Imad I. Hamdan

In vitro and in vivo acute antihyperglycemic effects of five selected indigenous plants from Jordan used in traditional medicine.

ETHNOPHARMACOLOGICAL RELEVANCE Achillea santolina L., Pistacia atlantica Desf, Rheum ribes L., Sarcopoterium spinosum (L.) Spach and Teucrium polium L. have traditionally been used as herbal antidiabetic medicines. However their alleged benefits and mechanisms remain elusive. AIM OF THE STUDY This study aimed to evaluate the effect of water extracts of these plants in in vitro and in vivo experiments. MATERIALS AND METHODS In vitro enzymatic starch digestion with aqueous extracts from plants at concentrations of 1, 5, 10, 12.5, 25, 50 and 100 mg/ml was assayed using α-amylase and α-amyloglucosidase. Acarbose was used as control and glucose liberation was determined by glucose oxidase method. Oral starch tolerance test (OSTT) and oral glucose tolerance test (OGTT) were determined for the plant extracts at concentrations 125, 250 and 500 mg/kg b.wt. on Sprague-Dawley rats. Blood glucose levels in rats treated with plant extracts and drugs (acarbose or metformin and glipizide) were measured at -30, 0, 45, 90 and 135 min. RESULTS Compared to acarbose (IC(50)=1.2 μg/ml), water extracts of Pistacia atlantica, Rheum ribes and Sarcopoterium spinosum exerted significant dose dependent dual inhibition of α-amylase and α-glucosidase in in vitro enzymatic starch digestion bioassay, with IC(50)s; 46.98, 58.9 and 49.9 mg/ml, respectively. Comparable in vivo results were obtained for starch-fed rats, exhibiting significant acute postprandial antihyperglycemic efficacies. While Achillea santolina and Teucrium polium extracts lacked any favourable in vitro anti-α-amylase and anti-α-glucosidase effect, other modes of action can possibly explain their substantial acute antihyperglycemic activities in starch-treated rats. Except for Pistacia atlantica extracts, none of the investigated extracts qualified for improving the glucose intolerance in fasted rats on glucose loading. CONCLUSIONS Pistacia atlantica, Rheum ribes and Sarcopoterium spinosum can be considered as potential candidates for amelioration/management of type 2 diabetes.

Spectroscopic and HPLC methods for the determination of alendronate in tablets and urine

Two methods (spectrophotometric and HPLC) have been developed and validated for the analysis of alendronate sodium in tablet dosage form. Both methods depend on the ability of alendronate sodium to react with o-phthalaldehyde (OPA) at basic pH to produce a light-absorbing derivative. The derivative was found to possess absorption maximum at 330nm where neither the derivatizing agent nor the analyte had any absorption. Thus, spectroscopic method was based on the derivatization-induced absorption of alendronate sodium at 333nm. The HPLC method was based on separation of the formed derivative from other ingredients in tablets with detection at 333nm. Both methods were satisfactory with regard to accuracy, prescion and linearity. Moreover, a HPLC method with fluorescence detection (HPLC-FD) was developed for the quantification of alendronate sodium in urine. The method was also based on the derivatization of alendronate with OPA, but fluorescence detection was employed. Linearity, recovery, selectivity, prescision and sensitivity were satisfactory for the proposed HPLC-FD method. Yet a new quantification limit (0.6ngml(-1)) for alendronate in urine was achieved.

Development and validation of a stability indicating capillary electrophoresis method for the determination of metformin hydrochloride in tablets

A simple and a stability indicating capillary electrophoresis method was developed and validated for the analysis of metformin hydrochloride in tablet formulation. The method employed 40 mM citrate buffer at pH 6.7 as a background electrolyte with an applied voltage of 15 kV (positive polarity). The capillary used was of 68.5 cm length (60 cm to detector) and the detection wavelength was 230 nm. The method was validated in accordance with the ICH requirements, which involved accuracy, precision, linearity, selectivity and both limit of detection and limit of quantitation. The current method was linear over the concentration range of 0.2-2.0mg/ml of metformin hydrochloride. The method was precise as reflected by inter-day and intra-day relative standard deviation (RSD) of less than 2%. The limit of detection and limit of quantitation were 2.0 microg/ml and 8.0 microg/ml, respectively. The stability indicating capability of the method was established by enforced degradation studies combined with peak purity assessment using photodiode array detection.

Evaluation of the acute antihyperglycemic effects of four selected indigenous plants from Jordan used in traditional medicine.

Context: Eryngium creticum Lam. (Umbelliferae), Geranium graveolens L.Her.exn Ait (Geraniaceae), Paronychia argentea Lam. (Caryophyllaceae), and Varthemia iphionoides Boiss (Compositae) have traditionally been used as antidiabetic phytomedicines. However, their alleged benefits and mechanisms remain elusive. Objectives: To evaluate the effect of these plants on in vitro and in vivo enzymatic starch digestion. Materials and methods: In vitro enzymatic starch digestion with acarbose or (1–50 or 100 mg/ml) plants aqueous extracts was assayed using α-amylase and α-amyloglucosidase. Oral starch tolerance tests and oral glucose tolerance tests were determined for the plant extracts at concentrations 125, 250, and 500 mg/kg body weight. Blood glucose levels in rats treated with plant extracts or drugs (acarbose or metformin and glipizide) were measured at −30, 0, 45, 90, and 135 min. Results and discussion: In vitro, acarbose, and water extracts of G. graveolens and V. iphionoides exerted significant dose-dependent dual inhibition of α-amylase and α-glucosidase, with respective IC50s of 1.2 μg/ml, 84.7, and 65.2 mg/ml. Comparable in vivo acute postprandial antihyperglycemic efficacies were obtained for G. graveolens and V. iphionoides in starch-fed rats. E. creticum exhibited substantial acute antihyperglycemic activities in starch-treated rats, despite lacking any favorable in vitro effectiveness. However, P. argentea lacked any inhibitory efficacy. None of the plant extracts qualified for improving the glucose tolerance in fasted rats on glucose loading. Conclusion: G. graveolens and V. iphionoides can be considered as potential candidates for therapeutic modulation of impaired fasting glycemia, impaired glucose tolerance, and type 2 diabetes.

Screening of Jordanian Flora for α-Amylase Inhibitory Activity

Thirty-five plant species that belong to different families were tested for α -amylase inhibitory activity. Four of the screened plants exhibited significant (more than 80%) α -amylase inhibitory activity. IC50 values for these plants were estimated based on the dried crude extract and found to be 0.08, 0.13, 0.32, and 0.47 mg/ml, for Osyris alba L. (Santalaceae), Sarcopotarium spinosum L. (Rosaceae), Hypericum triquetrifolium Turra (Hypericaceae) and Arbutus andrachne L. (Ericaceae), respectively. Amylase inhibitory activity was also evaluated in vivo using female rats. Results showed that the tested extracts had a favorable effect on blood glucose level after ingestion of a high dose of sucrose. The most potent extract in controlling the surge of glucose after sucrose ingestion appeared to be O. alba which also exhibited the highest α -amylase inhibitory activity in vitro.

Development and Validation of a HPLC Method for Determination of Potential Residual Cortisone Compounds in Timolol Maleate Eye Drops

Abstract One way of ensuring effective cleaning in the pharmaceutical industry is to analyze the batch of a product for a potential carry over contamination from a previously manufactured one. The potential cross contamination of timolol maleate (TM) eye drops with cortisone compounds, deserves special consideration, due to the well known deleterious effect of cortisones on glaucoma. Therefore, it was the objective of this study to develop and validate a HPLC method for the detection and quantitative determination of cortisones in TM eye drop preparations. The chromatographic behaviors of the five cortisones that were most likely to be present as contaminants in TM eye drops were characterized on different HPLC stationary phases (normal silica, C18, C8, and CN). Mobile phase and buffer constituents were further optimized on the best stationary phase material (C18). The final recommended mobile phase consisted of a mixture of THF:methanol:0.01M phosphate buffet at ratios of 15:25:60 and containing 0.01 M camphore sulfonic acid (pH adjusted to 4.2). The method was validated in light of ICH guidelines and applied to commercial samples with satisfactory results.

Capillary electrophoresis as a screening tool for alpha amylase inhibitors in plant extracts.

Capillary electrophoresis (CE) method was developed for screening plant extract for potential alpha amylase (AA) inhibitory activity. The method was validated against a well established UV method. Overall, the proposed method was shown able to detect plants with significant alpha amylase inhibitory activity but not those with rather clinically insignificant activities. Fifty plant species were screened using both the proposed CE method and the UV method and seven plant species were found to possess significant AA inhibitory activities. Two plant species were proved to have alpha amylase inhibitory activity for the first time.

In Vitro and In Vivo Evaluation of Casein as a Drug Carrier for Enzymatically Triggered Dissolution Enhancement from Solid Dispersions

Due to its unique properties, such as biodegradability, biocompatibility, high amphiphilic property, and micelle formation, casein (CS) has been increasingly studied for drug delivery. We used CS as a drug carrier in solid dispersions (SDs) and evaluated the effect of its degradation by trypsin on drug dissolution from the dispersions. SDs of CS and mefenamic acid (MA) were prepared by physical mixing, kneading, and coprecipitation methods. In comparison to pure MA, the dispersions were evaluated for drug–protein interaction, loss of drug crystalinity, and drug morphology by differential scanning calorimetry, X-ray diffractometry, Fourier transform infrared spectroscopy, and scanning electron microscopy. Drug dissolution from the dispersions was evaluated in simulated intestinal fluid as enzyme free and trypsin-enriched media. Furthermore, in vivo drug absorption of MA from CS-MA coprecipitate was evaluated in rats, in comparison with a reference SD of polyethylene glycol and MA (PEG-MA SD). Relative to other CS preparations, CS-MA coprecipitate showed the highest loss of drug crystallinity, drug micronization, and CS-MA interaction. CS remarkably enhanced the dissolution rate and extent of MA from the physical and kneaded mixtures. However, the highest dissolution enhancement was obtained when MA was coprecipitated with CS. Trypsin that can hydrolyze CS during dissolution resulted in further enhancement of MA dissolution from the physical and kneaded mixtures. However, a corresponding retardation effect was obtained for the coprecipitate. In correlation with in vitro drug release, CS-MA coprecipitate also showed significantly higher MA bioavailability in rats than PEG-MA SD.

Diclofenac-bismuth complex: Synthesis, physicochemical, and biological evaluation

Diclofenac-bismuth complexation was attempted by mixing diclofenac sodium (Na) and bismuth-subcitrate aqueous solutions at diclofenac:bismuth molar ratio of 3:1. A solid precipitate was obtained and isolated. The precipitate was characterized for stoichiometric ratio of diclofenac-bismuth complexation using capillary electrophoresis, which showed 1:1 complexation. In addition, nuclear magnetic resonance and Fourier transform infrared analysis were performed for the isolated solid complex and indicated that bismuth was in coordinate bond formation with the carboxylate group of diclofenac. In comparison with diclofenac Na powder, the complex was evaluated as an aqueous suspension for in vitro drug dissolution. The complex exhibited a faster dissolution rate than and similar dissolution extent as diclofenac Na. In comparison with an aqueous solution of diclofenac Na and an aqueous suspension of physical mixture of diclofenac acid (suspended) and bismuth-subcitrate (dissolved), the aqueous complex suspension was evaluated for ulcerogenic effect in rats upon oral administration. The complex led to more gastric ulceration than diclofenac Na, which was not in accordance with the antiulcer properties of bismuth. This antiulcer effect was shown as the physical mixture administration was accompanied with lower gastric ulceration than diclofenac Na administration. These gastric ulceration results were explained in terms of the difference in particle size between solid diclofenac acid formed as a result of the complex breakdown in an acidic medium (0.1 M HCl to simulate the gastric fluid) and that formed as a result of diclofenac Na neutralization. Diclofenac acid particles formed from the complex breakdown were of average size, three times smaller of those formed as a result of diclofenac Na protonation. This difference in particle size was correlated with the higher gastric ulceration associated with the complex than with diclofenac Na in terms of higher coverage of the gastric mucosa with diclofenac, and consequently, higher local ulceration.

A New HPLC Approach for the Determination of Hydrophilic and Hydrophobic Components: The Case of Pseudoephedrine Sulfate and Loratadine in Tablets

Effective isocratic separations of decongestants and antihistamines is a challenging analytical task due to wild differences in their lipohilicities (hydrophilic decongestants and hydrophobic antihistamines). In this paper a new approach for resolving such a problem is described taking pseudoephedrine sulfate and loratadine as an example. The chromatographic behavior of pseudoephedrine sulfate and loratadine on RP C18 and C8 columns were studied in presence and absence of sodium lauryl sulfate (SLS). The effect of combining two different types of stationary phases (cyano and C18 or C8) on the relative retention of the two compounds was investigated. In conclusion, it was found that the combination of a C18 column followed by a standard cyano column provides a stationary phase that separates both compounds effectively and within a reasonable time. This approach was compared to a literature method and demonstrated to have superior selectivity.