Accelerated wound healing and anti-inflammatory effects of physically cross linked polyvinyl alcohol–chitosan hydrogel containing honey bee venom in diabetic rats

Diabetes is one of the leading causes of impaired wound healing. The objective of this study was to develop a bee venom-loaded wound dressing with an enhanced healing and anti-inflammatory effects to be examined in diabetic rats. Different preparations of polyvinyl alcohol (PVA), chitosan (Chit) hydrogel matrix-based wound dressing containing bee venom (BV) were developed using freeze–thawing method. The mechanical properties such as gel fraction, swelling ratio, tensile strength, percentage of elongation and surface pH were determined. The pharmacological activities including wound healing and anti-inflammatory effects in addition to primary skin irritation and microbial penetration tests were evaluated. Moreover, hydroxyproline, glutathione and IL-6 levels were measured in the wound tissues of diabetic rats. The bee venom-loaded wound dressing composed of 10 % PVA, 0.6 % Chit and 4 % BV was more swellable, flexible and elastic than other formulations. Pharmacologically, the bee venom-loaded wound dressing that has the same pervious composition showed accelerated healing of wounds made in diabetic rats compared to the control. Moreover, this bee venom-loaded wound dressing exhibited anti-inflammatory effect that is comparable to that of diclofenac gel, the standard anti-inflammatory drug. Simultaneously, wound tissues covered with this preparation displayed higher hydroxyproline and glutathione levels and lower IL-6 levels compared to control. Thus, the bee venom-loaded hydrogel composed of 10 % PVA, 0.6 % Chit and 4 % BV is a promising wound dressing with excellent forming and enhanced wound healing as well as anti-inflammatory activities.     

Quality by design: understanding the formulation variables and optimization of metformin hydrochloride 750?mg sustained release tablet by Box–Behnken design

This study examined the formulation variables and optimization of a sustained-release metformin hydrochloride 750?mg tablet using a combination of hydroxypropyl methylcellulose (HPMC), polyvinylpyrrolidone (PVP K30), and glyceryl behenate with a computer optimization technique based on a response surface methodology (RSM) utilizing a polynomial equation. A three-factor, three-level Box?CBehnken design was used for the optimization procedure with the contents of HPMC (X ₁), PVP K30 (X ₂), and glyceryl behenate (X ₃) as independent variables. The drug release percentages at 1, 3, and 10?h were the target responses and were restricted to 25?C35?% (Y ₁), 55?C65?% (Y ₂), and 85?C100?% (Y ₃), respectively. The quadratic model fit the data well and the resulting equation was used to predict the responses in the optimal region. The optimized formulation was achieved with 165?mg HPMC, 40?mg PVP K30, and 22.5?mg glyceryl behenate, and the observed responses of the optimized formulation were very close to the values predicted using the RSM optimization technique. Furthermore, the drug release rate and dissolution curve shape of the optimized formulation were similar to those of the commercial product.     

Effect of γ-aminobutyric acid and nattokinase-enriched fermented beans on the blood pressure of spontaneously hypertensive and normotensive Wistar–Kyoto rats

In this study we have evaluated the changes in arterial blood pressure in spontaneously hypertensive rats (SHR) caused by the short-term intake of Bacillus subtilis B060-fermented beans with significant γ-aminobutyric acid (GABA) and nattokinase activity. After being weaned, 7-week-old male SHR and 7-week-old male Wistar–Kyoto (WKY) rats were randomized into seven groups. Until the 8^th week of life, the rats in each group were given one of the following: Group 1, high dose of GABA and nattokinase in the SHR (SHD); Group 2, medium dose of GABA and nattokinase in the SHR (SMD); Group 3, low dose of GABA and nattokinase in the SHR (SLD); Group 4, negative control in the SHR (SD); Group 5, positive control in the SHR (SM); Group 6, high dose of GABA and nattokinase in the WKY (WHD); and Group 7, negative control in the WKY (WD). Distilled water served as the negative control, and captopril (50 mg/kg), a known ACE inhibitor, served as the positive control. Systolic blood pressure and diastolic blood pressure values were measured weekly from the 8^th week to the 16^th week of life using the tail-cuff method. A definite decrease in systolic and diastolic blood pressure values could be observed in the rats treated with captopril and in the rats that received GABA and nattokinase. The greatest antihypertensive effect was observed when the pharmacological treatment was administered. The effect of the daily intake of fermented beans containing GABA and nattokinase may be helpful in controlling blood pressure levels in hypertensive model animals. The fermentation of beans with B. subtilis B060 may therefore constitute a successful strategy for producing a functional food with antihypertensive activity.     

Delayed adverse effects of neonatal exposure to polymeric nanoparticle poly(ethylene glycol)-block-polylactide methyl ether on hypothalamic–pituitary–ovarian axis development and function in Wistar rats

We studied delayed effects of neonatal exposure to polymeric nanoparticle poly(ethylene glycol)-block-polylactide methyl ether (PEG-b-PLA) on the endpoints related to pubertal development and reproductive function in female Wistar rats from postnatal day 4 (PND4) to PND 176. Female pups were injected intraperitoneally, daily, from PND4 to PND7 with PEG-b-PLA (20 or 40 mg/kg b.w.). Both doses of PEG-b-PLA accelerated the onset of vaginal opening compared with the control group. In the low-dose PEG-b-PLA-treated group, a significantly reduced number of regular estrous cycles, increased pituitary weight due to hyperemia, vascular dilatation and congestion, altered course of hypothalamic gonadotropin-releasing hormone-stimulated luteinizing hormone secretion, and increased progesterone serum levels were observed. The obtained data indicate that neonatal exposure to PEG-b-PLA might affect the development and function of hypothalamic–pituitary–ovarian axis (HPO), and thereby alter functions of the reproductive system in adult female rats. Our study indicates a possible neuroendocrine disrupting effect of PEG-b-PLA nanoparticles.     

Solid-State Stability of Human Insulin I. Mechanism and the Effect of Water on the Kinetics of Degradation in Lyophiles from pH 2–5 Solutions

Purpose. Previous studies have established that in aqueous solution at low pH human insulin decomposition proceeds through a cyclic anhydride intermediate leading to the formation of both deamidated and covalent dimer products. This study examines the mechanism and kinetics of insulin degradation in the amorphous solid state (lyophilized powders) as a function of water content over a similar pH range. Methods. Solutions of 1.0 mg/mL insulin were adjusted to pH 2–5 using HC1, freeze-dried, then exposed to various relative humidities at 35°C. The water content within the powders was determined by Karl Fischer titration, and the concentrations of insulin and its degradation products were determined by HPLC. Degradation kinetics were determined by both the initial rates of product formation and insulin disappearance. Results. Semi-logarithmic plots of insulin remaining in lyophilized powders versus time were non-linear, asymptotically approaching non-zero apparent plateau values, mathematically describable by a reversible, first-order kinetic model. The rate of degradation of insulin in the solid state was observed to increase with decreasing apparent pH (pH) yielding, at any given water content, solid-state pH-rate profiles parallel to the solution pH-rate profile. This pH dependence could be accounted for in terms of the fraction of the insulin A21 carboxyl in its neutral form, with an apparent pKa of 4, independent of water content. Aniline trapping studies established that the mechanism of degradation of human insulin in lyophilized powders between pH 3–5 and at 35°C involves rate-limiting intramolecular nucleophilic attack of the Asn_A21 C-terminal carboxylic acid onto the side-chain amide carbonyl to form a reactive cyclic anhydride intermediate, which further reacts with either water or an N-terminal primary amino group (e.g., Phe_B1, and Gly_Al) of another insulin molecule to generate either deamidated insulin (Asp_A21) or an amide-linked covalent dimer (e.g., [Asp_A21-Phe_B1] or [Asp_A21-Gly_A1]), respectively. The rate of insulin degradation in lyophilized powders at 35°C increases with water content at levels of hydration well below the suspected glass transition and approaches the rate in solution at or near the water content (20–50%) required to induce a glass transition. Conclusions. The decomposition of human insulin in lyophilized powders between pH 3–5 is a water induced solid-state reaction accelerated by the plasticization effect of sorbed water. The formation of the cyclic anhydride intermediate at A21 occurs readily even in the glassy state, presumably due to the conformational flexibility of the A21 segment even under conditions in which the insulin molecules as a whole are largely immobile.