Biotransformations of rifamycins: process possibilities

Rifampicin, an important antibiotic, is manufactured by chemical conversion of rifamycin S which is obtained by the chemical modification of rifamycin B. Rifamycin B is a product of Nocardia mediterranei fermentations. The chemical conversion of rifamycin B to rifamycin S has many disadvantages: Strong acidic conditions are required, heavy foam formation accompanies transformation and the yields are low. This review highlights the developments in alternative, biochemical transformations using enzymes and cells; the main focus is on transformations carried out by rifamycin oxidase.     

Pharmacokinetics and biotransformation of rifamycins in the body of experimental animals]

The study on distribution of 14C-rifampicin and 14C-rifamycin S in experimental animals after intramuscular administration of the drugs showed that concentrations of rifampicin in the organs and blood were higher than those of rifamycin S. Biotransformation products of both antibiotics, such as 25-deacetylrifampicin, N-oxide of rifampicin, 3-phormylrifamycin SV, rifamycin SV and others were found in the liver, kidneys, bile and urine. No products of the antibiotic metabolism were found in the blood, lungs and spleen.     

Allosteric modulation of the RNA polymerase catalytic reaction is an essential component of transcription control by rifamycins

Rifamycins, the clinically important antibiotics, target bacterial RNA polymerase (RNAP). A proposed mechanism in which rifamycins sterically block the extension of nascent RNA beyond three nucleotides does not alone explain why certain RNAP mutations confer resistance to some but not other rifamycins. Here we show that unlike rifampicin and rifapentin, and contradictory to the steric model, rifabutin inhibits formation of the first and second phosphodiester bonds. We report 2.5 A resolution structures of rifabutin and rifapentin complexed with the Thermus thermophilus RNAP holoenzyme. The structures reveal functionally important distinct interactions of antibiotics with the initiation sigma factor. Strikingly, both complexes lack the catalytic Mg2+ ion observed in the apo-holoenzyme, whereas an increase in Mg2+ concentration confers resistance to rifamycins. We propose that a rifamycin-induced signal is transmitted over approximately 19 A to the RNAP active site to slow down catalysis. Based on structural predictions, we designed enzyme substitutions that apparently interrupt this allosteric signal.     

Chromatographic separation of rifamycins on a thin sorbent thin layer]

A method of simultaneous separation of rifampicins O, S and rifamicin derivates S, SV in a thin layer of silica gel KSK No. 2 impregnated with acetate buffer, pH 4.6 in a solvent system chlorophorm-methanol (100:5) was developed. The minimum detectable amount of the compounds studied was 1--2gamma, when the coloured zones were estimated visually.     

Actions of growth factors in the follicle

In this paper we have examined the possibility that soluble factors produced by the thecal and granulosa cells may be essential local modulators of follicular development. The observations that insulin could influence both the growth and the differentiation of granulosa cells were important in establishing the concept that peptides could act as amplifiers of the actions of gonadotrophins. Insulin alone did not influence aromatase activity significantly but acted synergistically with FSH to augment aromatase activity in rat granulosa cells. Unlike aromatase activity, insulin alone was able to significantly stimulate 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) activity, the maximum level achieved approaching that obtained with high concentrations of FSH. To determine if insulin could influence other parameters of granulosa cell function in addition to steroidogenesis, we measured a component of extracellular matrix, fibronectin, previously shown to be inhibited by FSH. Treatment with insulin independently inhibited the increase in fibronectin secretion observed in control cultures. Also, insulin alone was able to stimulate quiescent bovine granulosa cells to incorporate [3H]thymidine into DNA under serum-conditions. The concentration of insulin required in these experiments was higher than physiological levels suggesting that other insulin-like growth factors may be involved. Our work and that of others has shown that IGF1 can mimic the actions of insulin and is effective at lower concentrations. A possible source of IGF1 production in the follicle was sought initially by collecting rat granulosa cell conditioned medium, and assessing biological activity and immunoreactivity. Conditioned medium augmented the actions of FSH on aromatase activity and alone stimulated 3 beta-HSD, indicating the presence of insulin-like bioactivity. A positive reaction on immunoblots using specific antiserum confirmed the presence of immunoreactive IGF1. Conditioned medium from thecal cells contained a growth-promoting activity (TcGF) that did not augment FSH-induced aromatase activity. The production of growth factors locally within the follicle may represent the self-amplifying mechanism that enables the dominant follicle to complete its developmental program and ovulate.     

Production of oxytetracycline and rifamycins B and SV with cells immobilized on glass wool

The antibiotic producing microorganisms Streptomyces rimosus PFIZER 18234-2 and Amycolatopsis mediterranei CBS 42575 were immobilized in glass wool for the production of oxytetracycline and rifamycins B and SV. The growth of the immobilized cells was dependent on the type of the microorganism and on the amount of glass wool as well. After 5 exchanges of the medium, 530 mg O.T.C., 2138 mg rifamycin B and 8672 mg rifamycin SV were maintained. The periodic batch culture was continued for 20 days.     

Electrophysiological Actions of Oxytocin on the Rabbit Myometrium

The electrical activities of myometrial cells of the pregnant rabbit uterus have been studied by means of sucrose-gap and intracellular micro-electrode recording techniques. The resting potential of the myometrial cell was about -50 mv, and it is unaffected by the duration of pregnancy or placental attachment. Action potentials of the myometrium, although dependent on external Na⁺, were not always of the regenerative type; preparations from nonparturient uteri often produce mainly small spikes. The mean spike amplitude was 35 mv, rising at a mean maximum rate of 3 v/sec. Oxytocin, in concentrations less than 500 µU/ml, increased the mean spike amplitude to 48 mv and the mean maximum rate of rise to 7 v/sec, without affecting the resting potential. The relation between membrane potential and dV/dt of the spike was steepened by oxytocin, suggesting that oxytocin increased the number of normally sparse sodium gates in the myometrial membrane. By this action, oxytocin is believed to increase the probability of successful regenerative spikes and thereby initiate electrical activity in quiescent preparations, increase the frequency of burst discharges, the number of spikes in each burst, and the amplitude of spikes in individual cells.     

Trends in the Actions of Mammalian Masticatory Muscles

The actions of the masticatory muscles of a variety of mammalsin which feeding behavior and the configuration of the masticatoryapparatus differ have been reported. The most common approachused in these studies involves (1) obtaining a good anatomicalperception of the musculature, (2) deriving a theoretical modelof the actions of these muscles during jaw movement, and (3)testing this model by recording muscle activity and jaw movementssimultaneously. A catalogue of the activity patterns in eleven species of mammalsduring food reduction reveals certain trends in the actionsof the masticatory muscles. Horizontal jaw movements are generatedprimarily by differential activities of the deep temporalis,superficial masseter, and medial pterygoid. Vertical movementsand the maintenance of tooth to food contact apparently areproduced by action of the superficial temporalis, deep masseter,and zygomaticomandibularis. Thus, horizontal movements are seeminglygenerated by muscles having fibers arranged in marked anteroposteriordirection, whereas vertical movements are generated by muscleshaving more or less vertically arranged fibers. The asymmetry of jaw movement and the muscular activity generatingit suggest that mastication involves an interactionbetween anunbalanced and flexible functional unit (muscles) and a balancedand stable structural unit (skull and teeth). Thus, any unbalancingof the structural unit results in a further unbalancing of themasticatory process.