Interaction of the polyene antibiotic nystatin with bivalent copper ions

Characteristic features of interaction of nystatin with bivalent copper salts in water, methanol and dimethylsulfoxide were studied. It was shown that stable compounds of copper and nystatin at ratios of 1 : 2 and 1 : 4 formed in the above solvents. The antibiotic in these compounds was in an inert, native or activated state. Physicochemical and biological properties of the compounds were investigated.     

Sterol composition of nystatin-resistantCandida maltosa mutants

Composition of sterol fractions of nystatin-resistantCandida maltosa strains was determined. Using UV-spectrometry, TLC and GLC-MS it was demonstrated that resistance to nystatin is connected with the composition alterations of yeast cell sterols. Block of different stages of ergosterol biosynthesis was revealed in some mutants,viz. C-24-transmethylation, Δ⁸→Δ⁷, 14α-demethylation, C-5(6)-dehydrogenation, reduction of C-14(15) and C-24(28) double bonds.     

Interaction of amphotericin B and nystatin with phospholipid bilayer membranes:effect of cholesterol.

Sodium-22 efflux was measured in multilamellar liposomes, exposed to one of the two polyene antibiotics amphotericin B or nystatin. Polyene mediated 22Na transport progressively rises with membrane sterol concentrations up to about 20 mol %, but falls with higher cholesterol concentrations. The polyene induced 22Na movement in cholesterol rich liposomes could be 'restored' by the addition of either dibucaine or propranolol (two local anesthetics) to the aqueous solution. These observations are interpreted in terms of the model of De Kruijff and Demel (Biochim. Biophys. Acta, 339, 57-70, 1974). In this model, nystatin and amphotericin B first complex with cholesterol and then these complexes aggregate to form transmembrane channels. It is here proposed that the aggregation of these complexes is inhibited by a high cholesterol content (decreased membrane fluidity) but that the two local anesthetics, by disrupting phospholipid-sterol interactions (increased membrane fluidity), can 'restore' this process of aggregation.     

Cooperative partition model of nystatin interaction with phospholipid vesicles

Nystatin is a membrane-active polyene antibiotic that is thought to kill fungal cells by forming ion-permeable channels. In this report we have investigated nystatin interaction with phosphatidylcholine liposomes of different sizes (large and small unilamellar vesicles) by time-resolved fluorescence measurements. Our data show that the fluorescence emission decay kinetics of the antibiotic interacting with gel-phase 1,2-dipalmitoyl-sn-glycero-3-phosphocholine vesicles is controlled by the mean number of membrane-bound antibiotic molecules per liposome, . The transition from a monomeric to an oligomeric state of the antibiotic, which is associated with a sharp increase in nystatin mean fluorescence lifetime from approximately 7-10 to 35 ns, begins to occur at a critical concentration of 10 nystatin molecules per lipid vesicle. To gain further information about the transverse location (degree of penetration) of the membrane-bound antibiotic molecules, the spin-labeled fatty acids (5- and 16-doxyl stearic acids) were used in depth-dependent fluorescence quenching experiments. The results obtained show that monomeric nystatin is anchored at the phospholipid/water interface and suggest that nystatin oligomerization is accompanied by its insertion into the membrane. Globally, the experimental data was quantitatively described by a cooperative partition model which assumes that monomeric nystatin molecules partition into the lipid bilayer surface and reversibly assemble into aggregates of 6 +/- 2 antibiotic molecules.     

热带假丝酵母产生子囊孢子的条件及单倍体分离的研究

本文研究了诱导热带假丝酵母子囊孢子产生和子囊破壁的条件,以及单倍体分离和鉴别方法.结果表明:NaAc、KCl这两种成分即可满足热带假丝酵母的产孢营养要求,在含有NaAc 8.2 g/L、KCl 1.8 g/L的琼脂培养基中,28℃培养7 d,热带假丝酵母细胞的产孢率可达到47.5%;单纯使用蜗牛酶对破除热带假丝酵母子囊壁的效率甚低,酶解液加入适量的石英砂同时振荡处理4 h左右,可以显著提高对热带假丝酵母子囊破壁速率.用这种方法处理,绝大多数子囊壁均可破除.子囊破壁处理后再以52℃处理10 min杀死残余的营养体细胞,分离物的单倍体孢子比例可由灭活处理前的48%,提高到76%以上.     

Transglycosyl-Amylase of Candida tropicalis

Transglucosyl-amylase was purified 96-fold and partially characterized. The K_m value with dextrin as substrate was 9.1 mg/ml. Glycerol, an acceptor of d-glucose, appeared to inhibit dextrin hydrolysis noncompetitively. The energy of activation of the enzyme was 7,920 cal/mole. Indirect determinations showed that synthesis of d-glucosyl glycerol was significantly affected by the nature of the amylaceous substrate. Glucosyl-glycerol synthesis did not increase as incubation temperature was raised from 50 to 60 C. Direct determinations by gas-liquid chromatography indicated that the synthesis of glucosyl glycerol, as a function of the concentration of either enzyme, substrate, or glycerol, traced a curvilinear path approaching 15 mg/ml as the maximum. When enzyme, substrate, and glycerol at high concentrations were varied in all possible combinations, however, conditions for producing as much as 47.5 mg/ml of glucosyl glycerol were established.     

Trnasglucosyl-amylase of Candida tropicalis

Transglucosyl-amylase was purified 96-fold and partially characterized. The K(m) value with dextrin as substrate was 9.1 mg/ml. Glycerol, an acceptor of d-glucose, appeared to inhibit dextrin hydrolysis noncompetitively. The energy of activation of the enzyme was 7,920 cal/mole. Indirect determinations showed that synthesis of d-glucosyl glycerol was significantly affected by the nature of the amylaceous substrate. Glucosyl-glycerol synthesis did not increase as incubation temperature was raised from 50 to 60 C. Direct determinations by gas-liquid chromatography indicated that the synthesis of glucosyl glycerol, as a function of the concentration of either enzyme, substrate, or glycerol, traced a curvilinear path approaching 15 mg/ml as the maximum. When enzyme, substrate, and glycerol at high concentrations were varied in all possible combinations, however, conditions for producing as much as 47.5 mg/ml of glucosyl glycerol were established.     

Stabilization of standard samples of levorin and nystatin with antioxidants]

To prolong the storage period of the reference samples of levorin and nystatin, polyenic macrolide antibiotics, their effect of 23 antioxidants was studied by using rapid inactivation. The antioxidants belonged to compounds of different classes. The inactivation was performed at 20, 37 and 50 degrees C in the presence of 1, 2 or 5 per cent of the antioxidants. An antioxidant of the class of partially hydrated oxyquinolines was shown to have the highest stabilizing action on levorin and nystatin. It may be recommended for stabilizing the levorin and nystatin reference samples.     

Double-layered mucoadhesive tablets containing nystatin

The objective of this work was to design a mucoadhesive tablet with a potential use in the treatment of oral candidosis. A 2-layered tablet containing nystain was formulated. Lactose CD (direct compression), carbomer (CB), and hydroxypropylmethylcellulose (HPMC) were used as excipients. Tablets were obtained through direct compression. Properties such as in vitro mucoadhesion, water uptake, front movements, and drug release were evaluated. The immediate release layer was made of lactose CD (100 mg) and nystatin (30 mg). The CB:HPMC 9∶1 mixture showed the best mucoadhesion properties and was selected as excipient for the mucoadhesive polymeric layer (200 mg). The incorporation of nystatin (33.3 mg) in this layer affected the water uptake, which, in turn, modified the erosion front behavior. Nystatin showed a first-order release. The polymeric layer presented an anomalous kinetic (n=0.82) when this layer layer was individually evaluated. The mucoadhesive tablet formulated in this work releases nystatin quickly from the lactose layer and then in a sustained way, during approximately 6 hours. from the polymeric layer. The mixture CB:HPMC 9∶1 showed good in vitro mucoadhesion. A swelling-diffusion process modulates the release of nystatin from this layer. A non-Fickian (anomalous) kinetic was observed.     

Distribution of pore sizes in black lipid membranes treated with nystatin

Resistance changes were measured on artificial membranes made of oxidized cholesterol and treated with nystatin. The experimental data were smoothed and then fitted to a mixture of Gaussian functions using the maximum likelihood method; the best fit was to a mixture of four gaussians centered approximately at 4, 6, 9 and 12 molecules per pore, leading to the conclusion that these are the most likely pore sizes.     

Phagocyte-mediated killing of Candida tropicalis

Human peripheral monocytes (MO), neutrophils (PMN), and lymphocytes (PBL) were tested for their ability to kill Candida tropicalis. With incubation times between 30 min and 2 h, unstimulated MO and PMN, but not PBL, were efficient killers of C. tropicalis. Both leukocyte subsets were able to kill at minimum 2.5 1 effector to target ratios. Pre-incubation of MO for 24 h with interferon-gamma or tumor necrosis factor (TNF) increased their ability to kill yeast targets. TNF alone had no effect on C. tropicalis targets at concentrations up to 1000 U/ml. PBL activated for 4 d with interleukin-2 did not kill yeast targets. PMN exhibited more cytocidal efficiency per cell than MO in these assays. Direct contact of effectors and targets was required; no significant killing by PMN or MO supernatants was measured. PMN-mediated killing, but not MO killing, was inhibited by a mixture of catalase and Superoxide dismutase suggesting that oxygen-dependent killing mechanisms were partially responsible for candidacidal activity.     

Characteristics of trehalase inCandida tropicalis

Summary The trehalase content of different yeasts varies widely. A strain ofCandida tropicalis was found to be the best source of this enzyme among the yeasts tested. The trehalase activity in this yeast could be increased 8.5 times by growing it on trehalose rather than glucose. Thus trehalase is an adaptive enzyme inC. tropicalis. It was found that the amount of trehalase which could be solubilized increased with increasing pH during autolysis of the cells, none being released from the cell debris at pH 4.5 and most at pH 6.3. Some evidence was obtained to show that the solubilization was caused by an enzyme. The stability of trehalase under various conditions was studied. A partial purification was achieved by precipitation with 40% ethanol at a temperature of −18°C. The maximum temperature of the enzyme was 48°C., and the optimum pH ranged from 4.1 to 5.3     

n-Alkane oxidation byCandida tropicalis

Summary The specific activity of the enzyme catalase was investigated in batch cultures ofCandida tropicalis on the following substrates: Gelsenberg 14/18, n-hexandecane, 1-octadecene, oleyl alcohol, oleic acid and glucose. The catalase activity does not change with the different oxidation levels of the hydrocarbon substrates. A correlation between specific activity and growth rate was established.Inhibition experiments with 2-amino-1,3,4-triazol (AT), a specific catalase inhibitor, gave no evidence for two kinds of hydrogen peroxide-degrading enzymes in yeast cells.Growth on hydrocarbons instead of on glucose enhanced the specific activities of both isocitratelyase (ICL) and catalase to about the same extent.On the other hand the succinate-cytochrome C-oxidoreductase, a mitochondrial enzyme, showed more or less the same activity on both substrates.All these facts suggest that the catalase enzyme is related to the degradation (- or -oxidation) or to the gluconeogenesis (glyoxylate cycle) of fatty acids, but not to the initial oxidation of the alkanes.     

The Blomia tropicalis allergens

Blomia tropicalis allergens are the most important mite allergens in tropical regions. Most of them only have 30-40% sequence identity with their Dermatophagoides counterparts and they share low IgE cross reactivity and exhibit different immunobiology. Unlike the pyroglyphid counterparts, Blo t 5 is the major allergen whereas Blo t 1 only has modest allergenicity.