. Among them the grade I was commendatory and theothers were general. Mutual pattern was established from 11 batches in the grade I by using the fingerprint chromatogram software required to use by the Chinese Pharmacopoeia Committee. Similarity calculations were studied by comparing the Rhizoma Heterosmilax japonica and mutual pattern. The result of thecommendatory should be all over 0.90 and the other samples were the general.The chemical composition of fatty acid from Rhizoma Heterosmilacis Japonica have been studied by GC-MS method.3 constituents of fatty acid were got and 2 of them were identifed. They were palmitic acid(45.01%) and linoleic acid(53.79%). The total content of the two compounds was 98.98%. The method of quantitative determination of 2 kinds of fatty acids from Rhizoma Heterosmilacis Japonica was to develop. The samples was determined with capillary column HP-5(30 m×0.25 mm,0.25μm) by GC. The detector was FID; temperature programs was 200℃for 9 min and then programmed to 260℃at 10℃·min-1 for 10 min; Nitrogen was used as carrier gas(1.0 mL·min-1). GC method was applied for the determination of Palmitic acid and linoleic acid. The linear range for palmitic acid methylester and linoleic acid methylester were 0.042-0.844 mg·mL-1(r=0.9991) and 0.070-1.399 mg·mL-1 (r=0.9991). The average recoveries (n=9) were 102.5%(RSD=2.6%) and 96.3%(RSD=3.0%). The method is sensitive, simple and accurate for the determination of palmitic acid and linoleic acid in Rhizoma Heterosmilacis Japonica.Precolumn derivation HPLC method was to establish to determine the monosaccharides in the Rhizoma Heterosmilacis Japonica polysaccharide and theirmolar ratio. Rhizoma Heterosmilacis Japonica sample was hydrolyzed with trifluoroacetic(TFA) and then was derivated by 1-phenyl-3methyl-5-pyrazolone (PMP). The monosaccharide composition of Rhizoma Heterosmilacis Japonica polysaccharide was carried out by reversed-phase technique on a Kromasil C18 column with a mobile phase composed of 100 mmol·L-1 ammonium acetate buffer (pH5.5) and acetonitrile in the ratio of 77:23. The detection was carried out at 245nm. The Rhizoma Heterosmilacis Japonica polysaccharideis composed of mannose, rhamnose, glucose, galactose and xylose with amolar ratio of 66:1:7590:452:528. The HPLC method with precolumn derivatization is appropriate for the analysis of monosaccharide composition of Rhizoma Heterosmilacis Japonica polysaccharide and the method is simple, quick and accurate.