Isolation and structure determination of a novel complex of the teicoplanin family.

A new teicoplanin-like antibiotic was discovered when using Actinoplanes teichomyceticus strain 3/W, the fermentation medium containing Alburex, and the fermentation time 275 hours. The new product was separated from teicoplanin complex by polyamide resin chromatography and purified by Amberlite XAD-7 and affinity resin chromatographies. The structure was established on the basis of the physico-chemical characteristics of the complex and of its aglycone. The new structure is that of teicoplanin with a carbonyl group substituting for the CHNH2 group of amino acid 1. We hypothesize that the novel complex is a transformation product of teicoplanin due to a simple transamination reaction, as supported by its structure and by the concomitant decrease in teicoplanin concentration during its production.

Antibiotics A21459 A and B, new inhibitors of bacterial protein synthesis. II. Structure elucidation.

The structures of the antibiotics, active against a few Gram-negative bacteria and Clostridium difficile, were determined on the basis of physicochemical analyses on the intact molecules and on the acid hydrolysate of A21459 A. FAB-MS and 1H and 13C NMR investigations identified the amino acid units and determined their sequence. Antibiotics A21459 A and B are homodetic cyclic peptides constituted by eight amino acid units. They are glycine, methoxytryptophan, tryptophan, cysteine, alanine, sarcosine, dehydroalanine, and alpha-aminobutyric acid for A21459 A (alanine for A21459 B). Cysteine and alanine condensed to form a thiazole moiety, according to the biosynthesis of thiazole containing antibiotics.

Antibiotic GE37468 A: a novel inhibitor of bacterial protein synthesis. II. Structure elucidation.

GE37468 A is a novel antibiotic produced by Streptomyces sp. ATCC 55365. It has molecular mass 1309.48 and formula C59H52O12N14S5 and belongs to the thiazolyl peptide group of antibiotics. The structure was elucidated by 1H and 13C NMR and MS studies on intact molecule and its hydrolysis products. The antibiotic is a highly modified peptide containing a macrocycle and a side chain composed of a thiazole ring and two dehydroalanine units.

HPLC method for the quantitation of cispentacin enantiomers in rat urine.

The two enantiomers of cispentacin, an antifungal antibiotic, are determined by reversed phase HPLC, after derivatization with Marfey's reagent, in 24 h urine samples collected from rats treated sc and iv with 20 mg/kg cispentacin racemate obtained by synthesis. The application range of the method is 25-250 mg/L for each enantiomer with a precision of 4.0-9.0%. Comparison with an authentic sample of natural origin (-)-cispentacin indicated that (-) enantiomer is excreted unchanged in smaller percentage than (+) enantiomer, which is almost completely eliminated as such.

Isolation, structure determination and biological activity of A-16686 factors A' 1, A' 2 and A' 3 glycolipodepsipeptide antibiotics.

When Actinoplanes strain ATCC 33076, the producer of A-16686 A1, A2 and A3 complex, is fermented in a suitable medium three additional factors, designated A' 1, A' 2 and A' 3 are produced. These were isolated and characterized, and were shown to differ from the parent components of the original complex by lacking one mannose unit. Bioconversion of A factors into A' factors was achieved by incubation with the mycelium of Actinoplanes ATCC 33076. Factor A' 2 has better antibacterial activity than A2 against some bacteria.

Quantitative determination of zetidoline, a new antipsychotic agent, in human blood plasma and saliva using capillary gas chromatograph-mass spectrometry.

A specific and sensitive assay is described. Zetidoline is extracted with ethyl ether from 1 ml of plasma or saliva added with the internal standard. The extract is carefully purified and injected into a gas chromatography-mass spectrometry system equipped with a crosslinked capillary column and operated in single ion monitoring mode by electron impact. Quantitative response is linear in the range 2-50 ng/ml. The detection limit is 1 ng/ml.

Teicoplanin metabolism in humans.

Teicoplanin, a lipoglycopeptide antibiotic, consists of five major components (A2-1 through A2-5), one hydrolysis component (A3-1), and four minor components (RS-1 through RS-4). All the major components contain an N-acyl-beta-D-glucosamine, but they differ in the lengths and branchings of their acyl-aliphatic chains. Previous studies with radiolabeled teicoplanin in rats and humans have shown that the drug is eliminated by the renal route and that metabolic transformation is very minor, about 5%. A possible metabolic transformation of teicoplanin into A3-1 was also suggested. In the present study in humans, two metabolites (metabolites 1 and 2; 2 to 3% of total teicoplanin) were isolated after intravenous administration of radiolabeled teicoplanin. After purification, their structures were determined by fast atom bombardment mass spectroscopy and 1H nuclear magnetic resonance spectroscopy on the basis of the well-known correlations established in this field, and they were found to be new teicoplaninlike molecules, bearing 8-hydroxydecanoic and 9-hydroxydecanoic acyl moieties. This metabolic transformation is likely due to hydroxylation in the omega-2 and omega-1 positions for metabolites 1 and 2, respectively, of the C-10 linear side chain of component A2-3. This might explain the low extent of metabolism of teicoplanin if we consider that only component A2-3 has a linear chain that is susceptible to such oxidation.

Determination of the acyl moieties of the antibiotic complex A40926 and their relation with the membrane lipids of the producer strain.

Structures of the fatty acid residues characterizing the various components of A40926 were determined by gas chromatography/mass spectrometry on the methyl esters obtained by methanolysis of the complex. The results confirm the residues previously assigned to Factor A (n-undecanoic acid) and B (10-methyl-undecanoic acid) and establish the residues of Factor A1 (9-methyl-decanoic acid), B1 (n-dodecanoic acid), RS1 (8-methyl-nonanoic acid), RS2 (n-decanoic acid), and RS3 (n-tridecanoic acid). As the Actinomadura species contain in their mycelia large quantities of C15-C17 fatty acid residues as membrane phospholipids, these mycelia were saponified and the fatty acids obtained were analyzed as above. There is a close correlation between the fatty acid content of A40926 complex and that of the longer homologues in the producer mycelia.

Microbial de-mannosylation and mannosylation of teicoplanin derivatives.

The single components of the teicoplanin complex, glycopeptide antibiotics active against Gram-positive bacteria, can be converted in the corresponding de-mannosyl derivatives by cultures of Nocardia orientalis NRRL 2450 or Streptomyces candidus NRRL 3218. Conversely, teicoplanin aglycone and other teicoplanin de-mannosyl derivatives can be converted in the corresponding teicoplanin mannosyl derivatives by cultures of Actinoplanes teichomyceticus ATCC 31121. The biological transformation yields are approximately 40% for de-mannosylation and 90% for mannosylation. The processes allow for the preparation of gram quantities of the de-mannosyl derivatives of teicoplanin and of teicoplanin mannosyl derivatives. De-mannosyl teicoplanin and teicoplanin mannosyl-pseudoaglycone were not amenable to preparation by either acidic or basic chemical hydrolysis.

Factors affecting the normal and branched-chain acyl moieties of teicoplanin components produced by Actinoplanes teichomyceticus.

Teicoplanin, a glycopeptide antibiotic produced by Actinoplanes teichomyceticus, comprises five main components, denoted T-A2-1 to T-A2-5, differing in the structure of their acyl side chain, which is linear in T-A2-1 and T-A2-3 and branched in the other components. Production of T-A2-1, characterized by a linear C10:1 acyl moiety, is entirely dependent on the presence of linoleate in the fermentation medium. Addition to the medium of oleic acid esters at 2 g l-1 increases the yields of T-A2-3, characterized by a linear C10:0 acyl chain, about threefold. The antibiotic linear side chains thus appear to originate from C18 unsaturated acid by beta-oxidation degradation. The percentage of T-A2-2, T-A2-4 and T-A2-5, bearing the iso-C10:0, anteiso-C11:0 and iso-C11:0 acyl moieties, respectively, is strongly influenced by the presence in the medium of the amino acids known to be precursors of branched-chain fatty acids. Thus, valine increases the production of T-A2-2 whereas isoleucine or leucine increase the relative yields of T-A2-4 or T-A2-5, respectively. Analysis of the total cell lipids upon addition of the same amino acid shows corresponding increases in the proportion of the iso-C16:0, iso-C15:0 or anteiso-C17:0. A mutant A. teichomyceticus strain, which produces a novel teicoplanin with a linear C9:0 chain, differs from the wild strain in the presence of the linear C17:1 acid in its lipids.(ABSTRACT TRUNCATED AT 250 WORDS)

Teicoplanin metabolism in rats.

This study was done to see whether teicoplanin undergoes metabolic transformation in rats. Sprague-Dawley rats were given 7.6 mg of [14C]teicoplanin (904 U/mg, 7.6 muCi/mg) per kg intravenously; 73.2 +/- 4.0% of the administered radioactivity and 59.1 +/- 4.8% of the administered microbiological activity were recovered in the 24-h urine samples. The difference between these two values was due to adsorption of teicoplanin to the urinary sediment, making some of the antibiotic not available for microbiological determination. In fact, after a sample or urine was filtered through an Acrodisc (0.45 micron pore size), radiochemical and microbiological data for the filtrate were very similar. Possible metabolites were looked for by high-performance liquid chromatographic analysis of the teicoplanin complex composition in the urine samples, with both UV and radioactivity detection. The quantitation, based on the 14C percentage in each peak, showed that no more than 3 to 5% of the [14C]teicoplanin underwent metabolic and/or chemical transformation.

Liquid chromatographic-mass spectrometric studies on rifamycin antibiotics.

The use of a direct liquid introduction type liquid chromatographic-mass spectrometric interface to study highly thermally labile rifamycin antibiotics is described. Using negative ionization, abundant molecular ions were observed, and the spectra, also contained structurally significant fragments. Variation of the high-performance liquid chromatographic parameters did not change the spectra, thus making it easy to change chromatographic conditions. In quantitative studies, a surprising correlation was found, indicating that the mass spectrometric signal was proportional to the square of the sample concentration.

Laser-induced vaporization mass spectrometry of rifamycins.

The mass spectra of many rifamycins cannot be obtained by electron ionization (EI) owing to their thermal decomposition. When a laser beam is used to vaporize the sample through an optic fibre inserted in a hollow probe which reaches the sample cup, decomposition is minimized and the EI spectra show abundant molecular ions and fragments of structurally high diagnostic value. These ionic species are easily observed owing to the lack of chemical noise often present in soft ionization methods, such as direct liquid chemical ionization and fast atom bombardment.

A42867, a novel glycopeptide antibiotic.

A42867 is a new glycopeptide antibiotic of the ristocetin-vancomycin class active against aerobic and anaerobic Gram-positive bacteria. A42867 is produced by a strain of Nocardia nov. sp. ATCC 53492. A42867 was isolated during a screening program aimed at the discovery of new members of this glycopeptide class of antibiotics, by affinity chromatography based on an acyl-D-alanyl-D-alanine probe. The structure of A42867 was elucidated by fast atom bombardment MS, high field 2D 1H NMR spectroscopy, and HPLC analysis of the hydrolyzed carbohydrates. A42867 differs from vancomycin in the sugar portion and in the presence of only one chlorine atom in the peptide core. Its biological activity on Gram-positive aerobic and anaerobic bacteria is similar to that of other antibiotics of this group.

Isolation and structure determination of two new analogs of teicoplanin, a glycopeptide antibiotic.

Teicoplanin is an antibiotic produced by fermentation of Actinoplanes teichomyceticus as a complex formed by five closely related glycopeptides characterized by different fatty acid chains of ten and eleven carbon atoms. In addition, minor quantities of related substances are present. Two of them, named RS-1 and RS-2, were shown to be teicoplanins having as fatty acid chains 10-methylundecanoic acid and n-dodecanoic acid, respectively. Other two related substances, named RS-3 and RS-4, have now been isolated and purified starting from fermentation broths of a mutant of the same microorganism producing them in substantial amounts. This was achieved by semipreparative reversed-phase liquid chromatography carried out on high-pressure scale. The structures were assigned on the basis of 1H NMR spectra and homonuclear COSY 2D experiments and fast atom bombardment MS spectrometry, in comparison with the large mass of data till now accumulated on teicoplanin. RS-3 and RS-4 are teicoplanins having as fatty acid chains 6-methyloctanoic acid and n-nonanoic acid, respectively.

Liquid chromatography-mass spectrometry coupling and its application in pharmaceutical research.

The advantages and technical problems related to the on-line connection of liquid chromatography and mass spectrometry are described, with special emphasis on applications in pharmaceutical chemistry. The most important characteristics of various interfaces, such as direct liquid introduction, Magic, continuous flow fast atom bombardment and thermospray, are discussed. Special applications to the study of thermally very labile compounds are considered.

Isolation and structure determination of the main related substances of teicoplanin, a glycopeptide antibiotic.

Teicoplanin is a complex formed by five closely related glycopeptides and by a small amount of a hydrolysis product. Minor quantities of related substances are also present. Two of them (named RS-1 and RS-2) were isolated and purified starting from the tailing fractions of a teicoplanin batch. Preparative reversed-phase liquid chromatography on large low-pressure and medium high-pressure scales, concentration, desalting, and freeze-drying steps were applied. 300 mg of RS-1 and 900 mg of RS-2 were obtained in a purity grade (about 90%) sufficient for structural investigation. Starting from considerations on the HPLC retentivity and on biosynthesis, the structures were assigned on the basis of 1H-N.M.R. spectra and homonuclear CO-SY 2D experiments, FAB-MS spectrometry, and GC-MS of the esters of the fatty acids obtained by hydrolysis. RS-1 and RS-2 are teicoplanins having 10-methyl-undecanoic acid and n-dodecanoic acid, respectively as fatty acid chains. No major difference in the in vitro activity of these teicoplanins emerged in comparison with teicoplanin complex.