Isolation and identification of two galangin metabolites from rat urine and determination of their in vitro hypolipidemic activity

Purpose: To investigate the lipid-lowering activity of two metabolites of galangin, namely, galangin-3-Oβ-D-glucuronic acid (GG-1) and galangin-7-O-β-D-glucuronic acid (GG-2). Methods: Female Sprague-Dawley rats were orally administered with galangin. The two metabolites of galangin were isolated from urine sample and purified using Sephadex LH-20 and semi-preparative high performance liquid chromatography (HPLC). The structures of the metabolites were identified by analyzing spectroscopic data. Hypolipidemic activity was evaluated in HepG2 cells. The down- or upregulation of lipogenic genes was detected using real-time quantitative polymerase chain reaction (qPCR). Results: Both metabolites of galangin showed hypolipidemic activity. These activities are closely associated with the down-regulation of lipogenic genes such as SREBP-1a, SREBP-1c, and SREBP-2 transcription factors, and the downstream genes such as FAS, ACC, and HMGR were revealed by realtime qPCR data. Conclusion: The results show that both metabolites possess better lipid-lowering activities than galangin. These hypolipidemic activities are closely associated with inhibiting key genes or proteins that regulated the biosynthesis of both cholesterol and triglycerides.

Antitumor effect and mechanism of action of polysaccharides extracted from Polygonum perfoliatum L whole plant in human lung carcinoma A549 cell line

Purpose: To optimize the extraction conditions of polysaccharides from Polygonum perfoliatum L. (PSDP) and to evaluate their anti-tumor activities on A549 cell line. Methods: Extraction of PSDP was optimized using Box-Behnken design (BBD). Three factors of response surface methodology (RSM) including extraction time, ratio of water to raw material and number of extractions were employed to optimize the yield of PSDP. The cytotoxic effect of PSDP on human lung carcinoma A549 cell line was evaluated in vivo, while its effects on expressions of caspase3, caspase-9, Bcl-2 and Bax were determined by western blot assay. Result: BBD was significant and applicable to PSDP extraction. Based on the contour plots, response surface plots and variance analysis, it predicted that the optimum conditions for PSDP extraction were: 1.58 h (extraction time); 30.18 mL/g (ratio of water to raw material); and 2.02 (number of extractions). PSDP had significant inhibitory effect on the growth of A549 cells in a concentration- and timedependent manner (p < 0.05). After treatment with PSDP, caspase-3, caspase-9 and Bax were significantly up-regulated (p < 0.05), whereas Bcl-2 was down-regulated, all concentration-dependently. Conclusion: RSM analysis is an appropriate method to optimize PSDP extraction. The results also indicate that PSDP has significant anti-tumor effect against A549 cells, most likely via inducing mitochondria-mediated apoptosis.

Anticonvulsant and sedative effect of Fufang Changniu pills and probable mechanism of action in mice

Purpose: To investigate the anticonvulsant and sedative effects of Fufang Changniu Pills (FCP) and its probable mechanism of action in mice. Methods: The water decoction of FCP was prepared and the main constituents were determined by high performance liquid chromatography (HPLC). The anticonvulsant activities of FCP were evaluated by maximal electroshock (MES) and pentylenetetrazole (PTZ)-induced seizures in mice. Pentobarbital sodium-induced sleeping time and locomotor activity measurements were performed to evaluate the sedative effects of FCP in mice. Finally, PTZ-induced chronic seizures were established, and expressions of gamma-aminobutyric acid A receptor (GABA-A) and glutamic acid decarboxylase 65 (GAD65) in the brains of the mice were assayed by western blot in order to explore the probable mechanisms of action of the drug. Results: Gallic acid, liquiritin, cinnamyl alcohol, cinnamic acid and glycyrrhizic acid were detected in FCP decoction. FCP (50, 100 and 200 mg/kg) showed significant anticonvulsant and sedative effects on epileptic mice induced by MES (p < 0.05) and PTZ (p < 0.05). Moreover, pentobarbital sodium-induced sleeping time and locomotor activity tests showed that FCP possesses sedative effect (p < 0.05). Western blot data indicate that FCP significantly up-regulated GABA-A and GAD 65 in the brains of chronic epileptic rats (p < 0.05). Conclusion: FCP has significant anticonvulsant and sedative effects, and the mechanism of its action may be related to the up-regulation of GABA-A and GAD 65 in mice brain.

Bioprospecting for culturable actinobacteria with antimicrobial properties isolated from rivers in Colombian Orinoquia

Purpose: To Isolate and characterize Actinobacteria with antimicrobial activity from Guaviare River (Colombia). Methods: Water and sediment samples were collected from Guaviare River. Direct plating, heat and CaCO3 methods were used to isolate Actinobacteria. Six bacterial strains were tested using T-Streak method: Escherichia coli ATCC 23724, Staphylococus aureus ATCC 25923, Acinetobacter baumannii ATCC 19606, Bacillus subtilis ATCC 21556, Klebsiella pneumoniae ATCC 700603, Chromobacterium violaceum ATCC 31532. Strains of Fusarium sp. H24, Trichoderma harzianum H5 and Colletotrichum gloeosporioides were tested using Kirby-Bauer method. Isolates with high antimicrobial activity were selected for further taxonomic identification. Results: A total of 374 actinobacteria isolates were obtained. Seven isolates exhibited high antimicrobial activity (p < 0.05) and were confirmed as members of Streptomycetaceae family. Of these, three isolates showed differential phenotypic and genotypic profiles, indicating that they may represent new species. Conclusions: To date, this is the first study of this type in Colombian Orinoquia and indicates that this promising source of Actinobacteria from aquatic sediments with the ability to produce antimicrobial secondary metabolites.

Synthesis and antibacterial study of some s-substituted aliphatic analogues of 2-mercapto-5-(1-(4-toluenesulfonyl) piperidin-4-yl)-1,3,4-oxadiazole

Purpose: To synthesize a series of analogues of 1,3,4-oxadiazole and to evaluate their antibacterial activity. Methods: Ethyl piperidin-4-carboxylate (1) was mixed with 4-toluenesulfonyl chloride (2) in benignant conditions to yield ethyl 1-(4-toluenesulfonyl)piperidin-4-carboxylate (3) and then 1-(4- toluenesulfonyl)piperidin-4-carbohydrazide (4). Intermolecular cyclization of 4 into 2-mercapto-5-(1-(4- toluenesulfonyl) piperidin-4-yl)-1,3,4-oxadiazole (5) was obtained on reflux with CS2 in the presence of KOH. Molecule 5 was stirred with alkyl halides, 6a-i, in DMF in the presence of LiH to synthesize the final compounds, 7a-i. The structures of these molecules were elucidated by Fourier transform infra-red (FTIR) spectroscopy, proton nuclear magnetic resonance (1H-NMR) and electron impact mass spectrometry (EI-MS). Antibacterial activity was evaluated against five bacterial strains, namely, Salmonella typhi, Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus and Bacillus subtilis, with ciprofloxacin used as standard antibacterial agent. Results: Out of nine synthesized derivatives, compound 7a was the most active against three bacterial strains, S. typhi, E. coli and P. aeruginosa, with minimum inhibitory concentration (MIC) of 9.11 ± 0.40, 9.89 ± 0.45 and 9.14 ± 0.72 μM, respectively, compared with 7.45 ± 0.58, 7.16 ± 0.58 and 7.14 ± 0.18 μM, respectively, for the reference standard (ciprofloxacin). Similarly, compounds 7a - 7c showed relatively good antibacterial activity against B. subtilis strain while compound 7e - 7g revealed good results against S. typhi bacterial strain. Conclusion: The results indicate that S-substituted derivatives of the parent compound are more effective antibacterial agents than the parent compound, even with minor differences in substituents

Determination of acrylamide levels in selected commercial and traditional foods in Syria

Purpose: To determine acrylamide (AA) levels in different brands of commercial and traditional foodstuffs available in Syria by ultra-performance liquid chromatography-mass spectrometery (UPLCMS). Methods: A total of 63 samples were analyzed. Food samples were defatted by hexane and then extracted with methanol 98 % in a vortex mixer. Thereafter, Carrez I and Carrez II were added to precipitate proteins from the co-extractives and then centrifuged to obtain a clear aqueous extract that was evaporated to dryness. The extract was dissolved in 1 mL of water, eluted through a preconditioned Oasis HLB cartridge and then filtered. The filtrate was analyzed by UPLC-MS/MS to determine AA content. Results: Among the commercial foods tested, the highest acrylamide quantity was found in potato products (396 ± 3.59 – 1844 ± 5.29 μg kg−1) and the lowest in corn products (183 ± 2.64 – 366 ± 4.58 μg kg−1). This was followed by biscuits (57 ± 2.64 – 1433 ± 2.51 μg kg−1), breakfast cereals (121 ± 8.73 – 245 ± 3.60 μg kg−1), bread (119 ± 1.73 – 263 ± 3.60 μg kg−1) and then coffee (113 ± 2.64 - 64 ± 3.05 μg kg−1). Regarding traditional foods, the highest level of AA was found in AL- Mshabak (481 ± 2.08 μg kg−1) and AL-Awamat (421 ± 2.64 μg kg−1) followed by AL-Namora (282 ± 4.35 μg kg−1) and AL-Kenafa (242 ± 2.64 μg kg−1). It was also observed that the lowest amount of AA was in fried bread (230 μg kg−1), AL-Fatayer (192 ± 3.51 μg kg−1) and AL-Baqlawa (172 ± 4.35 μg kg−1) while Eid Aqras (130 ± 4.58 μg kg−1) and AL-Brazeq (167 ± 3.78 μg kg−1) contained the least amount of AA. Conclusion: The results indicate that the highest levels of AA are found in the most commonly consumed foods. There was significant difference (p < 0.05) in AA levels among different food items and within different brands of the same product.

Liquid chromatographic-mass spectrometric method for determination of drug content uniformity of two commonly used dermatology medications in a split-tablet dosage form

Purpose: To develop and validate a simple, efficient and reliable Liquid chromatographic-mass spectrometric (LC-MS/MS) method for the quantitative determination of two dermatological drugs, Lamisil® (terbinafine) and Proscar® (finasteride), in split tablet dosage form. Methods: Thirty tablets each of the 2 studied medications were randomly selected. Tablets were weighed and divided into 3 groups. Ten tablets of each drug were kept intact, another group of 10 tablets were manually split into halves using a tablet cutter and weighed with an analytical balance; a third group were split into quarters and weighed. All intact and split tablets were individually dissolved in a water: methanol mixture (4:1), sonicated, filtered and further diluted with mobile phase. Optimal chromatographic separation and mass spectrometric detection were achieved using an Agilent 1200 HPLC system coupled with an Agilent 6410 triple quadrupole mass spectrometer. Analytes were eluted through an Agilent eclipse plus C8 analytical column (150 mm × 4.6 mm, 5 μm) with a mobile phase composed of solvent A (water) containing 0.1% formic acid and 5mM ammonium formate pH 7.5, and solvent B (acetonitrile mixed with water in a ratio A:B 55:45) at a flow rate of 0.8 mL min-1 with a total run time of 12 min. Mass spectrometric detection was carried out using positive ionization mode with analyte quantitation monitored by multiple reaction monitoring (MRM) mode. Results: The proposed analytical method proved to be specific, robust and adequately sensitive. The results showed a good linear fit over the concentration range of 20 - 100 ng mL-1 for both analytes, with a correlation coefficient (r2) ≥ 0.999 and 0.998 for finasteride and terbinafine, respectively. Following tablet splitting, the drug content of the split tablets fell outside of the proxy USP specification for at least 14 halves (70 %) and 34 quarters (85 %) of FIN, as well as 16 halves (80 %) and 37 quarters (92.5 %) of TBN. Mean weight loss, after splitting, was 0.58 and 2.22 % for FIN half- and quarter tablets, respectively, and 3.96 and 4.09 % for TBN half- and quarter tablets,respectively. Conclusion: The proposed LC-MS/MS method has successfully been used to provide precise drug content uniformity of split tablets of FIN and TBN. Unequal distribution of the drug on the split tablets is indicated by the high standard deviation beyond the accepted value. Hence, it is recommended not to split non-scored tablets especially, for those medications with significant toxicity

Determination of borneol and other chemical compounds of essential oil of Dryobalanops aromatica exudate from Malaysia

Purpose: To determine borneol and other chemical compounds of essential oil derived from the exudate of Dryobalanops aromatica aromatica in Malaysia. Methods: Exudate was collected from D. aromatica and subjected to fractional distillation to obtain essential oil. Gas chromatography-mass spectrometry (GC-MS) was performed to characterize the composition of the isolated essential oil. Results: Essential oil (7.58 %) was obtained with the highest yield (3.24 %) in the first 2 h of fractional distillation. Thirty compounds which accounted for 97.56 % of total essential oil composition were identified by GC-MS, and they include sesquiterpenes (46.87 %), monoterpenes (31.05 %), oxygenated monoterpenes (16.76 %) and oxygenated sesquiterpenes (2.13 %). Borneol (0.74 %) was also detected in the essential oil. Conclusion: Borneol and other terpenoid compounds are present in the essential oil of the exudate of D. aromatica

Identification of metabolites of kurarinone from Sophora flavescens Ait in rat urine by ultra-performance liquid chromatography with linear ion trap orbitrap mass spectrometry

Purpose: To study the in vivo metabolism of kurarinone, a lavandulyl flavanone which is a major constituent of Kushen and a marker compound with many biological activities, using ultra-performance liquid chromatography coupled with linear ion trap Orbitrap mass spectrometry (UPLC-LTQ-Orbitrap- MS). Methods: Six male Sprague-Dawley rats were randomly divided into two groups. First, kurarinone was suspended in 0.5 % carboxymethylcellulose sodium (CMC-Na) aqueous solution, and was given to rats (n = 3, 2 mL for each rat) orally at 50 mg/kg. A 2 mL aliquot of 0.5 % CMC-Na aqueous solution was administered to the rats in the control group. Next, urine samples were collected over 0-24 h after the oral administrations and all urine samples were pretreated by a solid phase extraction (SPE) method. Finally, all samples were analyzed by a UPLC-LTQ-Orbitrap mass spectrometry coupled with an electrospray ionization source (ESI) that was operated in the negative ionization mode. Results: A total of 11 metabolites, including the parent drug and 10 phase II metabolites in rat urine, were first detected and interpreted based on accurate mass measurement, fragment ions, and chromatographic retention times. The results were based on the assumption that kurarinone glucuronidation was the dominant metabolite that was excreted in rat urine. Conclusion: The results from this work indicate that kurarinone in vivo is typically transformed to nontoxic glucuronidation metabolites, and these findings may help to characterize the metabolic profile of kurarinone.

Structural determination and gynecological tumor diagnosis using antibody chip captured proteins

Purpose: To identify markers for gynecological tumor diagnosis using antibody chip capture. Methods: Marker proteins, including cancer antigen 153 (CA153), CA125, and carcinoembryonic antigen (CEA), were analyzed using antibody chip capture of serum samples. Fifteen agglutinin types that specifically recognized five common glycans (fucose, sialic acid, mannose, N - acetylgalactosamine, and N-acetylglucosamine) were used to detect marker protein glycan levels. The levels of CA153, CA125, and CEA from 49 healthy control samples, 31 breast cancer samples, 24 cervical cancer samples, and 19 ovarian cancer samples were used to measure the glycan levels of these marker proteins. Results: In breast cancer samples, CA153 and CA125 were down-regulated (p < 0.01), while differences in ovarian cancer samples were not statistically significant (p > 0.01). The total accuracy was 85.1 %, with 96.8 % accuracy for breast cancer, 75 % in cervical cancer, and 78.9 % in ovarian cancer. Cross-validation analyses showed that breast cancer had 93.5 % accuracy, cervical cancer was 66.7 %, and ovarian cancer was 68.4 %, leading to 78.4 % total accuracy (58/74). Conclusions: The results indicate that better clinical diagnosis of gynecological tumors can be obtained by monitoring changes in glycan levels of serum proteins and types of proteoglycan changes.

Factors associated with hypoglycemia episodes in hospitalized type 2 diabetes mellitus patients in a tertiary health facility in Malaysia

Purpose: To determine the factors associated with severity of hypoglycemia in hospitalized type 2 diabetes mellitus patients in a tertiary health facility in Malaysia. Methods: This retrospective study involved 207 hospitalised T2DM patients with hypoglycaemia episodes from January 2008 to December 2012 and was conducted in University Malaya Medical Centre, Petaling Jaya, Malaysia. Patients were classified into 2 groups, viz, those who had hypoglycaemia on admission and those who had hypoglycaemia during hospital stay. Patients with hypoglycemia on admission were those admitted due to hypoglycemia while patients with hypoglycemia during hospital stay were those admitted due to other causes but subsequently developed hypoglycemia during hospitalization. Results: The results for the 207 patients investigated show that most of the patients (72.2 %) were asymptomatic during hypoglycemic episodes. The majority of the episodes (57.4 %) experienced by the patients were mild hypoglycemia (< 3.9 mmol/L). Old age (p = 0.011) and presence of stroke (p = 0.033) were found to be significantly associated with severe hypoglycemia (< 2.2 mmol/L) while concurrent use of opioid (p = 0.008) was associated with mild hypoglycemia. Conclusion: The identification of the underlying factors associated with severity of hypoglycemia may help in preventing and resolving hypoglycemia in T2DM patients.

Complications associated with malnutrition in elective surgical patients in a Malaysian setting

Purpose: To identify the level of malnutrition and complications observed in Malaysia. Methods: A prospective, observational study was conducted with the objectives of identifying the degree of malnutrition, complications and the need for nutritional support in elective surgical patients. Collection of data was performed in local tertiary hospitals using a Patient Generated Subjective Global Assessment (PG - SGA) nutritional screening tool and medical records. Results: A total of 50 patients electively admitted for surgery were included. The results demonstrated that there was a significant increase in malnourished patients post-surgery compared to pre-surgery (p = 0.0001). Most interesting was the significant number of complications observed post-surgery compared to pre-surgery (p = 0.035) which was associated to the poor level of nutrition. The most common type of complications noted post-surgery were nosocomial infection, wound infection and readmission. Conclusion: Malnutrition is prevalent pre- and post-operatively in Malaysia. Therefore, focus on the outcome of these malnourished patients should be performed to reduce complications associated with poor nutrition.

Prevalence of chronic diseases in private healthcare sector of South Africa: A threat to public health

Purpose: To evaluate the prevalence of patients suffering from registered chronic disease list (CDL) conditions in a section of the South African private health sector from 2008 - 2012. Methods: This study was a retrospective analysis of the medicine claims database of a nationally (South African) representative Pharmacy Benefit Management (PBM) company data between 2008 and 2012. Statistical analysis was used to analyse the data. Descriptive analysis was performed to calculate the prevalence of CDL conditions for the entire population, and stratified by age and gender. However, MIXED linear modelling was used to determine changes in the average number of CDL conditions per patient, adjusted for age and gender from 2008 - 2012. Results: An increase of 0.20 in chronic diseases was observed from 2008 - 2012 in patients having any CDL condition, with an average of 1.57 (1.57 - 1.58, 95 % CI) co-morbid CDL conditions in 2008 and 1.77 (1.77 - 1.78, 95 % CI) in 2012. This increase in average number of CDL conditions per patient between 2008 and 2012 was statistically significant (p < 0.05), but with no large practical significance (d < 0.8). Conclusion: Prevalence of patients with CDL conditions along with risk of co-morbidity has been increasing with time in the private health sector of South Africa. Risk of increased co-morbidity with age and among different genders was prevalent.

Clinical effect of intravenous thrombolysis combined with nicorandil therapy in patients with acute ST-segment elevation myocardial infarction

Purpose: To evaluate the effectiveness of intravenous thrombolysis in combination with nicorandil in the treatment of acute ST-segment elevation myocardial infarction (STEMI). Methods: Patients who developed acute STEMI and underwent intravenous thrombolysis in the hospital were selected and divided into observation group (n = 128) and control group (n = 114). Besides thrombolytic therapy, the observation group was also given 20 mg of nicorandil. The control group received conventional thrombolytic therapy only. Clinical effects and rehabilitation of patients were observed. Results: Cardiac troponin I (cTNI) level of the observation group was 4.0 ± 1.5, 8.3 ± 2.8 and 9.8 ± 3.9 after 4, 12 and 24 h, respectively, which is much lower than 5.8 ± 1.4, 11.4 ± 2.7 and 13.2 ± 4.2 in the control group (p < 0.05). ST-segment resolution of observation group was higher (44 ± 14, 52 ± 17, 69 ± 21 and 80 ± 18) % at different time points, compared with the control group (p < 0.05). The proportion of patients with Curtis-Walker score > 3 points, and ventricular wall motion score (4.70 %; 1.38 ± 0.11) in the observation group were both lower than those of the control group (21.00 %; 1.43 ± 0.15) (p < 0.05). The difference in adverse cardiac events between the observation group (N = 6, 4.70 %) and control group (N = 12, 10.50 %) was not statistically significant (p > 0.05) Conclusion: Combining intravenous thrombolysis with nicorandil therapy can enhance myocardial perfusion level, reduce myocardial damage, improve cardiac function and decrease risk of arrhythmia for acute STEMI patients.

Natural antioxidant vitamins: A review of their beneficial roles in management of diabetes mellitus and its complications

Diabetes mellitus is a complex and progressive metabolic disease which is associated with multiple complications. Chronic hyperglycaemia is the defining characteristic of diabetes mellitus. Hyperglycaemia leads to generation of free radicals and induces oxidative stress, which has become the chief factor that leads to diabetic complications. This review supports the use of antioxidant vitamins as therapeutic agents in the management of diabetes mellitus and its complications, and also provides an insight into the potential pharmacological effects of natural antioxidant vitamins in diabetic conditions. These antioxidant vitamins can be used as safe supplements to manage the occurrence and complications of the disease. Selected studies have reported on the beneficial effects of antioxidant vitamins in experimental models. The involvement of oxidative stress in diabetes and its complications has made the use of natural antioxidant vitamins (free radical scavengers) from plants inevitable as they may be very effective and safer in the management of diabetes.