Testing Neurotransmitters for Toxicity with a Luminescent Biosensor: Implications for Microbial Endocrinology – Pages 88-94

Alexander V. Oleskin1, Elena V. Sorokina2, Alevtina P. Zarubina3 and Inna M. Parkhomenko4
1Doctor of Biological Sciences, Professor, Department of General Ecology, School of Biology, Lomonosov Moscow State University, Moscow, Russia; 2School of Biology, Lomonosov Moscow State University, Moscow, Russia; 3Department of Microbiology, School of Biology, Lomonosov Moscow State University, Moscow, Russia; 4Department of Biophysics, School of Biology, Lomonosov Moscow State University, Moscow, Russia
DOI: https://doi.org/10.6000/1927-5951.2017.07.03.3

Abstract: Background: The human organism is a complex superorganism including numerous eukaryotic, eubacterial, and archaean cells. The qualitative and quantitative assessment of the microbiota toxicity of chemical agents, i.e., their inhibitory effects on the microbial inhabitants of the human organism in health and disease, seems to hold much value in this context. In this work, a bacterial luminescence-based express test system for microbiota toxicity is applied to neurotransmitters such as serotonin, dopamine, norepinephrine, and histamine.

Methods: The biosensor was based on a GM Escherichia coli K12 strain (TGI) that contained the lux operon of the luminescent soil bacterium Photorhabdus luminescencens ZMI. The biosensor was exposed to the action of the tested neurotransmitters for 5 to 60 minutes The intensity of bacterial luminescence (counts.sec-1) was monitored in the control and the experimental samples with a Biotoks 6 ms luminometer (Russia); the toxicity index (T) of the neurotransmitters was determined.

Results: A marked toxic effect on bioluminescence was produced by serotonin, histamine, and dopamine at concentrations exceeding 80 µg/ml, 100 µg/ml, and 1 mg/ml, respectively. At lower concentration, these neurotransmitters were “negatively toxic”, i.e. stimulatory in terms of the effect on bacterial luminescence. In contrast, norepinephrine inhibited luminescence at all concentrations tested.

Conclusions: The bacterial luminescence-based testing method is applicable to the assessment of the destructive and stimulatory effects of neurotransmitters; the data obtained are of microbiological and clinical relevance.

Keywords: Bioassay, bioluminescence, toxicity, neurotransmitters, serotonin, dopamine, histamine, norepinephrine.

Inspection of Phytochemical Content and In Vitro Antioxidant Profile of Gnaphalium luteoalbum L.: An Unexplored Phytomedicine – Pages 136-146

Md. Sahab Uddin1, G.M. Sala Uddin1, Mst. Marium Begum2, Yesmin Begum1, Oscar Herrera-Calderon3, Md. Manjurul Islam4 and Mohamed M. Abdel-Daim5
1Department of Pharmacy, Southeast University, Dhaka, Bangladesh; 2Department of Pharmacy, East West University, Dhaka, Bangladesh; 3Academic Department of Pharmaceutical Sciences, Faculty of Pharmacy and Biochemistry, Universidad Nacional San Luis Gonzaga de Ica, Ica, Peru; 4Department of Pharmacy, University of Development Alternative, Dhaka, Bangladesh; 5Pharmacology Department, Faculty of Veterinary Medicine, Suez Canal University, Ismailia, Egypt
DOI: https://doi.org/10.6000/1927-5951.2017.07.03.10

Abstract: Background: Oxidative stress is intensely linked with several pathological manifestations. Searching for medicinal plant with the superior safety profile for the treatment of oxidative stress related disorders are ongoing due to multiple unwanted effects associated with synthetic antioxidants. Therefore the purpose of this study was to examine the phytochemical content, in vitro antioxidant potentiality of crude methanol extract (CME), carbon tetrachloride fraction (CTF), petroleum ether fraction (PEF), chloroform fraction (CLF) and ethyl acetate fraction (EAF) of aerial parts of Gnaphalium luteoalbum (GL) L.

Methods: The aerial parts of the GL were extracted with methanol followed by fractionation using carbon tetrachloride, petroleum ether, chloroform and ethyl acetate. The phytochemical screening of this plant was performed by using standard methods to evaluate the existence of alkaloids, carbohydrates, phenols, flavonoids, saponins, tannins, terpenoids and fixed oils. Antioxidant potentiality was estimated by, 2,2-diphenyl-1-picrylhydrazyl (DPPH), hydroxyl (OH) and nitric oxide (NO) radical scavenging tests. Total antioxidant capacity (TAC), total phenolic content (TPC) and total flavonoid content (TFC) were also measured.

Results: Phytochemical analysis of the aerial parts of GL confirmed the presence of carbohydrates, phenols, flavonoids and saponins in crude extract and its all fractions. The CME showed the highest scavenging activity (43.28%) with IC50 of 398.49 μg/mL in the DPPH radical scavenging test. The IC50 values of EAF, CME were statistically significant (P < 0.05, P < 0.01) with respect to ascorbic acid (ACA). For OH and NO radical scavenging tests maximum scavenging (48.39%, 69.64%) was also reported for CME compared to CTF, PEF, CLF and EAF. Compared to ACA, in case of OH and NO radical scavenging activities the IC50 values of CME were markedly significant (P < 0.01, P < 0.05). In the TAC test, CME showed the highest antioxidant activity (absorbance, 2.6 nm) related to other fractions. TPC was found to be the highest in the CME (115.96 mg of gallic acid equivalent/g of dried extract) rather than other fractions. The ranking order of CTF, PEF, CLF, EAF and CME for TFC was 48.67 < 55.75 < 65.29 < 71.35 < 82.29 mg quercetin equivalent/g of dried extract.

Conclusion: The existing study suggested that CME of the aerial parts of GL can be used as a natural source of antioxidant which might be effective towards preventing or slowing oxidative stress related disorders.

Keywords: Phytochemical, Antioxidant activity, Gnaphalium luteoalbum, Oxidative stress, Synthetic antioxidants.

Evaluation of Protective Effect of Methyl Sulfonyl Methane on Colon Ulcer Induced by Alendronate – Pages 130-135

Safaa William Mdawar1, Shaza Anwar Al Laham1 and Ahmad Izzat Al-Manadili2
1Pharmacology & Toxicology Department, Faculty of Pharmacy, Damascus University, Damascus, Syria; 2Oral Histopathology Department, Faculty of Dentistry, Damascus University, Damascus, Syria
DOI: https://doi.org/10.6000/1927-5951.2017.07.03.9

Abstract: Background: Bisphosphonates represent a new class of drugs that have shown very promising therapeutic efficacy in the treatment of a number of diseases associated with abnormally accelerated bone resorption including osteoporosis, Paget’s disease, and hypercalcemia of malignancy. As an increasing number of these drugs become available for clinical use in the treatment of the millions of individuals with these skeletal diseases, evidence is becoming available that their chronic usage may be associated with a number of gastrointestinal side effects, such as diarrhea, abdominal pain and inflammation, and erosions and ulceration of the upper gastrointestinal tract. Methyl sulfonyl methane (MSM) which is used for osteoarthritis was used to prevent bisphosphonates ulceration. Our objective was to study the preventive effect of MSM against colon ulcer induced by Alendronate (ALN) in rats. The experiments had been done on 8 white wistar rats for each group. The gastric ulcer has been induced by administration of Alendronate (20mg/kg/day) by gavage for 4 days. MSM (400mg/kg/day) has been given for the protective group for 4 days before administration of Alendronate. The ulcers in rats’ colon were examined histologically and microscopically. The results showed that administration of MSM before Alendronate inducing ulcer led to a reduction in ulceration and showed significant difference comparing with morbidity group.

Conclusion: MSM (400 mg/kg/day) has protective effect of colon ulcer induced by alendronate.

Coumarin and Safrole Content in Cinnamon-Flavored Food Products on the Syrian Market – Pages 124-129

Raw’aa Solaiman and Joumaa Al-Zehouri
Department of Food Control and Analytical Chemistry, College of Pharmacy, Damascus University, Damascus, Syria
DOI: https://doi.org/10.6000/1927-5951.2017.07.03.8

Abstract: Some plants that are processed into foods often contain natural substances that may be hazardous to human health. One example is coumarin, which is known to cause liver and kidney damage in rats, mice and probably humans. Coumarin is found in different Cinnamomum species such as Cinnamomum cassia, Cinnamomum loureiroi, and Cinnamomum burmannii; all commonly referred to as cassia. Another hazardous substance is Safrole. Safrole is reasonably anticipated to be a human carcinogen based on sufficient evidence of carcinogenicity from studies in experimental animals; and is mainly present in cinnamon leaf oil and could be a possible contaminant in cinnamon powder. European Council’s Directive on food flavourings 88/388/EEC limits safrole in foodstuffs to 1ppm. The content of coumarin is regulated in the European Regulation (EC) No 1334/2008. In the present study, coumarin and safrole levels were analyzed in locally bought cinnamon samples and cinnamon-flavored food products using a validated HPLC method with diode array detector (DAD). Appreciable amounts of coumarin were found in bakery products with concentrations up to 39.466 mg/kg in certain kinds of cookies, whereas safrole was undetectable. Our exposure data on coumarin in bakery products show that there is still a need for a continued regulation of coumarin in foods. A toxicological re-evaluation of coumarin with the aim to derive scientifically founded maximum limits should be conducted with priority.

Keywords: Cinnamon, Coumarin, Safrole, High-performance liquid chromatography, cinnamon-flavored foods.

Food Extrusion Technology: Initiatives to Address Food and Nutrition Insecurity in South Africa – Pages 116-123

Evanie D. Deenanath and Abdulkadir Egal
Centre of Sustainable Livelihoods, Vaal University of Technology, Private Bag X021, Vanderbijlpark, 1900, Science and Technology Park, Sebokeng, South Africa
DOI: https://doi.org/10.6000/1927-5951.2017.07.03.7

Abstract: The use of extrusion can be regarded as beneficial due to its short production time and wide variety of foods produced by this method. South Africa as a developing country has been involved in food extrusion since the 1980’s and this technology is gaining momentum in academic research areas. A number of research efforts related to extrusion in South Africa have shown the consumption of extruded dry beans can reduce plasminogen activator inhibitor levels in hyperlipidaemic men; the production of sorghum-cowpea extruded instant porridge resulted in a nutritional acceptable product and can be used to supplement the diet of young children to assist with protein deficiencies. Furthermore, research has proven extruder parameters play a role in the outcome of the product and can influence product properties. Based on these research initiatives, Vaal University of Technology/Centre of Sustainable Livelihoods (VUT/CSL) has acquired an Extrusion Pilot Plant to implement interdisciplinary research of nutrition and engineering science. The research will look at process optimisation studies to obtain maximum product output and evaluating nutritional compositions of the products under various conditions. It is hoped the future research efforts at VUT/CSL will address food and nutrition insecurity and showcase the pilot plant as a testing facility and potential advancement to commercialisation.

Keywords: Extrusion, Research, South Africa, Production, Nutrition.

Sliding Scale Regular Human Insulin for Identifying Critically Ill Patients Who Require Intensive Insulin Therapy and for Glycemic Control in those with Mild to Moderate Hyperglycemia – Pages 106-115

Sarah V. Cogle1, Susan E. Smith2, George O. Maish III3, Gayle Minard3, Martin A. Croce3 and Roland N. Dickerson2
1Department of Pharmacy Practice, Auburn University Harrison School of Pharmacy, Auburn, AL, 2Departments of Clinical Pharmacy and 3Surgery, University of Tennessee Health Science Center, Memphis, TN, USA
DOI: https://doi.org/10.6000/1927-5951.2017.07.03.6

Abstract: Two sliding scale regular human insulin (RHI) algorithms (SSI) were retrospectively evaluated to identify those who develop severe hyperglycemia (blood glucose (BG) > 180 mg/dL) and for glycemic management of continuously-fed, critically ill trauma patients with mild to moderate hyperglycemia (BG 126 to 179 mg/dL). Assignment of low or high SSI was based upon anticipated severity of difficulty in glycemic control. BG was obtained every 3 to 6 hours. Target BG range was 70 to 149 mg/dL. Patients who were unable to achieve a BG < 150 mg/dL with SSI and who required a continuous intravenous RHI infusion were identified. Twenty-five of 121 patients (21%) failed SSI necessitating more intensive insulin therapy. The low and high intensity SSI groups exhibited a baseline BG of 123 + 33 mg/dL and 164 + 20 mg/dL (P = 0.001). Average BG for each group was 129 ± 14 mg/dL and 145 ± 21 mg/dL (P = 0.001). Each group spent 20 ± 4 and 16 ± 5 hours/day within the target BG range (P = 0.001), respectively. Mild hypoglycemia (BG 40 – 60 mg/dL) occurred in 11% and 7% of patients from each group (P = N.S.). Severe hypoglycemia (BG < 40 mg/dL) occurred in zero and two (5%) patients, respectively (P = N.S). SSI served as a useful technique to identify those requiring more intensive insulin therapy and was safe and efficacious for continuously-fed, critically ill trauma patients with mild to moderate hyperglycemia.

Keywords: Hyperglycemia, hypoglycemia, insulin, critical care, trauma, enteral nutrition, parenteral nutrition.

Phase II Clinical Trial to Establish Efficacy of a Locally Appropriate Bivalent Anti Snake Venom in Pakistan – Pages 100-105

Naeem Quraishi1, Tasneem Ahmad1, Allah-Bux Ghanghro2, Abdullah Arejo3, Sharib S. Muhammad1 and Atta Chandio4
1ASV/ARV Serology Laboratory, Peoples Medical University Nawabshah, Pakistan; 2Institute of Biochemistry, Sindh University Jamshoro, Pakistan; 3Parasitology Department, Sindh Agriculture University, Tando Jam, Pakistan; 4Faculty of Community Medicine & Public Health Peoples University of Medical & Health Sciences for Women Nawabshah, Pakistan
DOI: https://doi.org/10.6000/1927-5951.2017.07.03.5

Abstract: Objective: This study was conducted to determine the efficacy of Snake anti-venom Immunoglobulin [IgG] manufactured by Anti-Snake Venom [ASV]/Anti-Rabies [ARV] Serology Laboratory, Health Department, Government of Sindh.

Methods: The prospective, observational single arm study was conducted after the approval of IRB. Study included six patients with viper [Echis carinatus sochureki] snakebites referred to the emergency ward of Peoples University of Medical & Health Sciences Hospital, Nawabshah and District Headquarter Hospital Mithi, Sindh, Pakistan with consultation of Clinical and Principal investigator. The study was conducted over a period of three months [August 2015 to November 2015]. All patients were given IV infusion of 10 mL [1 vial] investigational ASV diluted in 100 mL normal saline except one patient who received 5 mL management dose and 5 mL subsequent dose for the recovery of coagulopathy. The efficacy was assessed by Primary and secondary efficacy endpoints, i.e. the dose at which maximum no of patients were treated [permanent restoration of normal blood coagulation tested by 20-minute whole blood clotting test [20-minute WBCT] with minimum toxicity.

Results: All patients recovered from coagulopathy after receiving IV infusion of 10 mL investigational ASV diluted in 100 mL normal saline tested by 20-minute WBCT. Mean Recovery time was 9:15 ± 3:25 hours.

Conclusion: Safety and efficacy was assessed for the Bivalent Anti venom Immunoglobulin-NQ1 [IgG] manufactured by ASV/ARV Serology Laboratory, Health Department, Government of Sindh.

Keywords: Immunoglobulin [IgG], anti-snake venom [ASV], 20-minute WBCT, Echis carinatus sochureki [Lundi], Coagulopathy.

Isolation and Purification of β-Carotene from Morinda citrifolia as HPLC Standard and Active Pharmaceutical Ingredient – Pages 95-99

Rashidi Othman1, Fatimah Azzahra Mohd Zaifuddin2 and Norazian Mohd Hassan2
1International Institute for Halal Research and Training (INHART), Herbarium Unit, Department of Landscape Architecture, Kulliyyah of Architecture and Environmental Design, International Islamic University Malaysia, 53100 Kuala Lumpur, Malaysia; 2Department of Pharmaceutical Chemistry, Kulliyyah of Pharmacy, International Islamic University Malaysia, 25200 Kuantan, Malaysia
DOI: https://doi.org/10.6000/1927-5951.2017.07.03.4

Abstract: Qualitative and quantitative analysis of individual carotenoids content and composition are complicated, time consuming and in fact very costly. The crucial and vital part is the availability and reliability of the pure standards. Most of the individual carotenoids are commercially available either in natural or synthetic form but they are quite expensive and some of it not available in the market anymore. These problems strongly associated with the accuracy and reliability of High Performance Liquid Chromatography (HPLC) analysis data. Therefore, this study aimed to set up an analytical scheme of obtaining b-carotene standard from the leaves of Morinda citrifolia as one of the carotenoid standards for HPLC analysis. M. citrifolia has been selected due to its abundance throughout the year with tropical climate. The scheme via open column chromatography (OCC) established that the purity of β-carotene standard was 97% and the coefficient of correlation was 0.9923. However after 30 day storage period of time, the purity decreased to 95.46%. Although these had an effect on the carotenoid standard stability but it can be a reliable source of β-carotene standard for HPLC analysis as well as active pharmaceutical ingredient for cosmeceutical, nutraceutical, food and beverage industries.

Keywords: Carotenoid, β-carotene, Morinda citrifolia, Active pharmaceutical ingredient.

Characterisation of Carotenoid and Total Retinol Equivalent Content in Ulam and Medicinal Species as Alternative Food Intervention to Combat Vitamin A Deficiency – Pages 81-87

Rashidi Othman1, Fatimah Azzahra Mohd Zaifuddin2 and Norazian Mohd Hassan2
1International Institute for Halal Research and Training (INHART), Herbarium Unit, Department of Landscape Architecture, Kulliyyah of Architecture and Environmental Design, International Islamic University Malaysia, 53100 Kuala Lumpur, Malaysia; 2Department of Pharmaceutical Chemistry, Kulliyyah of Pharmacy, International, Islamic University Malaysia, 25200 Kuantan, Malaysia
DOI: https://doi.org/10.6000/1927-5951.2017.07.03.2

Abstract: Vitamin A deficiency (VAD) is one of the continuous leading causes of children and pregnant women death. To overcome this malnutrition which currently affected one-third of the world population, there is always renewed interest in exploring numerous dietary sources rich in carotenoids which some of them serve as pre-cursors to vitamin A (pro-vitamin A). It is important that affordable staple foods be as nutritious as possible because poverty limits food access for much of the developing world’s population. Therefore, this study was aimed to explore various dietary sources for carotenoids in 28 ulam and medicinal species which are commonly consumed by the local folks. Carotenoid extraction using organic solvents was performed and analysis employed in this study through High Performance Liquid Chromatography revealed seven types of carotenoids in the food matrices; neoxanthin, violaxanthin, lutein, zeaxanthin, β-cryptoxanthin, α-carotene and β-carotene. Interestingly, these carotenoids profiles were found in varying concentration and composition in different species as well as in different period or season. Total carotenoids content quantified in all of the samples lies between 1.315 ± 0.007 to 190.301 ± 3.427 µg/g DW where cekur manis has the highest content. The total vitamin A activity (in terms of retinol equivalent, RE) of every species is also included in this study. The results suggested that at least 20 of the ulam and medicinal species may be used as alternative food intervention to eliminate VAD as a public health concern.

Keywords: Carotenoid, Retinol Equivalent, Vitamin A Deficiency, pro-vitamin A, Ulam.

Effects of Antrodia Camphorata Mycelia Extract Containing Antroquinonol on Lowering Low-Density Lipoprotein Cholesterol: A Randomized Double-Blind Study – Pages 73-80

Miles Chih-Ming Chena, Pei-Ni Chena, Howard Hao-Yu Chenga, Wayne Ching-Cheng Weia, Ryuji Takedab, Mitsuko Moric and Kiichiro Mochidad
aDivision of Clinical Research, Golden Biotechnology Corp., Danshui Dist., 251, New Taipei City, Taiwan, R.O.C.; bDepartment of Nutritional Sciences for Well-being, Faculty of Health Sciences for Welfare, Kansai University of Welfare Sciences, 582-0026 Asahigaoka 3-11-1, Kashiwara city Osaka, Japan; cTokyo Branch of the Oriental Occupational Health Association, Oriental Ueno Health Checkup Center, Basement Floor 1, Suzunoya Building, 1-20-11, Ueno, Taito-shi, Tokyo, Japan; dRCT Japan Inc., Shibuya 2-21-1 8F, Shibuya Hikarie Building, Shibuya Ward, Tokyo, Japan
DOI: https://doi.org/10.6000/1927-5951.2017.07.03.1

Abstract: Objective: Antrodia camphorata is a type of true fungus that grows only on Cinnamomum camphora trees, also known as Cinnamomum kanehirae (“kashi”) in Taiwan. Antroquinonol is a characteristic component of A. camphorata mycelia extract and was previously shown to exhibit antitumor action and lower blood cholesterol (total cholesterol and low-density lipoprotein [LDL] cholesterol) in cellular and animal models. So, This study examined the ability of A. camphorata mycelia extract to reduce LDL cholesterol in humans.

Methods: We conducted a randomized double-blind trial in 26 subjects with either borderline LDL cholesterol (120–139 mg/dL; n = 11) or mildly elevated LDL cholesterol (140–159 mg/dL; n = 15). Participants ingested tablets containing either 25 mg of A. camphorata mycelia extract (antroquinonol: 0.68 mg; n = 14) or a placebo (n = 12) for 12 weeks.

Results: The test group showed a significant reduction in LDL cholesterol when compared with the placebo group after 12 weeks of tablet ingestion (p < 0.05), demonstrating the effects of A. camphorata mycelia extract on LDL cholesterol. A. camphorata mycelia extract also tended to reduce total cholesterol when compared with the placebo (p < 0.10). The borderline LDL cholesterol and mildly elevated LDL cholesterol subgroups showed a significant reduction in LDL cholesterol in subjects who ingested A. camphorata mycelia extract compared with those who ingested the placebo, again demonstrating the LDL cholesterol-lowering effect of the extract.

Conclusion: A. camphorata mycelia extract lowers LDL cholesterol in individuals with somewhat high LDL cholesterol levels.

This clinical trial was registered with the University Hospital Medical Information Network (UMIN no. # 000019670).

Keywords: Antrodia camphorate, Antroquinonol, LDL-cholesterol, LDL receptor genes, randomized placebo-controlled double-blind study.