Preparation, characterisation, pharmacokinetics and distribution of esculin microspheres administered via intravitreal injection into rabbit brain.

This study explored the distribution of esculin microspheres in rabbit brain tissue following intravitreal injection and investigated the possibility of direct entry of the drug into the brain through the eye, to develop a formulation with enhanced therapeutic efficacy against Parkinson's disease.Chitosan microspheres of esculin were prepared via an emulsification cross-linking method and their characteristics were evaluated, including angle of repose, bulk density, and swelling ratio. Furthermore, the pharmacokinetic parameters and brain tissue distribution in rabbits were compared among groups administered esculin eye drops, intravitreal esculin solution, and intravitreal esculin microspheres, to determine whether esculin could enter the brain through an ocular route.The results showed that the prepared esculin microspheres were spherical and had good fluidity. Notably, intravitreal administration enhanced the area under the curve (AUC) of esculin in the thalamus. Delivery through microspheres prolonged the drug retention time in both rabbit plasma and brain tissues, as well as the brain-targeting efficiency of esculin.The collective findings indicated that there may be a direct eye-brain pathway facilitating enter of esculin microspheres into brain tissue after intravitreal injection, supporting the utility of intravitreal esculin microspheres as an effective therapeutic formulation for Parkinson's disease, a long-term chronic condition.

Esculin prevents Lipopolysaccharide/D-Galactosamine-induced acute liver injury in mice.

Liver injury is an important cause of serious liver disease and is characterized by inflammatory and oxidative responses. Esculin, a coumarinic derivative found in Aesculus hippocastanum L., has been shown to exhibit anti-inflammatory and anti-oxidative effects. Here, we investigated the effects and molecular mechanism of esculin on Lipopolysaccharide/D-Galactosamine (LPS/D-Gal)-induced acute liver injury. A mouse model for acute liver injury was induced by intraperitoneal injection with D-Gal and LPS, and was assessed by histology, and serum transaminase analyses. The results showed that esculin significantly reduced the pathological symptoms of acute liver injury, as well as serum AST and ALT levels. LPS/D-Gal-induced liver myeloperoxidase (MPO) activity and malondialdehyde (MDA) content were also suppressed by esculin. Furthermore, LPS/D-Gal-induced liver tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß) production were attenuated by esculin. Our data demonstrate that esculin can inhibit nuclear factor kappa B (NF-κB) activation as well as increase nuclear factor E2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) expression. In conclusion, this paper demonstrates that esculin protects liver injury induced by LPS/D-Gal via inhibiting inflammatory and oxidative responses.

Preparation of Chitosan Okra Nanoparticles: Optimization and Evaluation as Mucoadhesive Drug Delivery System.

OBJECTIVE: To prepare chitosan-okra gum nanoparticles and their evaluation as mucoadhesive drug delivery system for intranasal delivery of esculin. METHOD: Esculin loaded chitosan-okra gum based nanoparticles were prepared using ionic gelation method. The preparation method was optimized using Box-Behnken experimental design employing okra gum concentration, chitosan concentration, pH and stirring speed as independent variables and particle size, encapsulation efficiency and zeta potential of the formulation were selected as dependent variables. The optimized formulation was characterized using FTIR, SEM and TEM. The nanoparticles were evaluated for their bioadhesive strength and in vitro drug release studies. The optimized intranasal formulation was administered to rats and the pharmacokinetic profile and biodistribution studies were carried out to calculate the targeting efficiency of the formulation in the brain. RESULTS: The nanoparticles were found to depict particle size in the range of 294.0 to 613.4 nm. The concentration of gums was found to significantly influence the particle size and encapsulation efficiency. The nanoparticles depicted bioadhesive strength of 32±2%. The in vitro drug release studies showed 96.4±4.2% release of esculin from nanoparticles in 4h. The drug targeting of NPs to brain was found to be nearly double that of esculin given as simple solution. CONCLUSION: The nanoparticles prepared using chitosan-okra gum were found to possess good mucoadhesive strength with and high brain targeting efficiency.

Colloidal characteristics and functionality of rationally designed esculin-loaded hydrogel microcapsules.

This work reports the development of different types of alginate hydrogel microparticles designed specifically for the esculin (ESC) payload. Negatively charged alginate (ALG) microspheres were prepared by the ionotropic gelation technique, and an oppositely charged polyelectrolyte (PE) shell as a compatible polycation (chitosan (CHIT) or gelatin (GEL)) or a synthetic PEs (poly(allylamine hydrochloride) (PAH) and poly(4-styrenesulfonate) (PSS)) were adsorbed using electrostatic complexation. Thorough characterization of microparticles was performed with advanced microscopic techniques (scanning electron, fluorescence and confocal), followed by stability studies, ESC encapsulation efficacy determination and in vitro release kinetics measurements. We provide an in-depth investigation of the relationships between the properties (thickness, viscosity, areal mass, zeta potential) of the outer shell and the retaining and release abilities of the fabricated microcarriers, using quartz crystal microbalance with dissipation monitoring technique (QCM-D), spectroscopic ellipsometry and streaming potential measurements, combined in a new approach that was not attempted before for micrometric particles. The PAH-PSS and GEL coatings provided sufficient protection against ESC release under simulated gastric conditions that followed a two-stage Corrigan-Gallagher model with a marginal release rate in the first (lag) stage. This seems to be an interesting outcome, since it is rather peculiar for a low-molecular weight hydrophilic compound encapsulated in a highly porous microhydrogel to be released in such a manner.

Aesculin modulates bone metabolism by suppressing receptor activator of NF-κB ligand (RANKL)-induced osteoclastogenesis and transduction signals.

Aesculin (AES), a coumarin compound derived from Aesculus hippocasanum L, is reported to exert protective role against inflammatory diseases, gastric disease and cancer. However, direct effect of AES in bone metabolism is deficient. In this study, we examined the effects of AES on osteoclast (OC) differentiation in receptor activator of NF-κB ligand (RANKL)-induced RAW264.7 cells. AES inhibits the OC differentiation in both dose- and time-dependent manner within non-toxic concentrations, as analyzed by Tartrate Resistant Acid Phosphatase (TRAP) staining. The actin ring formation manifesting OC function is also decreased by AES. Moreover, expressions of osteoclastogenesis related genes Trap, Atp6v0d2, Cathepsin K and Mmp-9 are decreased upon AES treatment. Mechanistically, AES attenuates the activation of MAPKs and NF-κB activity upon RANKL induction, thus leading to the reduction of Nfatc1 mRNA expression. Moreover, AES inhibits Rank expression, and RANK overexpression markedly decreases AES's effect on OC differentiation and NF-κB activity. Consistently, AES protects against bone mass loss in the ovariectomized and dexamethasone treated rat osteoporosis model. Taken together, our data demonstrate that AES can modulate bone metabolism by suppressing osteoclastogenesis and related transduction signals. AES therefore could be a promising agent for the treatment of osteoporosis.

Gastroprotective effect of esculin on ethanol-induced gastric lesion in mice.

The gastroprotective effect of esculin was investigated in a mouse model of ethanol-induced gastric lesion. Administration of esculin at doses of 5, 10, and 20 mg/kg body weight prior to ethanol ingestion led to significant gastroprotection compared with untreated mice. Gastric mucosal lesions were evaluated by macroscopic and histopathological alterations, lesion index, and myeloperoxidase (MPO) activity. Pretreatment with esculin significantly reduced macroscopic and histopathological damage, gastric lesion index, and MPO activity in a dose-dependent manner. Moreover, esculin significantly reduced nitric oxide (NO) production, inducible NO synthase (iNOS) levels, and nuclear factor-kappa B (NF-κB) p65 protein expression in gastric tissues after ethanol challenge. Analysis of inflammatory cytokines indicated that esculin pretreatment markedly suppressed the increased expression of tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) in ethanol-treated mice. The results demonstrate a protective effect of esculin against gastric injury and suggest that the underlying mechanism might be associated with inhibition of NF-κB activation, which subsequently reduces expression of iNOS, TNF-α, and IL-6.

Esculin improves dyslipidemia, inflammation and renal damage in streptozotocin-induced diabetic rats.

BACKGROUND: Increasing studies have shown that dyslipidemia and inflammatory responses play important roles in the progression of microvascular diabetic complications. Esculin (ES), a coumarin derivative, was extracted from Fraxinus rhynchophylla. The present study was to evaluate the potential effects of ES on lipid metabolism, inflammation responses and renal damage in streptozotocin (STZ)-induced experimental diabetic rats and explore the possible mechanism. METHODS: Diabetic rat model was established by administration high-glucose-fat diet and intraperitoneal injection of STZ 45 mg/kg. ES was administrated to diabetic rats intragastrically at 10, 30 and 90 mg/kg for 10 weeks respectively. The levels of triglycerides (TG), total cholesterol (T-CHO), low density lipoproteins (LDL), and high-density-cholesterol (HDL-C) in serum were measured. IL-1, IL-6, ICAM-1, NO, NAGL, and AGEs level in serum were detected by ELISA assay. The accumulation of AGEs in kidney tissue was examined by immunohistochemistry assay. RESULTS: The results showed that ES could decrease TG, T-CHO, LDL levels in serum of diabetic rats in a dose dependent manner. ES also decreased IL-1, IL-6, ICAM-1, NO and NGAL levels in serum of diabetic rats in a dose dependent manner. Furthermore, ES at 30 and 90 mg/kg significantly decreased AGEs level in serum and alleviated AGEs accumulation in renal in diabetic rats. CONCLUSIONS: Our findings indicate that ES could improve dyslipidemia, inflammation responses, renal damage in STZ-induced diabetic rats and the possible mechanism might be associated with the inhibition of AGEs formation.

Protective effect of esculin on streptozotocin-induced diabetic renal damage in mice.

The present study investigated the presence and mechanism of esculin-mediated renoprotection to assess its therapeutic potential. Esculin was orally administered at 20 mg/kg/day for 2 weeks to streptozotocin-induced diabetic mice, and its effects were compared with those of the vehicle in normal and diabetic mice. After oral administration of esculin to mice, the concentrations of esculin and esculetin in blood were 159.5 ± 29.8 and 9.7 ± 4.9 ng/mL at 30 min, respectively. Food and water intake were significantly increased in the diabetic mice compared to normal mice but attenuated in mice receiving esculin. The elevated blood glucose level and hepatic glucose-6-phosphatase expression were significantly reduced in esculin-treated diabetic mice, supporting the antidiabetic effect of esculin. Esculin also increased the uptake of glucose and induced the insulin-evoked phosphorylation of insulin receptor, Akt, and glycogen synthase kinase 3ß in C2C12 myotubes, indicating a potential for improvement of insulin sensitivity. In addition, esculin lessened the elevated blood creatinine levels in diabetic mice and ameliorated diabetes-induced renal dysfunction by reducing caspase-3 activation in the kidney. Data support the beneficial effect of esculin against diabetes and oxidative stress-related inflammatory processes in the kidney.

Protective effect of esculin against prooxidant aflatoxin B1-induced nephrotoxicity in mice.

The study was designed to investigate the protective effect of esculin against pro-oxidant aflatoxin B1 (AFB1)-induced nephrotoxicity in mice. In this study toxicity was developed by oral administration of AFB1 at a dose of 66.60 µg/kg bw/day for 90 days in male Swiss albino mice. Esculin (150 mg/kg bw/0.2 ml/day) and standard compound ascorbic acid (300 mg/kg bw/0.2 ml/day) was given after 30 min of AFB1 administration for 90 days. Protective efficacy was assessed by measuring the levels of lipid peroxidation (LPO) and non-enzymatic antioxidants such as reduced glutathione (GSH) and also by measuring activities of enzymatic antioxidants such as glutathione peroxidase (GPX), glutathione-S-transferase (GST), glutathione reductase (GR), superoxide dismutase (SOD) and catalase (CAT) in kidney. Results were analysed at the 30(th), 60(th) and 90(th) day of the daily treatments, which showed a decrease in the level of LPO and an increase in the levels of enzymatic and non-enzymatic antioxidants. The protective effect of esculin was further proved by histopathological findings as it exhibited regenerative activities in mice renal tubules against AFB1-induced nephrotoxicity. The results obtained clearly demonstrate that the protective efficacy of esculin against pro-oxidant AFB1-induced nephrotoxicity in mice might be due to its antioxidants and free radical scavenging properties.

The effect of deacetylated gellan gum on aesculin distribution in the posterior segment of the eye after topical administration.

The aim of the present work was to evaluate the effect of deacetylated gellan gum on delivering hydrophilic drug to the posterior segment of the eye. An aesculin-containing in situ gel based on deacetylated gellan gum (AG) was prepared and characterized. In vitro corneal permeation across isolated rabbit cornea of aesculin between AG and aesculin solution (AS) was compared. The results showed that deacetylated gellan gum promotes corneal penetration of aesculin. Pharmacokinetics and ocular tissue distribution of aesculin after topical administration in rabbit eye showed that AG greatly improved aesculin accumulation in posterior segmentsrelative to AS, which was probably attributed to conjunctivital/sclera pathway. The area-under-the-curve (AUC) for AG in aqueous humor, choroid-retina, sclera and iris-ciliary body were significantly larger than those of AS. AG can be used as a potential carrier for broading the application of aesculin.

Determination of aesculin in rat plasma by high performance liquid chromatography method and its application to pharmacokinetics studies.

A sensitive and reproducible high performance liquid chromatography method with UV detection was described for the determination of aesculin in rat plasma. After deproteinization by methanol using metronidazole as internal standard (I.S.), solutes were evaporated to dryness at 40 degrees C under a gentle stream of nitrogen. The residue was reconstituted in 100 microl of mobile phase and a volume of 20 microl was injected into the HPLC for analysis. Solutes were separated on a Diamonsil C18 column (250 mm x 4.6 mm i.d., 5 microm particle size, Dikma) protected by a ODS guard column (10 mm x 4.0 mm i.d., 5 microm particle size), using acetonitrile-0.1% triethylamine solution (adjusted to pH 3.0 using phosphoric acid) (10:90, v/v) as mobile phase (flow-rate 1.0 ml/min), and wavelength of the UV detector was set at 338 nm. No interference from any endogenous substances was observed during the elution of aesculin and internal standard (I.S., metronidazole). The retention times for I.S and aesculin were 10.4 and 12.4 min, respectively. The limit of quantification was evaluated to be 57.4 ng/ml and the limit of detection was 24.0 ng/ml. The method was used in the study of pharmacokinetics of aesculin after intraperitoneal injection (i.p.) administration in rats.

Randomized, double-blind trial comparing topical nitroglycerine with xylocaine and Proctosedyl in idiopathic chronic anal fissure.

OBJECTIVE: To compare symptomatic relief, healing, and changes in maximal anal resting pressure with the use of topical formulations in patients with chronic anal fissure. METHODS: Sixty-four consecutive patients with chronic anal fissure were randomized into 4 groups that received, in a double-blind manner, a topical ointment that contained 0.2% nitroglycerine (GTN), 5% xylocaine, Proctosedyl (hydrocortisone acetate, heparin, framycetin sulfate, esculoside, ethoform, butoform) or petroleum jelly (Vaseline), to be applied twice daily. Patients were reviewed at 2-week intervals for 6 weeks. Anal manometry was done before, and 20 minutes after, the first application of the ointment. RESULTS: There was significant (p < 0.0001) reduction in mean anal resting pressure after application of GTN, but not any other ointment. Of 16 patients receiving GTN, complete pain relief occurred in 6 and 15 patients after 2 and 4 weeks of treatment, respectively; this was more frequent than in the other 3 groups. At 6 weeks also, complete pain relief occurred more often with GTN than with Vaseline or xylocaine. After 4 weeks of treatment, 3 patients on GTN had complete healing of fissure as compared to one each in the xylocaine and Proctosedyl groups and none in the Vaseline group. At 6 weeks, healing of fissure had occurred in 15 of 16 patients receiving GTN as compared to 4 receiving Vaseline, 11 receiving xylocaine, and 12 on Proctosedyl. CONCLUSIONS: Topical nitroglycerine produces 'chemical sphincterotomy' with reduction in mean anal resting pressure. Pain relief and healing of fissure occurred earlier with GTN than with other treatments. GTN should be considered as the treatment of choice for the non-surgical management of patients with chronic anal fissure.

Effect of a total extract from Fraxinus ornus stem bark and esculin on zymosan- and carrageenan-induced paw oedema in mice.

This study investigates the total ethanol extract (TE) of the stem bark of Fraxinus ornus and its constituent esculin (EN). They inhibited classical pathway (CP) and alternative pathway (AP) of complement activation in mouse serum. After intraperitoneal administration the total extract displayed antiinflammatory activity in both zymosan- and carrageenan-induced paw oedema in mice. The results suggest that the traditional use of Fraxinus ornus stem bark extracts in the treatment of inflammatory disorders is at least partially due to its coumarin constituents.

Comparative efficacy and tolerability of two ointment and suppository preparations ('Uniroid' and 'Proctosedyl') in the treatment of second degree haemorrhoids in general practice.

A multi-centre general practice, open study was carried out in 89 patients with second degree haemorrhoids to compare the efficacy and tolerability of two antibiotic-corticosteroid combinations ('Uniroid' and 'Proctosedyl') in ointment and suppository formulations. Patients were allocated at random into 4 groups and received treatment with one of the trial preparations for 1, 2 or 3 weeks, as required, with weekly assessments of response. There were no significant differences between the various groups at the start of treatment. Significant improvement occurred in all groups during treatment. Both suppository and ointment formulations were broadly comparable and control of symptoms was achieved from Week 2 onwards, building up to levels in excess of 90% after 3 weeks of therapy. With regard to the symptoms of pain and itching, suppositories gave marginally greater relief in the early stages of treatment, while both ointment and suppositories were associated with similar reduction in bleeding from haemorrhoids. Whereas both suppository formulations were about equal in reducing anal discharge, 'Uniroid' ointment was clinically superior to 'Proctosedyl' ointment in controlling this symptom over the 3-week trial period. No unwanted effects were experienced attributable to treatment. No statistically significant differences between the two ointment and the two suppository formulations were identified in this study and all four preparations were found to be efficacious in the majority of patients studied.

[Critical study of the therapeutic effects of phlebotropic drugs on chronic venous insufficiency]. / Etude critique des effets thérapeutiques des médicaments phlébotropes sur l'insuffisance veineuse chronique.

The authors describe two types of test. Their aim was to verify the therapeutic potency of drugs used in the treatment of chronic venous disorders. The tests were carried out using both a single prescription and using successive prescriptions. The results showed that with strict clinical surveillance, phlebotropic drugs had definite therapeutic effects both on the subjective disturbances and on certain objective parameters.

Treatment of experimental brain oedema following sudden decompression, surgical wound, and cold lesion with vasoprotective drugs and the proteinase inhibitor "Trasylol".

The study was performed on 81 cats with three models of experimental brain oedema: sudden decompression, surgical wound, and cold injury. During the experiments blood pressure, central venous pressure, and intracranial pressure were recorded. The blood-brain-barrier was tested with Evans blue solution. The gray and white matter tissue was sampled at the end of the experiment, and the water content and sodium and potassium concentrations were determined. The animals with the same experimental model were divided into three groups: untreated, treated with the vasoprotective agents, and treated with the protease inhibitor Trasylol. In the sudden decompression model after balloon deflation, white matter haemorrhages and oedema development were found in gray matter and basal nuclei. In animals treated with the vasoprotective drugs, haemorrhages were not observed, and oedematous changes were less pronounced. The Trasylol effect on oedema development was not significant in this model. In the surgical wound model, oedematous changes were observed after 24 hours following the lesion. Oedema occurred in the white matter, as in the animals with cold lesions. In both models--surgical wound and cold lesion--the beneficial effect of Trasylol was shown, while the effect of Aescorin was less evident. The results obtained seemed to testify to the usefulness of both Trasylol and vasoprotective drugs in the prevention and treatment of brain oedema in neurosurgical patients.