| ObjectiveTo study Uyghur Medicine of diaphragma juglandis fructus in Xinjiang systematically. To clarify its basic treatment mechanism of deficiency of kidney-YANG and to screen the effective solvent parts; to study the extraction, isolation, purification, physical and chemical properties, structure determination, etc. of the chemical compositions from the effective solvent parts, which will provide scientific basis for new anti-deficiency of kidney-YANG drug research and comprehensive utilization of Uyghur Medicine of diaphragma juglandis fructus.Methods1. The preliminary study of the main chemical constituents of diaphragma juglandis fructus: by way of in vitro pre-test method, we study the main chemical constituents of diaphragma juglandis fructus preliminarily. For the first time, with Ginsenoside Rb1 and Glucose as reference substances, the contents of total Saponins and polysaccharides from diaphragma juglandis fructus were assayed respectively. Referring to pertinent literature, the contents distribution of total alkaloids, total flavonoids, total saponins, polysaccharides, total tannic acid and other chemical components of diaphragma juglandis fructus was studied.2. Effect of Uygur Medicine diaphragma juglandis fructus on mice model of kidney-YANG deficiency: After mice model of kidney-yang deficiency built with the high dose Hydrocortisone, the groups were treated with different solvent extracts (n-butanol group, 95% ethanol group and distilled water group) respectively. Investigate weight growth rates and organ coefficients, and evaluate the histopathological changes of kidney. The effects of different solvent extracts from Diaphragma Juglandis Fructus on Mice Model of Kidney-yang Deficiency were discussed, and the treatment mechanism of deficiency of kidney-YANG was analyzed.3. Extraction, isolation and purification of the effective solvent parts: the three effective solvent parts (n-butanol part, 95% ethanol part and distilled water part) were graded by polyamide column chromatograph, macroporous resin AB-8 column chromatograph respectively. All grades were separated and purified by column chromatograph of Sephadex LH-20 and polyamide. The purities of the chemical compounds from the effective solvent parts were examined by UV, column chromatograph of Sephadex LH-20, freeze thawing assay and TLC. The structure determination of those chemical compounds was identified by UV, IR and NMR, etc.Results1. Preliminary studies have shown that diaphragma juglandis fructus mainly consists of volatile oil, grease, alkaloids, flavonoids, saponins, tannins, organic acids or phenols, carbohydrate, amino acids, peptides or proteins and so on; and lactones or cumoric anhydride, cardiac glycosides, steroids or triterpenoids were not detected. Through comparative studies, the content of flavonoid is the highest, polysaccharide second, the results of which are basically consistent with the results of test tube method.2.①The systemic depletion induced by high dose cortical hormone was shown in Kidney-yang Deficiency group, but the three solvent extracts of Diaphragma Juglandis Fructus could improve the depletion at a different degree.②Compared with the control group, the body weights of the mice in Kidney-yang Deficiency group were lower, which had statistical significance (P<0.01). After the administration, compared with the control group, the body weights of the mice in Kidney-yang Deficiency group and n-butanol group (high, middle and low) were lower, which all had statistical significance (P<0.05). Compared with the Kidney-yang Deficiency group, the body weights of the mice in n-butanol group (high, middle and low) and distilled water group were higher, which all had statistical significance (P<0.01). Compared with the treated group, the body weights of the mice in the three groups were higher, which all had no statistical significance (P>0.05);③Compared with the control group, the adrenal coefficient of the mice in Kidney-yang Deficiency group were lower, which had statistical significance (P<0.01). Compared with the Kidney-yang Deficiency group, the adrenal coefficient of the mice in the treated group, n-butanol groups (high, middle and low) was higher, which all had statistical significance (P<0.01, P<0.05, P<0.05, P<0.05). Compared with the treated group, the adrenal coefficient of the mice in the three groups was higher, which all had no statistical significance (P>0.05);④The kidney glomerulus, kidney tubules and renal interstitium of the mice in the control group were normal, the structure of which were distinct and had no abnormal pathological characteristics. The glomerular capillaries of the mice in Kidney-yang Deficiency group expanded in midrange; and the renal tubular epithelial cells were in the condition of hydropic degeneration and mortification; renal interstitium had the symptom of bleeding. Besides hydropic degeneration of the renal tubular epithelial cell, the bleeding of the renal interstitium, there was no abnormal pathological characteristics. The n-butanol groups (high, middle and low) improved expansion of glomerular capillaries, hydropic degeneration of the renal tubular epithelial cell and the bleeding of the renal interstitium in different degree. The extracts of 95% ethanol and distilled water had no obvious effects on the kidney functions of kidney-yang deficiency mice model.3. Nine compounds, Fr.Ⅰ(24.0mg), Fr.Ⅱ, Fr.Ⅲ(20.3mg), Fr.Ⅳ(9.3mg), Fr.Ⅴ(20.2mg), FrⅥ(15.4mg), Fr.Ⅶ(8.8mg), Fr.Ⅷ(30.5mg) and Fr.Ⅸ(29.8mg), were obtained from diaphragma juglandis fructus. Based on chemical reactions and spectral analysis (SephadexLH-20, UV, IR, MS, 1H-NMR, 13C-NMR, etc.), two of them were elucidated, which is 5,7,8,3',4'- flavonol-3-O-α-D- rhamnoside.Conclusion1. For the first time, the pre-test of diaphragma juglandis fructus chemical constituents has been fulfilled, which is benefit for the research of extraction, isolation and purification of the active components, and which could be offer foundation for the research and exploitation of the resource of diaphragma juglandis fructus. The results of contents of the main components are basically consistent with the results of test tube method.2. Diaphragma juglandis fructus has definite effects on mice model of kidney-yang deficiency induced by high dose of hydrocortisone. The extract of n-butanol, chief components of which is glucosides such as flavonoid glycoside, has obvious pharmaco-activity, which are probably the effective components. Diaphragma juglandis fructus has reverse effects on pathological characteristics of kidney of kidney-yang deficiency mice model at some extent.3. Six homogeneous polysaccharides were obtained from diaphragma juglandis fructus and their structures were elucidated for the first time. All of the homogeneous polysaccharides were preliminarily elucidated; two identical flavonoid glycosides, which is 5,7,8,3',4'- flavonol-3-O-α-D-rhamnoside. |
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