Metabolism of cystathionine, N-monoacetylcystathionine, perhydro-1,4-thiazepine-3,5-dicarboxylic acid, and cystathionine ketimine in the liver and kidney of D,L-propargylglycine-treated rats

Experimental cystathioninuria was induced by injection of D,L-propargylglycine in rats. The novel cystathionine metabolites, N-monoacetylcystathionine (NAc-cysta), perhydro-1,4-thiazepine-3,5-dicarboxylic acid (PHTZDC), and cystathionine ketimine (CK), were identified previously in the urine of patients with cystathioninuria and D,L-propargylglycine-treated rats. In this study, we identified these compounds in the liver and kidney of D,L-propargylglycine-treated rats using liquid chromatography-mass spectrometry with an atmospheric pressure chemical ionization interface system (LC/APCI-MS) and an amino acid analyzer. The metabolism of these compounds in the liver and kidney of D,L-propargylglycine-treated rats was also studied. PHTZDC, NAc-cysta, and CK were accumulated in the rat tissues in proportion to the content of cystathionine after D,L-propargylglycine administration. The concentrations of these compounds in the liver were higher than those in the kidney, and these compounds reached maxima earlier in the liver than in the kidney.     

Isolation and identification of mercapturic acids of cinnamic aldehyde and cinnamyl alcohol from urine of female rats

Rats dosed with cinnamic aldehyde (I) excreted two mercapturic acids in the urine. The major one was identified as N-acetyl-S-(1-phenyl-3-hydroxypropyl)cysteine (V). The minor one was identified as N-acetyl-S-(1-phenyl-2-carboxy ethyl)cysteine (VI). The ratio appeared to be V : VI = 4 : 1. The hydroxy mercapturic acid (V) was also isolated from urine of rats dosed with cinnamyl alcohol (II). The total mercapturic acid excretion as percentage of the dose was 14.8 +/- 1.9% for cinnamic aldehyde (250 mg/kg) (n = 4) and 8.8 +/- 1.7% for cinnamyl alcohol (n = 4) (125 mg/kg). Inhibition of the alcohol dehydrogenase by pyrazole (206 mg/kg) diminished the thioether excretion of cinnamyl alcohol to 3.3 +/- 1.4% of the dose (n = 8). Cinnamic aldehyde has been proposed to be an intermediate in the mercapturic acid formation of cinnamyl alcohol.     

A study of metabolites isolated from the urine samples of cats and dogs administered orbifloxacin

Urine samples of cats and dogs collected for 24 hr after a subcutaneous injection of orbifloxacin (OBFX) were analyzed. The metabolites were examined using HPLC. In the dog urine, 87% of total was the parent compound and 13% glucuronide compound of OBFX and 96% was parent and 4% metabolite in the cat urine. The metabolite of cat urine was identified as N-hydroxy OBFX, determined by comparison of the extraction of urine with chloroform with the standard compound of N-hydroxy OBFX, using LC/APCIMS. N-hydroxy OBFX had a weaker antibacterial activity against fluoroquinolone sensitive bacteria than the parent compound.     

A new oxisuran metabolite

1. An unidentified oxisuran metabolite which had been observed in animal urine was biosynthesized by incubating [14C]oxisuran with rat liver cytosol. 2. The metabolite, isolated by preparative t.l.c. and extraction, was identified as oxisuran alcohol sulphide by mass fragmentography. Confirmation of this identification was obtained by biosynthesis of the same compound from oxisuran sulphide. 3. The 9000 g supernatant liquid from rat liver was less effective than cytosol in reducing oxisuran to its alcohol sulphide. Neither rat liver fraction reduced oxisuran alcohol sulphoxides to sulphide. 4. The 9000 g fraction oxidized oxisuran and oxisuran alcohol sulphoxide to oxisuran alcohol sulphone.     

Structure of the ring cleavage product of 1-hydroxy-2-naphthoate, an intermediate of the phenanthrene-degradative pathway of Nocardioides sp. strain KP7

1-Hydroxy-2-naphthoate (compound I) is a metabolite of the phenanthrene-degradative pathway in Nocardioides sp. strain KP7. This singly hydroxylated aromatic compound is cleaved by 1-hydroxy-2-naphthoate dioxygenase. In this study, the structure of the ring cleavage product generated by the action of homogeneous 1-hydroxy-2-naphthoate dioxygenase was determined upon separation by high-performance liquid chromatography at pH 2.5 by using nuclear magnetic resonance (NMR) and mass spectroscopic techniques. The ring cleavage product at this pH existed in equilibrium between two forms, 2-oxo-3-(3-oxo-1,3-dihydro-1-isobenzofuranyl)propanoate (compound III) and 2,2-dihydroxy-3-(3-oxo-1, 3-dihydro-1-isobenzofuranyl)propanoate (compound IV). After the pH of the solution was raised to 7.5, the structure of the major species became (E)-4-(2-carboxylatophenyl)-2-oxo-3-butenoate (compound II; common name, trans-2'-carboxybenzalpyruvate), which was in equilibrium with compound III. Direct monitoring of the enzymatic formation of the ring cleavage product by 1H-NMR in a deuterated potassium phosphate buffer (pH 7.5) detected only compound II as a product, and the proton on carbon 3 of compound II was not exchanged with deuterium. Thus, compound II is likely to be the first stable product of dioxygenation of 1-hydroxy-2-naphthoate.     

Psychosocial aspects of palliative care

The metabolites of [2,3-14C]acrolein in the urine and feces of Sprague-Dawley rats were identified after either intravenous administration in saline at 2.5 mg/kg or oral administration by gavage as an aqueous solution as either single or multiple doses at 2.5 mg/kg or as a single dose of 15 mg/kg. Selected urine and feces samples were pooled by sex and collection interval and profiled by combinations of reverse-phase, anion-exchange, cation-exchange, and ion-exclusion high-performance liquid chromatography (HPLC). Feces were also profiled by size-exclusion chromatography. Metabolites were identified by comparison with well-characterized standards by HPLC and by mass spectrometry. The urinary metabolites were identified as oxalic acid, malonic acid, N-acetyl-S-2-carboxy-2-hydroxyethylcysteine, N-acetyl-S-3-hydroxypropylcysteine, N-acetyl-S-2-carboxyethylcysteine, and 3-hydroxypropionic acid. The fecal radioactivity from the oral dose groups was partitioned into methanol-soluble, water-soluble, and insoluble radioactivity, some of which could be liberated by dilute acid hydrolysis. HPLC analysis of these extracts revealed no discrete metabolites. Size-exclusion chromatography indicated a molecular weight range of 2,000 to 20,000 Da for the radioactivity, which was unaffected by hydrolysis at reflux with 6 M acid or base. This radio-activity was thought to be a homopolymer of acrolein, which was apparently formed in the gastrointestinal tract. The pathways of acrolein metabolism were epoxidation followed by conjugation with glutathione, Michael addition of water followed by oxidative degradation, and glutathione addition to the double bond either following or preceding oxidation or reduction of the aldehyde. The glutathione adducts were further metabolized to the mercapturic acids.     

A novel melatonin metabolite, cyclic 3-hydroxymelatonin: a biomarker of in vivo hydroxyl radical generation

In the current study, we characterized a urinary melatonin metabolite which could provide a safe and effective method to monitor generation of HO* in humans. Using mass spectrometry (MS), proton nuclear magnetic resonance (1H NMR), COSY 1H NMR analysis, and calculations on the relative thermodynamic stability, a novel melatonin metabolite was identified as cyclic 3-hydroxymelatonin (3-OHM). 3-OHM is the product of the reaction of melatonin with HO* which was generated in two different cell-free in vitro systems. Interestingly, this same metabolite, 3-OHM, was also identified in the urine of both rats and humans. A proposed reaction pathway suggests that 3-OHM is the footprint product that results when a melatonin molecule scavenges two HO*. When rats were challenged with ionizing radiation which results in HO* generation, urinary 3-OHM increased dramatically compared to that of controls. These results strongly indicate that the quantity of 3-OHM produced is associated with in vivo HO* generation. Since melatonin exists in virtually all animal species and has a wide intracellular distribution and 3-OHM is readily detected noninvasively in urine, we suggest that 3-OHM is a valuable biomarker that can be used to monitor in vivo HO* levels in humans and other species. The measurement of urinary 3-OHM as a biomarker of HO* generation could provide clinical benefits in the diagnosis and treatment of diseases.     

Inactivation of 4-oxalocrotonate tautomerase by 2-oxo-3-pentynoate

The compound, 2-oxo-3-pentynoate, has been synthesized and tested as an inhibitor of the enzyme 4-oxalocrotonate tautomerase. The enzyme is rapidly and irreversibly inactivated by the acetylenic product analogue in a time-dependent fashion. The enzyme displays saturation kinetics and is protected from inactivation by the presence of substrate. These observations are consistent with inactivation taking place at the active site. Partial reactivation ( approximately 18%) occurs by incubating the inactivated enzyme with 10 mM hydroxylamine (pH 7.3). The partition ratio, determined to be approximately 0.4, suggests that the inactivation of 4-OT by 2-oxo-3-pentynoate shows half-of-the-sites stoichiometry. The same phenomenon is observed in the inactivation of 4-OT by 3-bromopyruvate and can be explained by examination of the crystal structure. Mass spectral analysis shows that a single residue is modified on the enzyme which has been localized to the nine residue amino-terminal fragment Pro-1 to Glu-9. It can be reasonably concluded that Pro-1 is the site of covalent attachment. Inactivation of 4-OT can occur by either a Michael addition of 4-OT to C-4 of 2-oxo-3-pentynoate or by the enzyme-catalyzed rearrangement of 2-oxo-3-pentynoate to an allene derivative which alkylates Pro-1. These results provide the foundation for the use of 2-oxo-3-pentynoate in future mechanistic studies and as a ligand in an inactivated 4-OT complex that can be studied by X-ray crystallography. Finally, 2-oxo-3-pentynoate is an acetylene analogue of a variety of 2-oxo acids and as such may have general utility as an inhibitor of reactions that bind and process these compounds.     

Role of renal cysteine conjugate beta-lyase in the mechanism of compound A nephrotoxicity in rats

BACKGROUND: The sevoflurane degradation product compound A is nephrotoxic in rats, in which it undergoes extensive metabolism to glutathione and cysteine S-conjugates. The mechanism of compound A nephrotoxicity in rats is unknown. Compound A nephrotoxicity has not been observed in humans. The authors tested the hypothesis that renal uptake of compound A S-conjugates and metabolism by renal cysteine conjugate beta-lyase mediate compound A nephrotoxicity in rats. METHODS: Compound A (0-0.3 mmol/kg in initial dose-response experiments and 0.2 mmol/kg in subsequent inhibitor experiments) was administered to Fischer 344 rats by intraperitoneal injection. Inhibitor experiments consisted of three groups: inhibitor (control), compound A, or inhibitor plus compound A. The inhibitors were probenecid (0.5 mmol/kg, repeated 10 h later), an inhibitor of renal organic anion transport and S-conjugate uptake; acivicin (10 mg/kg and 5 mg/kg 10 h later), an inhibitor of gamma-glutamyl transferase, an enzyme that cleaves glutathione conjugates to cysteine conjugates; and aminooxyacetic acid (0.5 mmol/kg and 0.25 mmol/kg 10 h later), an inhibitor of renal cysteine conjugate beta-lyase. Urine was collected for 24 h and then the animals were killed. Nephrotoxicity was assessed by light microscopic examination and biochemical markers (serum urea nitrogen and creatinine concentration, urine volume and urine excretion of protein, glucose, and alpha-glutathione-S-transferase [alpha GST], a marker of tubular necrosis). RESULTS: Compound A caused dose-related nephrotoxicity, as shown by selective proximal tubular cell necrosis at the corticomedullary junction, diuresis, proteinuria, glucosuria, and increased alpha GST excretion. Probenecid pretreatment significantly (P < 0.05) diminished compound A-induced increases (mean +/- SE) in urine excretion of protein (45.5 +/- 3.8 mg/24 h vs. 25.9 +/- 1.7 mg/24 h), glucose (28.8 +/- 6.2 mg/24 h vs. 10.9 +/- 3.2 mg/24 h), and alpha GST (6.3 +/- 0.8 micrograms/24 h vs. 1.0 +/- 0.2 microgram/24 h) and completely prevented proximal tubular cell necrosis. Aminooxyacetic acid pretreatment significantly diminished compound A-induced increases in urine volume (19.7 +/- 3.5 ml/24 h vs. 9.8 +/- 0.8 ml/24 h), protein excretion (37.2 +/- 2.7 mg/24 h vs. 22.2 +/- 1.8 mg/24 h), and alpha GST excretion (5.8 +/- 1.5 vs. 2.3 micrograms/24 h +/- 0.8 microgram/24 h) but did not significantly alter the histologic pattern of injury. In contrast, acivicin pretreatment increased the compound A-induced histologic and biochemical markers of injury. Compound A-related increases in urine fluoride excretion, reflecting compound A metabolism, were not substantially altered by any of the inhibitor treatments. CONCLUSIONS: Intraperitoneal compound A administration provides a satisfactory model of nephrotoxicity. Aminooxyacetic acid and probenecid significantly diminished histologic and biochemical evidence of compound A nephrotoxicity, whereas acivicin potentiated toxicity. These results suggest that renal uptake of compound A-glutathione or compound A-cysteine conjugates and cysteine conjugates metabolism by renal beta-lyase mediate, in part, compound A nephrotoxicity in rats.     

Absorption, disposition kinetics, and metabolic pathways of cyclohexene oxide in the male Fischer 344 rat and female B6C3F1 mouse

Cyclohexene oxide (CHO) is a monomer intermediate used in the synthesis of pesticides, pharmaceuticals, and perfumes. Although CHO has a variety of industrial uses where direct human exposure is possible, very little is known about its fate in the body. Therefore, the objectives of this study were to determine the absorption, distribution, metabolism, and excretion of cyclohexene oxide after oral, intravenous, and dermal exposure in male Fischer 344 rats and female B6C3F, mice. After intravenous administration of [14C]CHO (50 mg/kg), CHO was rapidly distributed, metabolized, and excreted into the urine. Plasma concentrations of CHO rapidly declined and were below the limit of detection within 60 min. Average (+/- SD) values for terminal disposition half-life, apparent volume of distribution at steady-state, and systemic body clearance were: 19.3 +/- 1.6 min; 0.44 +/- 0.08 liter/kg; and 31.3 +/- 0.5 ml/kg * min, respectively. After oral administration of [14C]CHO (10 and 100 mg/kg), it was found that 14C-equivalents were rapidly excreted in the urine of both species. At 48 hr, the majority of the dose (73-93%) was recovered in urine, whereas fecal elimination accounted for only 2-5% of the dose. At no time after oral administration was parent CHO detected in the blood. However, its primary metabolite cyclohexane-1,2-diol was present for different lengths of time depending on the dose. Four metabolites were detected and identified in mouse urine by MS: cyclohexane-1,2-diol; cyclohexane-1,2-diol-O-glucuronide; N-acetyl-S-(2-hydroxycyclohexyl)-L-cysteine; and cyclohexane-1,2-diol-O-sulfate. The sulfate conjugate was not present in rat urine. Topical application of [14C]CHO (60 mg/kg) provided poor absorption in both species. The majority of 14C-equivalents applied dermally were recovered from the charcoal skin trap (approximately 90% of the dose). Only 4% of the dose was absorbed, and the major route of elimination was via the urine. To evaluate the toxicity of CHO, animals were given daily doses of CHO orally and topically for 28 days. No statistically significant changes in final body weights or relative organ weights were noted in rats or mice treated orally with CHO up to 100 mg/kg or up to 60 mg/kg when given topically. Very few lesions were found at necropsy, and none were considered compound related. In conclusion, regardless of route, CHO is rapidly eliminated and excreted into the urine. Furthermore, after either oral or dermal administration, it is unlikely that CHO reaches the systemic circulation intact due to its rapid metabolism, and is therefore unable to cause toxicity in the whole animal under the test conditions used in this study.     

Photochemotherapy: identification of a metabolite of 4, 5', 8-trimethylpsoralen

A compound, 4, 8-dimethyl, 5'-carboxypsoralen (DMeCP), has been identified in mouse urine as a major metabolite of the photoactive drug, 4, 5', 8-trimethylpsoralen (TMeP). This drug is widely used in the treatment of vitiligo and psoriasis. DMeCP is fluorescent, and nonphotosensitizing when tested on guinea pig skin. DMeCP also occurs in the urine of human patients receiving TMeP orally.     

Study on major metabolites of 3H-1,2-dihydro-2-(4-methylphenylamino)-methyl-1-pyrrolizinone in rabbits]

The metabolites of a 750 mg oral dose of Z-47 [3H-1, 2-dihydro-2-(4-methylphenylamino) methyl-1-pyrrolizinone], a new anti-inflammatory and analgesic agent, in rabbit urine were separated and detected with high performance liquid chromatographic method. On basis of the chromatographic behavior of Z-47 metabolites and biotransformation pathways of drugs with partial structure of Z-47, the carboxylic derivative of Z-47 [4-(3H-1, 2-dihydro-1-pyrrolizinone-2-methylamino) benzoic acid] was proposed as a potential metabolite so that the compound was synthesized. The authentic substance was then compared with one of the metabolites by the chromatographic retention time and the ratio of their UV-absorbances at two wavelengths. The enzyme-hydrolyzed product of another metabolite was also analysed. It was consequently confirmed that the carboxylic derivative of Z-47 and its acyl beta-D-glucuronide are major metabolits of Z-47 in rabbits.     

The primitive protozoon Trichomonas vaginalis contains two methionine gamma-lyase genes that encode members of the gamma-family of pyridoxal 5'-phosphate-dependent enzymes

Methionine gamma-lyase, the enzyme that catalyzes the breakdown of methionine by an alpha,gamma-elimination reaction and is a member of the gamma-family of pyridoxal 5'-phosphate-dependent enzymes, is present in high activity in the primitive protozoan parasite Trichomonas vaginalis but is absent from mammals. Two genes, mgl1 and mgl2, encoding methionine gamma-lyase, have now been isolated from T. vaginalis. They are both single copy, encode predicted proteins (MGL1 and MGL2) of 43 kDa, have 69% sequence identity with each other, and show a high degree of sequence identity to methionine gamma-lyase from Pseudomonas putida (44%) and other related pyridoxal 5'-phosphate-dependent enzymes such as human cystathionine gamma-lyase (42%) and Escherichia coli cystathionine beta-lyase (30%). mgl1 and mgl2 have been expressed in E. coli as a fusion with a six-histidine tag and the recombinant proteins (rMGL1 and rMGL2) purified by metal-chelate affinity chromatography. rMGL1 and rMGL2 were found to have high activity toward methionine (10.4 and 0.67 mumol/min/mg of protein, respectively), homocysteine (370 and 128 mumol/min/mg of protein), cysteine (6.02 and 1.06 mumol/min/mg of protein), and O-acetylserine (3.74 and 1.51 mumol/min/mg of protein), but to be inactive toward cystathionine. Site-directed mutagenesis of an active site cysteine (C113G for MGL1 and C116G for MGL2) reduced the activity of the recombinant enzymes toward both methionine and homocysteine by approximately 80% (rMGL1) and 90% (rMGL2). In contrast, the activity of mutated rMGL2 toward cysteine and O-acetylserine was increased (to 214 and 142%, respectively), whereas that of mutated rMGL1 was reduced to 39 and 49%, respectively. These findings demonstrate the importance of this cysteine residue in the alpha,beta-elimination and alpha, gamma-elimination reactions catalyzed by trichomonad methionine gamma-lyase.     

Metabolism of Ro 21-5998 (mefloquine) in the rat (author's transl)

After administration of 15 mg/kg 14C-Ro 21-5998/001 i.p. to rats, the metabolite patterns in feces, urine, bile and blood were compared. Metabolites from feces were identified by GC/MS, in all other cases by TLC. The main component in the feces consists of mefloquine (Ro 21-5998). In addition the acid Ro 21-5104, a derivative of mefloquine with a hydroxy group in the piperidine moiety (M 12), the alcohol Ro 14-0518 and a metabolite (M 4a), which is supposed to be a lactam, were shown to be present. The acid Ro 21-5104 is the main metabolite in the urine. The bile contains the parent compound Ro 21-5998, the acid Ro 21-5104 and the alcohol Ro 14-0518, partially as conjugates. The structurally investigated components account for about 40% of the administered dose. The blood contains the parent compound Ro 21-5998 and the acid Ro 21-5104 as main components. In comparing the various metabolite patterns, it can be summarized that that in urine is slightly different from those in bile (after hydrolysis of the conjugates), feces and blood which show more similarities between each other.     

Biotransformation of the naturally occurring isothiocyanate sulforaphane in the rat: identification of phase I metabolites and glutathione conjugates

Sulforaphane (SFN) is a naturally occurring isothiocyanate present in cruciferous vegetables, such as broccoli, that has been identified as a potent inducer of glutathione S-transferase activities in laboratory animals. The present studies were carried out to elucidate the metabolic fate of SFN in the rat. Particular emphasis was placed on glutathione (GSH)-dependent pathways because conjugation with GSH is a major route by which many isothiocyanates are eliminated in mammals. Male Sprague-Dawley rats were administered a single dose of SFN (50 mg kg-1 ip), and bile and urine were collected over ascorbic acid. Analysis of biological fluids was carried out by ionspray LC-MS/MS using the neutral loss (129 Da) and precursor ion (m/z 164) scan modes to detect GSH and N-acetylcysteine (NAC) conjugates, respectively. In bile, five thiol conjugates (designated M1-M5) were detected. Metabolites M2 and M4 were identified as the GSH conjugates of SFN and erucin (ERN, the sulfide analog of SFN), respectively, by comparing their LC-MS/MS properties with those of standards obtained by synthesis. M1 was characterized as the GSH conjugate of a desaturated metabolite of SFN (tentatively assigned the structure of delta 1-SFN), suggesting that the parent compound also undergoes oxidative metabolism. Metabolites M3 and M5 were identified as the NAC conjugates of SFN and ERN, respectively, and together with the NAC conjugate of delta 1-SFN, these species also were detected in urine. Quantitative determination of the former two mercapturates in urine indicated that approximately 60% and approximately 12% of a single dose of SFN is eliminated in 24 h as the NAC conjugates of SFN and ERN, respectively. The corresponding figures in rats dosed with ERN were approximately 67% and approximately 29%. When the GSH conjugate of SFN was incubated with phosphate buffer (pH 7.4, 37 degrees C), < 1% of the conjugate dissociated to liberate free SFN. On the other hand, the conjugate underwent a facile thiol exchange reaction (> 70% conversion) when incubated in the presence of excess cysteine, thereby acting as an effective carbamoylating agent. It is concluded that SFN undergoes metabolism by S-oxide reduction and dehydrogenation and that GSH conjugation is the major pathway by which the parent compound and its phase I metabolites are eliminated in the rat.     

The interaction of myristylated peptides with the catalytic domain of protein kinase C revealed by their sequence palindromy and the identification of a myristyl binding site

The nature of products of contamination intake were investigated in cattle dosed with [14C]di-n-butylphthalate (DBP). Radio-labelled metabolites were extracted from bile, faeces, plasma and urine onto solid-phase media, fractionated by ion-exchange chromatography, separated by reverse phase HPLC and analysed by negative ion atmospheric pressure chemical ionization mass spectrometry(n) (LCQ, Finnigan). All matrices contained a common major metabolite [deprotonated molecular ion (M-H)- m/z 221] which coeluted with and had an identical daughter ion spectrum to reference monobutylphthalate (MBP). MBP was metabolised to a beta-glucuronidase sensitive compound (M-H)- m/z 397 whose spectrum contained daughter ions (m/z 175 and 221) consistent with the parent glucuronide. A further three beta-glucuronidase resistant radio-labelled metabolites were also produced (M-H- m/z 165, 193 and 237); comparison of daughter ion spectra with those of reference MBP and phthalic acid indicated identity with phthalic acid, monoethylphthalate (MEP) and monohydroxybutylphthalate (MHBP) respectively. The presence of a benzoate daughter ion (m/z 121) in all spectra was indicative of side chain biotransformation. Both MBP and MEP contained a phthalate daughter ion (m/z 165) indicating loss of a butyl and ethyl side chain respectively. A daughter ion of m/z 89 derived from the side chain provided evidence that the third metabolite was MHBP. Incubation of DBP with isolated bovine hepatocytes produced the same metabolites and provided relatively clean samples for LC/MSn analysis. Detection of these DBP metabolites in meat or dairy food products will provide evidence for environmental exposure and biotransformation in vivo, whereas the presence of the parent compound would suggest contamination during food processing and packaging.     

Reactivity of haloketenes and halothioketenes with nucleobases: reactions in vitro with DNA

A novel secondary metabolite SCH 42282 (1), with antifungal activity was isolated from the fermentation broth of a soil actinomycete identified as a Microtetraspora sp. The active compound was separated from the fermentation broth by butanol extraction and purified on a silica gel column and by multi-coil counter current chromatography. The compound was identified as a novel macrolactam trisaccharide related to SCH 38518 (4). The structure was established by hydrolysis of the parent compound and spectroscopic studies of the acetate derivative. The compound is active against Candida spp. (Geometric Mean MIC's. 18 micrograms/ml) but less active SCH 42729 (3). the disaccharide (Geometric Mean MIC's, > or = 10.7 micrograms/ml and SCH 38518 (4), the monosaccharide (Geometric Mean Mic's, 3.8 micrograms/ml.     

Action of DNA-gyrase inhibiting derivatives of 4-oxo-1, 4-dihydro-3-pyridinecarboxylic acid against Trypanosoma brucei

Ten derivatives of 4-oxo-1,4-dihydro-3-pyridinecarboxylic acid as DNA-gyrase inhibitors were evaluated in vitro and in vivo against Trypanosoma brucei brucei IPP. Two compounds were were active at 100 microM in vitro after 1 h incubation time. After 24 h incubation, 8 compounds were active and the most interesting compound, 1-(4-hydroxy-2-methyl-phenyl)-6-[2-(4,5-dichloro-phenyl)-ethenyl] -4-oxo-1, 4-dihydropyridine-3-carboxylic acid (compound No. 8) was trypanocidal at 10 microM. The trypanocidal effects were not noted in vivo after subcutaneous treatment administered as a single dose of 50 mumols/kg. The in vitro trypanocidal effect is correlated with the DNA-gyrase inhibition.     

In vivo evaluation of the Lillehei-Kaster heart valve prosthesis

Three decomposition products of Morazone (ingredient of the pharmaceutical preparation Rosimon-Neu) were observed following heat treatment in acid medium (hydrochloric or tartaric acid). These products were isolated by TLC and identified as bis-antipyryl-methane, phenmetrazine and 4-hydroxymethyl-antipyrine by mass spectrometry and IR-spectroscopy and 1H-nuclear magnetic resonance. Morazone and the metabolite phenmetrazine may be extracted from alkaline urine using chloroform, however acid hydrolysis (pH 1) of the urine before alkaline extraction will improve the sensitivity of detection of morazone by producing the metabolite phenmetrazine in addition to bis-antipyrylmethane. The metabolite 4-hydroxymethyl-antipyrine is barely detectable by TLC from alkaline extraction of urine.     

Study of factors affecting the determination of total plasma 7-fluorobenzo-2-oxa-1,3-diazole-4-sulfonate (SBD)-thiol derivatives by liquid chromatography

A flunixin metabolite, a hydroxylated product, has been identified in camel urine and plasma samples using gas chromatography-mass spectrometry (GC-MS) and GC-MS-MS in the electron impact and chemical ionization modes. Its major fragmentation pattern has been verified by GC-MS-MS in daughter ion and parent ion scan modes. The method could detect flunixin and its metabolite in camel urine after a single intravenous dose of 2.2 mg of flunixin/kg body weight for 96 and 48 h, respectively, which increases the reliability of antidoping control analysis.