Two new steroidal saponins from the rhizomes of Dioscorea panthaica and their cytotoxic activity.

Two new steroidal saponins, dioscoresides C (1) and D (2), along with a new natural product, pregnadienolone 3-O-beta-gracillimatriose (3), and two known compounds, pregnadienolone 3-O-beta-chacotrioside (4) and pseudoprotodioscin (5), were isolated from the rhizomes of Dioscorea panthaica Prain et Burkill. On the basis of extensive NMR studies and chemical evidence, dioscoresides C and D were determined to be 26-O-beta-D-glucopyranosyl-3 beta,26-dihydroxy-23(S)-methoxy-25(R)-furosta-5,20(22)-dien-3-O-alpha-L-rhamnopyranosyl-(1-->2)-[alpha-L-rhamnopyranosyl-(1-->4)]-beta-D-glucopyranoside and 26-O-beta-D-glucopyranosyl-3 beta,26-dihydroxy-20,22-seco-25(R)-furosta-5-en-20,22-dine-3-O-alpha-L-rhamnopyranosyl-(1-->2)-[alpha-L-rhamnopyranosyl-(1--> 4)]-beta-D-glucopyranoside. These compounds showed mild cytotoxicity against the cancer cell lines, A375, L929, and HeLa, in a dose-dependent manner.

[Research on the transformation of ginsenoside Rg1 by intestinal flora].

OBJECTIVE: The decomposition of Ginsenoside Rg1(Rg1) by intestinal bacteria in rats or human was investigated both in vitro and in vivo. METHOD: The decompositions were investigated by means of thin-layer chromatography and Electron spurt ion mass. RESULT: It was found that Rg1 was converted into two metabolites [Rh1 and 20(S)-protopanaxatriol(20S-Ppt)] by human intestinal bacteria in vitro, while three compounds [Rh1, F1, and 20(S)-Ppt] were detected in rat intestinal bacteria metabolism in vitro and in vivo. CONCLUSION: The decomposition and/or metabolism of Rg1 in rat and human digestive tract was confirmed. The mode of metabolism in rat is Rg1-->Rh1(F1)-->Ppt, while in human it is Rg1-->Rh1-->Ppt.

[Studies on metabolism of oxymatrine by human intestinal bacteria].

OBJECTIVE: To study the oxymatrine metabolism induced by human intestinal bacteria and its absorbed metabolites in blood. METHOD: TLC and HPLC were used to examine oxymatrine and its metabolites, and UV, IR, NMR and MS were used to confirm the chemical structures of the metabolites. RESULT: Oxymatrine was transformed into matrine by human intestinal bacteria metabolism in vitro. Rats were given orally oxymatrine 100 mg.kg-1 and were decapitated 3 hours after administration, and their blood was taken to examine serum metabolites by TLC and HPLC, which revealed that oxymatrine and matrine were absorbed into blood. CONCLUSION: Oxymatrine will be transformed into matrine when it is given orally and both of the alkaloids can be absorbed to blood.

[A comparison of chemical composition and bioactivity of polypeptides from velvet antlers of Cervus nippon Temminck and Cervus elaphus Linnaeus].

OBJECTIVE: To compare the chemical composition and bioactivity of polypeptides(PPs) isolated from velvet antlers of sika deer (Cervus nippon Temminck) and red deer (Cervus elaphus Linnaeus). METHOD: The two kind of polypeptides were isolated from the above mentioned velvet antlers with same technology. The chemical composition was determined using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and MALDI-TOF mass spectrometry. Stimulant activity of cells proliferation was measured by [3H] TdR incorporation into DNA. RESULT: The graphs of SDS-PAGE and MALDI-TOF MS of velvet antler polypeptides (VAPPs) from Chinese and New zealand red deer were very similar, but there were obvious difference in respect of graph between sika deer and red deer. VAPPs 25-50 mg.L-1 showed marked proliferation-promoting activity for rabbit costed chondrocytes, either sika deer or red deer. However, the activity of sika deer VAPPs 12.5 mg.L-1 for epidermal cells was weaker than that of red deer (12.5 mg.L-1). CONCLUSION: The chemical property and bioactivity of VAPPs from sika deer and red deer are significantly different.

[Isolation and identification of steroidal saponins from Dioscorea panthaica Prain et Burkill].

AIM: To investigate the chemical constituents of the rhizomes of Dioscorea panthaica Prain et Burkill and look for new active compounds. METHODS: The compounds were isolated with silica gel and HPLC, and their structures were elucidated on the basis of chemical evidences and spectral analysis (IR, ESI-MS, 1HNMR, 13CNMR, DEPT, 1H-1H COSY, HMQC, HMBC, NOESY). RESULTS: Three steroidal saponins were isolated from the rhizomes of Dioscorea panthaica. They were elucidated as 26-O-beta-D-glucopyranosyl-3 beta, 26-diol-23(S)-methoxy-25(R)-delta 5,20(22)-diene-furosta-3-O-[alpha-L-rhamnopyranosyl-(1-->2)- O-alpha-L-rhamnopyranosyl-(1-->4)]-beta-D-glucopyranoside (I), pseudoprotodioscin (II), 26-O-beta-D-glucopyranosyl-3 beta, 26-diol-25 (R)-delta 5,20(22)-diene-furosta-3-O-alpha-L-rhamnopyranosyl- (1-->2)-O-beta-D-glucopyranoside (III). CONCLUSION: Compound I is a new steroidal saponin and was named as dioscoreside C. Compound II and III were obtained from this genus for the first time.

[Metabolism of ginsenoside Rb1 and panaxadiol saponins by fungi].

AIM: To study the metabolic process of ginsenoside Rb1 (G-Rb1) and panaxadiol saponins (PDS) by fungi. METHODS: Ten strains of fungi were incubated with G-Rb1 and PDS at a certain temperature with shaking. A portion was taken out at different time and mixed up with butanol. The butanol extract was analysed by thin layer chromatography (TLC), high performance liquid chromatography (HPLC) and electrospray ionazition mass spectrometry (ESI-MS). RESULTS: It was found that there were ginsenoside-Rd (G-Rd), ginsenoside-F2 (G-F2), compound K (CK) and 20(S) protopanaxadiol (Ppd) metabolites beside the prodrug G-Rb1 induced by fungi (No. 1, 2, 3, 6, 8, 9). CONCLUSION: The six strains of fungi have different degrees of ability to metabolize G-Rb1 and PDS. The possible metabolic process could be as follows: G-Rb1 (or PDS)-->G-Rd-->G-F2-->CK-->Ppd.

Effects of polysaccharides (FI0-b) from mycelium of Ganoderma tsugae on proinflammatory cytokine production by THP-1 cells and human PBMC (I).

AIM: To compare the effects of water-soluble polysaccharides, FI0-b, and its formic acid-modified derivative, FI0-b-H, on production of human proinflammatory cytokines. METHODS: The polysaccharides were modified by formic acid. Cytokine production was quantitated by radioimmunoassay. mRNA for cytokines was measured by semi-quantitative RT-PCR. RESULTS: FI0-b and FI0-b-H 4, 40, and 400 mg/L significantly downregulated interleukin-1 alpha (IL-1 alpha) production by THP-1 cells induced by lypopolysaccharide (LPS) 1 or 10 mg/L and phorbol myristate acetate (PMA) 200 nmol/L. At lower stimulation with LPS 10 mg/L and PMA 200 nmol/L, both polysaccharides significantly upregulated tumor necrosis factor alpha (TNF alpha) production by THP-1 cells. However, at higher stimulation with LPS 100 mg/L and PMA 200 nmol/L, they downregulated TNF alpha production. FI0-b-H downregulated interleukin-8 (IL-8) production by THP-1 cells at a lower-dose of LPS 1 mg/L and PMA 200 nmol/L, but upregulated IL-8 production at a higher-dose of LPS 10 mg/L and PMA 200 nmol/L. Production of cytokines (IL-1 alpha and TNF alpha) was transcriptionally or post-transcriptionally regulated by FI0-b and FI0-b-H. CONCLUSION: The water-soluble polysaccharides of Ganoderma tsugae mycelium have bidirectional immunomodulatory effects on cytokine production in different stimulatory conditions in a dose-dependent manner. Compared with FI0-b, FI0-b-H has more marked effects on human proinflammatory cytokine production.

Effects of polysaccharides (FI0-c) from mycelium of Ganoderma tsugae on proinflammatory cytokine production by THP-1 cells and human PBMC (II).

AIM: To study the effects of water-soluble polysaccharides. FI0-c, and its sulfated derivative, FI0-c-S, on production of human proinflammatory cytokines, interleukin-1 alpha (IL-1 alpha) and tumor necrosis factor alpha (TNF alpha). METHODS: The herbal polysaccharides were modified by chlorosulfornic acid in dimethyl sulfoxide (Me2SO). Cytokine production was measured by radioimmunoassay, mRNA for the cytokines was measured by semi-quantitative RT-PCR. RESULTS: FI0-c 4 mg/L itself induced IL-1 alpha production by THP-1 cells without stimulants, such as lipopolysaccharides (LPS) and phorbol myristate acetate (PMA). On the other hand, FI0-c and FI0-c-S inhibited the IL-1 alpha production by THP-1 cells with these stimulants. FI0-c and FI0-c-S significantly upregulated TNF alpha production by THP-1 cells without stimulants or at a low dose of LPS 10 mg/L and PMA 200 nmol/L, whereas these polysaccharides markedly downregulated the TNF alpha production by a high dose of LPS 100 mg/L and PMA. Human peripheral blood mononuclear cells (PBMC) responded to FI0-c and FI0-c-S in IL-1 alpha and TNF alpha production in a fashion similar to THP-1 cell responses. FI0-c 4 mg/L downregulated high-dose LPS- and PMA-induced IL-1 alpha or TNF alpha mRNA and their protein production by THP-1 cells. CONCLUSION: The water-soluble polysaccharides of Ganoderma tsugae mycelium have bidirectional immunomodulatory effects on cytokine production in different cell stimulatory conditions. Chemical modification of this polysaccharide changed the intensity of regulatory effect on cytokine production.

Velvet antler polypeptides promoted proliferation of chondrocytes and osteoblast precursors and fracture healing.

AIM: To study the effects of velvet antler (VA) total polypeptides (VATP) and VA polypeptides, VAP-A, VAP-B, and VAP-C on proliferation of chondrocytes and osteoblast precusors. METHODS: Chondrocytes (rabbit and human fetus) and osteoblast precusors (chick embryo) were incubated in the culture medium containing VATP or VAP-A, VAP-B, and VAP-C. [3H]TdR incorporation into DNA was measured. Fracture healing-promoting action of VATP was determined in rats. RESULTS: VATP 50-200 mg.L-1 and VAP-B 12.5, 25, and 50 mg.L-1 showed most marked proliferation-promoting activity for rabbit costed chondrocytes and increased incorporation of [3H]TdR from (73 +/- 9) Bq (control group) to (272 +/- 55), (327 +/- 38), and (415 +/- 32) Bq, respectively (P < 0.01). The activity of VAP-A was weaker than that of VAP-B, and VAP-C had no activity. VATP 10 and 20 mg.kg-1 by local injection into the cross-section fracture area accelerated healing of radial fracture. The healing rate of VATP-treated group was higher (75%) than that of control group (25%) (P < 0.05). CONCLUSION: VATP accelerated fracture healing by stimulating proliferation of chondrocytes and osteoblast precursors.

[Anti-inflammatory effects of pilose antler peptide].

Pilose antler peptide (PAP:MW:7200; amino acid residue:68) isolated from the antlers of Cervus nippon temminck 10 and 20 mg.kg-1 i.p. produced inhibitions towards acute and chronic inflammations in a dose-dependent manner. PAP reduced ascorbic acid and cholesterol contents in adrenal glands and decreased the serum hydrocortisone level of rats. The reduction of ascorbic acid and cholesterol contents were unaffected by the pretreatment of dexamethasone. PAP also showed an anti-inflammatory action on the swelling of carrageenan-induced hind paw in adrenalectomized rats. The results suggest that the anti-inflammatory effects of PAP do not depend absolutely on pituitary-adrenal system.

Purification and partial characterization of anti-inflammatory peptide from pilose antler of Cervus nippon Temminck.

An anti-inflammatory compound was purified and isolated from pilose antler of Cervus nippon Temminck by dialysis, gel filtration and ion-exchange chromatography techniques. HPLC and N-terminal amion acid analysis identified the compound as a homogeneous peptide. The peptide is composed of 68 amino acids and its molecular weight as determined by amino analysis, is about 7200.

Comparison of anti-lipid peroxidative effects of the underground parts of Notopterygium incisum and N. forbesii in mice.

Intraperitoneal administration of CCl4 to mice led to significant increases of thiobarbituric acid-reactive substances (TBA-RS), free malondialdehyde (MDA), lipid conjugated dienes and fluorescent lipid peroxidation products in the liver. However, subchronic pretreatment with oral doses of the MeOH extract of either the underground part of Notopterygium incisum or that of N. forbesii appreciably suppressed the formation of CCl4-induced lipid peroxidation products. The suppressing potency was more remarkable in the former.

[Effects of the ginseng polysaccharides on reducing liver glycogen].

The ginseng polysaccharides GH1 (100, 200 mg.kg-1) iv reduced liver glycogen and increased adenosine-3',5'-cyclic monophosphate (cAMP) level and adenyl cyclase (AC) activity in mice. The action of GH1 was completely antagonized by propranolol, inhibitor of adrenergic beta receptor. The stimulating effect of GH1 on AC activity was significant 2 and 4 h after iv GH1. However, GH1 at concentration of 20-120 mumol.L-1 in vitro showed no manifest effect on AC activity. GH1 stimulated the activities of 3',5'-cyclic-GMP phosphodiesterases (PDE) and calmodulin (CaM) in a dose-dependent manner. It is suggested that the reduction of liver glycogen induced by GH1 resulted from its obvious increase of cAMP which promoted glycogenolysis and decreased glycogenesis.

[Effects of ginseng polysaccharides on reducing blood glucose and liver glycogen].

Ginseng polysaccharides (GH1) 50-200 mg/kg ip or sc reduced blood glucose and liver glycogen of mice. Adrenalectomy did not affect this action. GH1 increased the content of pyruvic acid, but decreased the content of lactic acid by weakening the activity of lactate dehydrogenase. GH1 accelerated oxidative-phosphorylation of carbohydrate since the activities of succinate dehydrogenase (SDH) and cytochrome oxidase (CCO) were obviously stimulated. Besides the promotion of the activity of SDH in human embryonic lung fibroblasts (HELF), GH1 decreased the content of polysaccharides in HELF of the 24th age generation, but increased that of the 40th age generation. On the other hand, GH1 stimulated the release of insulin. It is suggested that the reduction of blood glucose and liver glycogen induced by GH1 be primarily due to the increase of carbohydrate utilization and the decrease of glycogenesis.

[Studies on the hypoglycemic effect of ginseng polypeptide].

Ginseng polypeptide (GPP) isolated from the root of Panax ginseng C. A. Meyer was demonstrated to decrease the levels of blood sugar and liver glycogen when injected intravenously to rats at doses of 50-200 mg/kg without affecting total blood lipid. when mice were injected subcutaneously daily at doses of 50 and 100 mg/kg for 7 successive days, GPP was also found to decrease blood glucose and liver glycogen. In addition, GPP was found to decrease various experimental hyperglycemia induced by injection of adrenaline, glucose and alloxan. However, GPP exhibited inhibiting effect on the glycogen enhancement induced by glucose, but strengthening effect on the glycogen decrease induced by adrenaline. When the levels of blood total lipid and liver glycogen were increased by alloxan, GPP was shown to inhibit these changes except its lowering of blood sugar. The toxicity of GPP is very low, its iv LD50 for mice was found to be 1.62 +/- 0.130 g/kg.

[Influence of the active compounds isolated from pilose antler on syntheses of protein and RNA in mouse liver].

The polyamines of pilose antler (PASPA) consist of putrescine (PU, 70.9%), spermidine (SPD, 26.3%) and spermine (SP, 2.8%). The incorporations of [3H] leucine into protein and [3H] uridine into RNA in mouse liver tissue were increased when PASPA was given orally to mice at the dose of 30 mg/kg for 4 successive days. The incorporations of [3H] leucine into liver protein and [3H] uridine into the cytosolic and nuclear RNA were also increased by treatment with PU (21 mg/kg). In addition, the RNA polymerase activity in the solubilized liver nuclear fraction of PU (21 mg/kg)-treated mice was increased. SPD only promoted the synthesis of protein in mouse liver tissue at the dose of 8 mg/kg. However, SP showed no effect on the synthesis of protein and RNA polymerase activity under the used dose (1 mg/kg). The results suggest that PASPA is the main active substance responsible for the promotion of the synthesis of protein and RNA in mouse liver.

[Studies on the mechanism of ginseng polypeptide induced hypoglycemia].

When ginseng polypeptide (GP) was administered iv or sc to mice or rats, the blood sugar and liver glycogen were decreased. The decrease of blood sugar and liver glycogen induced by GP in rats were inhibited by pretreatments with pentolamine and propranolol, respectively. The results suggest that the effect of GP on glucose metabolism may be related to adrenergic receptors. In addition, GP at doses which cause decreases of blood sugar and glycogen, inhibited LDH activity, and consequently produced decrease of lactic acid and increase of pyruvic acid. GP was also found to stimulate SDH and CCO activities (which are respiratory enzymes in the electron transport chain), consequently promote aerobic oxidation course.

[A tablet of Tripterygium wilfordii in treating lupus erythematosus].

Twenty-three cases of lupus erythematosus, including 15 cases of SLE and 8 cases of DLE treated with Tripterygium wilfordii (TW, three tablets thrice a day, each tablet contains 5 gm of crude TW) were reported. As controls, 19 cases of SLE were treated with prednisone alone at the same time. 9/15 cases in the former group and 6/19 in the latter got improvement after treatment. There was no significant difference between the two groups (P greater than 0.10). However, TW had beneficial effects to remission of arthralgia (7/10 in the former group and 7/15 in the latter group) and erythematosus rash (6/8 in the former group and 5/12 in the latter). Clinically, four out of eight cases of DLE improved after treatment with TW. TW had some toxic effects, such as decrease of peripheral blood leukocyte and kidney damage. Since the toxic dose and effective dose could be quite close, special attention should be paid when the drug was being used, especially on those patients with nephropathy. So the authors suggested that the indication of TW could include the patients of DLE and SLE without nephropathy.