Single-dose pharmacokinetics of a novel oral platinum cytostatic drug ([OC-6-43]-bis[acetato][1-adamantylamine]amminedichloroplatinum [IV]) in pigs.

The pharmacokinetics of total platinum (Pt) were investigated after a single oral dose of (OC-6-43-bis(acetato)(1-adamantylamine)amminedichloroplatinum (IV) (LA-12). A dose of 3 mg/kg (n = 3) and 30 mg/kg (n = 3) was given to two parallel groups of pigs (n = three each). Pt was measured in the blood, urine and feces by atomic absorption spectrometry. Blood was sampled at specified times until 240 h, urine was obtained through a catheter at 1-h intervals until 6 h, and feces were collected until 240 h after administration. LA-12 was rapidly absorbed, as indicated by a T(max) of total Pt within 0.5-1.5 h after administration. The mean (SEM) values for maximum plasma concentration (0.060 +/- 0.025 and 0.39 +/- 0.08 mg/l) and the area under the plasma concentration vs. time curve (12.6 +/- 5.6 and 36.3 +/- 2.0 mg.h/l) increased less than proportionally to the increase in the dose. The mean (SEM) Pt urinary excretion determined over a 6-h postdose period achieved only 1.9% and 0.8% of the administered doses, respectively. Within 2 h after dosing, the renal clearance of total Pt was approximately 2-fold higher than that of creatinine (CL(cr)). Thereafter, it steadily dropped and in the last collection interval (5-6 h postdose) its value was 50% less than CL(cr). Platinum recoveries in feces over 10 days after dosing reached 0.4% and 2.6% of the administered dose, respectively. This finding indicates that the extent of absorption of both doses was high. There were no changes in results of hematology and clinical biochemistry tests.

In vitro glycosidation potential towards olomoucine-type cyclin-dependent kinase inhibitors in rodent and primate microsomes.

Interspecies differences in glycosidation potential in mammalian tissues represent a factor contributing to ambiguity when endobiotic and/or xenobiotic metabolic pathways are extrapolated from animals to man. Using the TLC/autoradiographic technique, we conducted an in vitro investigation involving mouse, rat, monkey, as well as human liver and kidney microsomes to evaluate their glycoconjugation potential towards (3)H-labeled, purine-derived selective inhibitors of cyclin-dependent kinases such as olomoucine, bohemine, roscovitine, 6-(2-hydroxybenzyl)amino-2-(1-hydroxymethyl-2-methylpropyl)amino-9-isopropylpurine (compound A-4), and 6-(3-hydroxybenzyl)amino-2-[(1(R/S)-hydroxymethyl)propyl]amino-9-isopropylpurine (compound A-5) as aglycones. Principally, this study confirmed the aliphatic hydroxyl group of olomoucine-type inhibitors as a relatively suitable target for glucuronide, glucoside, xyloside, galactoside, and/or N-acetylaminoglucoside conjugation. Of the tissues examined, only the mouse microsomes were able to perform glucosidation and galactosidation reactions with the aglycones. On the other hand, monkey microsomes were superior to the mouse microsomes in a variety of glucuronide conjugates produced with compounds A-4 and A-5.

[Proteolytic enzymes from Streptomyces fradiae: a metalloendopeptidase, subtilisin-like, and trypsin-like proteinases]. / Proteoliticheskie fermenty Streptomyces fradiae: metalloéndopeptidaza, subtilizinopodobnaia i tripsinopodobnaia proteinazy.

Three proteolytic enzymes-the metalloproteinase, SFMP, and two serine proteinases, SFSP and SFTP-have been isolated and purified from the culture fluid of Streptomyces fradiae using chromatography on bacitracin-silochrome, bacitracin-Sepharose, DEAE-cellulose and fractionation by ammonium sulfate. Study of physico-chemical and functional properties of the enzymes and structural analysis revealed that SFMP is a cysteine-containing metalloendopeptidase with M(r) of 36 kDa, has a peak activity for synthetic substrates at pH 7.0-7.5 and at 60-65 degrees C and is stable at pH 7.0-9.0. The serine proteinase SFSP is related to subtilisin-like enzymes, has a M(r) of 29 kDa and a pH optimum at 7.5-8.5 at temperature up to 50 degrees C. The proteinase is stable at pH 4.0-9.0 and retains 30% of its activity at 70 degrees C. The other serine proteinase, SFTP, has a M(r) of 26 kDa and is related to trypsin-like enzymes. Its activity for synthetic substrates of trypsin is maximal at pH 6.8-8.8 at 50 degrees C. The enzyme is stable at pH 4.5-8.5 and at temperature below 50 degrees C. It has been shown that Streptomyces fradiae, like Streptomyces griseus and other Streptomycetes, possesses an ability to secrete serine proteinases (SFSP and SFTP) related to two evolutionally distinct families of serine proteinases, i.e., subtilisin and chymotrypsin families. SFMP and SFSP have been isolated and characterized for the first time.

[Primary structure of the intracellular serine proteinase from Bacillus amyloliquefaciens. III. Amino acid sequence of peptides obtained by hydrolysis with a Glu,Asp-specific proteinase. Reconstruction of the entire amino acid sequence of the proteinase]. / Pervichnaia struktura vnutrikletochnoi serinovoi proteinazy Bacillus amyloliquefaciens. III. Aminokislotnaia posledovatel'nost' peptidov, poluchennykh gidrolizom Glu,Asp-spetsifichnoi proteinazoi. Rekonstruktsiia polnoi aminokislotnoi posledovatel'nosti proteinazy.

Glu,Asp-specified protease hydrolysate of intracellular serine proteinase (ISP) was separated by ion-exchange chromatography on a sulphocationite resin followed by HPLC to yield 30 individual peptides. Their sequences, spanning to 243 amino acid residues, were determined by the manual Edman procedure. Four overlapping fragments were reconstructed by comparing their sequences with those of tryptic and chymotryptic peptides. To arrange these fragments in the proteinase polypeptide chain and to reconstruct the enzyme's total sequence, additional peptides were isolated from the tryptic hydrolysate and analysed. Primary structure of ISP, corresponding to 297 amino acid residues, was reconstructed. Its comparison with related serine proteinases revealed the following levels of homology: with Bacillus subtilis intracellular serine proteinase, 88%; with secretory subtilisin BPN' produced by B. amyloliquefaciens, 46%.

[Primary structure of an intracellular serine proteinase from Bacillus amyloliquefaciens. II. Amino acid sequence of peptides in a chymotrypsin hydrolysate]. / Pervichnaia struktura vnutrikletochnoi serinovoi proteinazy Bacillus amyloliquefaciens. II. Aminokislotnaia posledovatel'nost' peptidov khimotripticheskogo gidrolizata.

Chymotrypsin hydrolyzate of the intracellular serine protease was separated by ion-exchange chromatography on a sulphocationite resin followed by HPLC to yield fifty one individual peptides. Their sequences, corresponding in total to 381 amino acid residues, were determined by the manual Edman procedure.

[Primary structure of intracellular serine proteinase from Bacillus amyloliquefaciens. I. Isolation of the enzyme and amino acid sequence of peptides of tryptic hydrolysate]. / Pervichnaia struktura vnutrikletochnoi serinovoi proteinazy Bacillus amyloliquefaciens. I. Vydelenie fermenta i opredelenie aminokislotnoi posledovatel'nosti peptidov tripticheskogo gidrolizata.

Method of isolation of intracellular serine protease was modified. Gramicidin S-sepharose CL-4B with a higher content of the ligand, synthesized through a modified procedure, was used as an affinity sorbent which simplified the purification and led to the pure enzyme with high specific activity and 90% yield. Trypsin hydrolyzate of the protease was separated by ion-exchange chromatography on a sulphocationite resin followed by paper chromatography and paper electrophoresis to yield twenty-five individual peptides. Their complete or partial sequences, corresponding in total to 146 amino acid residues, were determined by the manual Edman procedure.

[Structure of a pepsin fragment containing the functionally important aspartic acid residue]. / Struktura fragmenta pepsina, soderzhashchego funktsional'noi vazhnyi ostatok asparaginovoi kisloty

The structure of B-5 fragment obtained under degradation of pig pepsin by cyanogen bromide is determined. B-5 fragment is a central part of pepsin molecule and it contains 46 amino acids, including a functionally important aspartic residue of the active site of the enzyme. Amino acid sequence of B-5 fragment was established by means of sequenator analysis and of studying the structure of peptides isolated from B-5 fragments.

[Application of the automatic Edman procedure to establishing the primary structure of swine pepsinogen and its fragments]. / Primenenie avtomaticheskogo metoda Edmana dlia ustanovleniia pervichnoi struktury svinogo pepsinogena, pepsina i ego fragmentov

The automatic Edman procedure was applied to elucidate N-terminal sequences of swine pepsinogen, pepsin, and the fragments of its degradation by BrCN, i. e. B-1 and B-5. A "Beckman" model 890 instrument was used in experiments. 50 amino acid residues were split off the pepsinogen molecule and identified and 55 amino acid residues-off the pepsin molecule by means of gas chromatography. A continuous N-terminal sequence of pepsinogen was 119 amino acids, in which the overlapping of the known peptide sequences with enzymic hydrolysers was taken into account. In B-1, B-4, B-5 fragments 22, 31 and 38, residues, respectively were analyzed with the sequencer.