K.H. Lee

Inhibition of inosine monophosphate dehydrogenase by sesquiterpene lactones

Inosine monophosphate (IMP) dehydrogenase had previously been determined to be a likely target enzyme for the sesquiterpene lactones, a class of potential anti-neoplastic drugs. IMP dehydrogenase was purified approx. 770-fold from the P-388 lymphocytic leukemia tumor cell line. The Km values for the substrates, IMP and NAD, were determined to be 12 microM and 25 microM, respectively. Xanthine monophosphate (XMP) was shown to be a competitive inhibitor with a Ki of 67 microM. Mycophenolic acid gave mixed-type inhibition with a Ki of 8 nM for the noncompetitive component and a Ki of 2 nM for the competitive component. Dissociation constants (Kd) and rate constants for inhibition of IMP dehydrogenase by nine different sesquiterpene lactones were determined. The highest Kd was seen with 2,3-dihydrohelenalin while the lowest Kd was observed with bis-helenalinyl malonate. Binding of the drugs by IMP dehydrogenase increased as the size of the drug increased. Also, changes in structure at position 6 had a relatively large effect on the Kd. There was no correlation with hydrophobicity, as determined by octanol/water partition. The first-order rate constants for the reaction of the sesquiterpene lactones with IMP dehydrogenase (k1) and the second-order rate constants for the reaction of the sesquiterpene lactones with glutathione (k2) were also determined. The rate constants for most of the sesquiterpene lactones with the alpha-methylene-gamma-lactone moiety were similar and were approximately twice as great as the rate constants for those sesquiterpene lactones with only the alpha, beta-unsaturated cyclopentenone ring. Microlenin had approximately 5-times the reactivity of the other sesquiterpene lactones towards IMP dehydrogenase, but had approximately the same reactivity towards glutathione, suggesting that it was bound to the enzyme in a way which facilitated its reaction with one or more essential sulfhydryls. The same procedure was used for a series of N-substituted maleimide compounds with the N-substituent ranging in size from a methyl group to a benzyl group. The binding of the maleimide compounds was generally tighter than for the sesquiterpene lactones and there was an increase in binding with size.

Regular articleAcute toxicity of helenalin in BDF1 mice

The acute toxicity of helenalin, a sesquiterpene lactone isolated from Helenium microcephalum, was examined in male BDF1 mice. The 14-day LD50 for a single ip dose of helenalin in male mice was 43 mg/kg. A single ip injection of 25 mg helenalin/kg increased serum alanine aminotransferase (ALT), lactate dehydrogenase (LDH), urea nitrogen (BUN), and sorbitol dehydrogenase within 6 hr of treatment. Multiple helenalin exposures, ip injection of 25 mg helenalin/kg for 3 days, increased differential polymorphonuclear leukocyte counts and decreased lymphocyte counts. Serum ALT, BUN, and cholesterol levels were also increased by multiple helenalin exposures at 25 mg helenalin/kg/day. Helenalin significantly reduced liver, thymus, and spleen relative weights and histologic evaluation revealed substantial effects of multiple helenalin exposures on lymphocytes of the thymus, spleen, and mesenteric lymph nodes. No helenalin-induced histologic changes were observed in the liver or kidney. Multiple helenalin exposures (25 mg/kg/day) significantly inhibited hepatic microsomal enzyme activities (aminopyrine demethylase and aniline hydroxylase) and decreased microsomal cytochromes P-450 and b5 contents. Three concurrent days of diethyl maleate (DEM) pretreatment (3.7 mmol DEM/kg, 0.5 hr before helenalin treatment) significantly increased the toxicity of helenalin exposure. The present studies indicate that the hepatic microsomal drug metabolizing system and lymphoid organs are particularly vulnerable to the effects of helenalin. In addition, helenalin toxicity is increased by DEM pretreatments which have been shown to decrease glutathione concentrations.

Antiviral Activity of Dammarane Saponins Against Herpes Simplex Virus I

AbstractTwo dammarane saponins, ginsenoside Rb1 (GRb1) and chikusetsusaponin III (Chi III), were investigated for antiviral activity against herpes simplex virus type I using an in vitro plaque elimination assay. Chi III prevented plaque formation at 50 μM, while GRb1 had no effect at 100 μM. Chi III was further examined for effects on virus yield and for cytotoxicity. The ID50 value for virus yield reduction was 29 μM, and cell growth was inhibited by 50% at 66 μM, indicating that Chi III exhibited marginal antiviral selectivity with a therapeutic index of about 2-fold. Although Chi III exhibited an in vitro virucidal effect with an ID50 value of 62 μM, drug treatment for 24 h reduced production of the cell-associated virus as well as the virus present in the medium, indicating Chi III has intracellular inhibitory activity. Intracellular levels of viral proteins were marginally affected at five hours post-infection, indicating Chi III has a minor impact on the early events of infection.

Anti-Inflammatory Agents. V. Amylose from Pinellia ternata

AbstractAn amylose was isolated from “Ban Xia”, the dried tuber of Pinellia ternata, which demonstrated marked anti-inflammatory activity in mice at 25 mgAg. This amylose demonstrated nmr characteristics consistent with a standard amylose of a molecular weight of 4,100, which also demonstrated antiinflammatory activity in the same potency range as the isolated product from “Ban Xia” in mice.