Microencapsulation of human insulin DEAE-dextran complex and the complex in liposomes by the emulsion non-solvent addition method

The blood levels of ethanol, acetaldehyde, acetate, methanol, acetone, lactate, pyruvate, and glucose were measured in 23 male alcohol-dependent patients on days 2 to 6 after hospitalization and in 22 healthy male blood donors. Correlations between the biochemical parameters and 17 symptoms of the alcohol-withdrawal syndrome (AWS) were calculated. Abnormally high levels of ethanol, methanol, acetate, and acetone as well as hypoglycaemia were found on day 2, but lactacidaemia and pyruvataemia were pronounced throughout the observation period. AWS severity correlated positively with the acetone content on day 2 and with the acetate content on days 2 to 6. Negative correlations were found between ethanol levels and craving for alcohol, methanol levels and craving for alcohol, and between psychopathologic disorders and the total AWS severity score. The results suggest that concentrations of blood ethanol, methanol, acetate, and acetone exceeding their normal endogenous levels can be used only as indicators of recent heavy drinking. Linear discriminant analysis using the levels of the nine parameters studied enabled the correct classification of 91% to 96% of alcoholic patients during 1 week of abstinence and 100% of control subjects. The most informative parameters in the discrimination between alcoholics and controls were lactate, pyruvate, the lactate/pyruvate ratio, acetate, and acetone.     

Bioconversion of hydrocortisone to prednisolone by immobilized bacterial cells in a two-liquid-phase system

Bioconversion of hydrocortisone to prednisolone by free, immobilized and reused immobilized cells of three bacterial strains (Bacillus sphaericus ATCC 13805, Bacillus sphaericus SRP III and Arthrobacter simplex 6946) in an aqueous and a two-liquid-phase system using different organic solvents was investigated. The experiments were carried out in a 125 cm³ shake flask at 27±2°C, 220 rpm for 96 h. The contents of prednisolone and hydrocortisone in samples taken at 0, 3, 6, 24, 48, 72, 96 and 144 h were determined by HPLC analysis. The immobilized bacterial cells showed higher prednisolone yield than the free form in an aqueous system. In the two-phase systems, the butyl acetate to aqueous media ratio of 1: 30 for all three bacterial strains in immobilized forms gave the highest prednisolone yields, at an incubation time of 144 h, of 87·6, 70·6 and 88·3% respectively. For an n-decane to aqueous ratio of 1: 6, moderate prednisolone yields of 81·8, 47·9 and 71·4% were obtained with shorter incubation times of 72, 96 and 6 h respectively. For cyclohexane and other alcohols, the organisms produced low yields of prednisolone (0–30%). Single reuse of all three immobilized bacterial cells gave a 3–20% lower yield of prednisolone than the non-reused cells. The increase in hydrocortisone concentration decreased the prednisolone production whereas increasing the n-decane to aqueous ratio from 1: 6 to 1: 3 caused no significant change in the productivity. © 1998 Society of Chemical Industry     

Chemical stability enhancement and cytotoxicity reduction of papain loaded in PLGA nanospheres

Poly(lactic-co-glycolic acid) (PLGA) nanospheres loaded with papain were prepared by the emulsion solvent diffusion in water (ESD) and the w/o/w emulsion solvent evaporation (ESE) methods. The nanosphere loaded with papain from the ESE method gave smaller particle sizes (220–232?nm) and higher encapsulation efficiency of about two-folds than those from the ESD method. The morphology of the nanospheres loaded with papain prepared by the ESE method exhibited spherical shape and smooth surface investigated by SEM and TEM. The release profile of papain from the PLGA nanospheres of the ESD and ESE method indicated two phases with an initial rapid phase of 6?h and followed by the slow release phase of 48?h. The unloaded PLGA nanospheres from the two methods did not show any cytotoxicity in human skin fibroblasts, while the unloaded papain gave toxicity more than the loaded papain of 1.5 times. Papain loaded in PLGA nanospheres prepared by the ESE method was more chemical stable than the unloaded papain of eight and three times when kept at 4°C and 25°C for 6 weeks, respectively. The developed stable and low cytotoxic nanosphere loaded with papain can be further developed as topical products.     

Relationship between biological activities and bioactive compounds in the fermented rice sap

Various rice (Oryza sativa L. var. indica), including white plain, purple plain, brown plain, white glutinous and purple glutinous rice, were fermented with Look Pang (a mixed culture of yeasts and molds). The sap samples were assayed for the bioactive compounds (unsaturated fatty acids, total phenolic compounds and total anthocyanin) and the biological activities (antioxidative, tyrosinase inhibition, cell proliferation and MMP-2 inhibition activities). The fermented purple plain sap samples at day 6th of the fermentation period showed high antioxidative, the highest tyrosinase inhibition and MMP-2 inhibition activities with low cytotoxicity to normal human skin fibroblast by SRB assay in comparing to other rice sap samples. This study has indicated the strong positive relationship between the bioactive compounds and the biological activities of the purple rice sap which can be further developed as functional foods and cosmetics.     

Antioxidant activities and skin hydration effects of rice bran bioactive compounds entrapped in niosomes

Bioactive compounds [ferulic acid (F), gamma-oryzanol (O) and phytic acid (P)] in rice bran have been widely used as antioxidants in skin care products. However, one of the major problems of antioxidants is the deterioration of their activities during long exposure to air and light. Niosomes have been used to entrap many degradable active agents not only for stability improvement, but also for increasing skin hydration. The objective of this study was to determine antioxidant activities [by in vitro ORAC (oxygen radical absorbance capacity) and ex vivo lipid peroxidation inhibition assay] and in vivo human skin hydration effects of gel and cream containing the rice bran extracts entrapped in niosomes. Gel and cream containing the rice bran extracts entrapped in niosomes showed higher antioxidant activity (ORAC value) at 20-28 micromol of Trolox equivalents (TE) per gram of the sample than the placebo gel and cream which gave 16-18 micromolTE/g. Human sebum treated with these formulations showed more lipid peroxidation inhibition activity than with no treatment of about 1.5 times. The three different independent techniques including corneometer, vapometer and confocal Raman microspectroscopy (CRM) indicated the same trend in human skin hydration enhancement of the gel or cream formulations containing the rice bran extracts entrapped in niosomes of about 20, 3 and 30%, respectively. This study has demonstrated the antioxidant activities and skin hydration enhancement of the rice bran bioactive compounds when entrapped in niosomes and incorporated in cream formulations.     

Characterization of amphotericin B liposome formulations

Liposomes composed of hydrogenated soya phosphatidylcholine (Emulmetik 950®)/cholesterol/charged lipids [dicetyl phosphate (-) or stearylamine (+)] were developed. The hydrogenated soya phosphatidylcholine/cholesterol/charged lipid liposomes at molar ratios of 1:1:0, 7:2:0, 7:2:1 (-), and 7:2:1 (+), with and without the entrapped amphotericin B (0.05 mg AmB/mg lipid), were prepared by a chloroform-film method with sonication. The charges of liposomes were characterized by a Zeta-Meter. The negative liposomes with and without the entrapped AmB showed higher surface charge density than other formulations. The size distribution of liposomes determined by standard error of the mean (SEM) was in the range of 0.115 to 0.364 µm. The smallest size was observed in the negative liposomes with the entrapped drug [7:2:1 (-) AmB]. The lamellarity of more than 15 layers was observed by transmission electron microscope (TEM) in the neutral liposomes with the entrapped drug [7:2 AmB]. The transition temperature and enthalpy of transition (ΔH) were determined by differential scanning calorimetry (DSC). Positive liposomes with the entrapped and unentrapped AmB demonstrated higher ΔH of the first peak than other formulations, indicating higher rigidity of liposomal membrane. The AmB contents in liposomes were determined by high-performance liquid chromatography (HPLC) with ultraviolet (UV) detection at 382 nm. The percentages of entrapment of AmB in all formulations were above 85%. The positive liposome [7:2:1 (+) AmB] formulation, which gave the highest thermal stability, was selected for further skin absorption evaluation.     

Effects of Gastrointestinal Administration of Human Insulin and a Human Insulin-Deae-Dextran Complex Entrapped in Different Compound Liposomes on Blood Glucose in Rats

The effects of gastrointestinal administration (oral, in duodenum and colon) of human insulin and a human insulin-DEAE (diethylaminoethyl) dextran complex entrapped in different compound liposomes in comparing to human insulin alone given subcutaneously on blood glucose level of normal and STZ-diabetic rats were investigated. The liposomes were prepared from a hydrogenated soy lecithin (Epikuron, E 200 H) and by a high pressure homogenization procedure. Samples were lyophilized and reconstituted in 0.067 M phosphate buffer, pH 7.4 before application. The complexed insulin (0.25 and 0.5 IU/kg insulin) showed no diffrences in blood glucose lowering profiles from the free insulin when both were administered intravenously in normal rats. When given orally, the complex (30 and 60 IU/kg Insulin) entrapped in positive liposomes (E 200 H/cholesterol/stearylamine = 7:2:1, in a molar ratio) indicated no effects in STZ rats. However, this complex liposome (6.0 IU/kg insulin) gave a retention effect of blood glucose lowering as % initial level of about 12% after 5 hours when injected in duodenum and showed two maximum effects of 21 and 22% at 100 and 300 minutes respectively when administered in colon of normal rats. When the free insulin entrapped in positive liposomes was given in duodenum In normal rats, the maximum effect of blood glucose lowering of 10% was observed at 2 hours (6 IU/kg insulin) and 1 hour (12 IU/kg insulin). For the free insulin (12 IU/kg) entrapped in other liposome systems given in duodenum of normal rats, both negative (E 200 H/cholesterol/dlcetyl phosphate=7:2:1, in a molar ratio) and neutral (E 200 H/cholesterol=1:1, in a molar ratio) liposomes indicated the maximum effect of about 30% at 120 minutes. Both cholesterol rich positive (E 200 H/cholesterol/stearylamlne=7:7:1, in a molar ratio) and negative (E 200 H/cholesterol/dicetyl phosphate=7:7:1, in a molar ratio) liposomes showed not only a maximum effect of about 20% at 2 hours, but also a retention of glucose lowering of 20% after 7 hours as well. This study suggested that a development of human Insulin by complexlng with the DEAE-dextran polymer and/or entrapping in liposomes, as a drug delivery system in duodenum and colon, is possible.     

Comparison of prednisolone production from cortexolone by free and immobilized isolated and standard collection strains using mixed culture techniques

The bioconversion of cortexolone (Reichstein's Compound S) to hydrocortisone or hydrocortisone to prednisolone by reference and isolated fungi and bacteria were investigated. The reference strains Cunninghamella echinulata ATCC 8688a and Bacillus sphaericus ATCC 13805, and the isolates C echinulata SRP III and B sphaericus SRP III were used as free or immobilized forms in single or mixed cultures by either sequential or concomitant mixing. In single cultures, immobilized fungi C echinulata ATCC 8688a and C echinulata SRP III showed less hydrocortisone yield than the free form, whereas immobilized B sphaericus ATCC 13805 and B sphaericus SRP III gave higher prednisolone yield than the free form. In mixed cultures, free fungi with immobilized bacteria of both isolated and standard strains showed the highest prednisolone yield both by concomitant and sequential mixing. The highest prednisolone yield of 72.5±0.78% with a bioconversion rate of 0.76% h ⁻¹ by sequential mixing was obtained with free C echinulata ATCC 8688a with immobilized B sphaericus ATCC 13805 at 96 h, whereas the free C echinulata SRP III with immobilized B sphaericus ATCC 13805 and the free C echinulata SRP III with immobilized B sphaericus SRP III gave 63.4±0.74% and 47.7±0.44% at 120 h with bioconversion rates of 0.53 and 0.40% h⁻¹ respectively. Free C echinulata SRP III with immobilized B sphaericus SRP III gave a maximum yield of 52.8±0.82% with a bioconversion rate of 0.44% h ⁻¹ at 120 h which was comparable to 59.2±0.44% with a bioconversion rate of 0.49% h ⁻¹ obtained by free C echinulata ATCC 8688a with immobilized B sphaericus ATCC 13805 using concomitant mixing. This study indicated high biotransformation activity of free fungal and immobilized bacterial isolates which showed comparable steroidal productivity to the standard strains both in single and mixed cultures by the concomitant mixing technique. © 1999 Society of Chemical Industry