Glucose oxidase as a model enzyme for antidiabetic activity evaluation of medicinal plants: In vitro and in silico evidence

Diabetes mellitus is a major public health problem in the world. In Africa, more than 80% of patients use plants for their treatment. However, the methods of validation of endogenous knowledge usually used are costly. The alternative method developed in this study aims at creating hyperglycemia <i>in vitro</i> and exploiting the metabolic pathway involving glucose oxidase for UV-visible spectrophotometric screening of medicinal plants’ antidiabetic activity. The evolution of glucose oxidation as a function of drug concentration is followed by UV-visible spectrophotometry. The formation of the stable complex between the enzyme and the inhibitor is studied using molecular docking. Drugs used (Gliben) and plant extracts exhibited an <i>in vitro</i> hypoglycemic effect by reducing exponentially, <i>in vitro</i>, the level of free glucose. The results also showed that <i>L. multiflora</i> is more active than <i>V. amygdalina</i> (IC₅₀: 1.36 ± 0.09 mg/mL Vs IC₅₀: 3.00 ± 0.54 mg/mL). Gliben (0.5 mg/mL) and <i>L. multiflora</i> (2 mg/mL) reduced both the rate of oxidation of glucose by glucose oxidase (catalytic power V_max: 0.84 ± 0.11 mg*mL^-1*min^-1 for Gliben and 1.72 ± 0.13 mg*mL^-1*min^-1 for ^{L. multiflora}); and the affinity of this enzyme for its substrate-glucose (K_M: 15.11 ± 2.72 mg*mL^-1 for Gliben and 9.17 ± 1.56 mg*mL^-1 for <i>L. multiflora</i>) when these results are compared to enzyme catalysis in the absence of inhibitor (V_max: 2.86 ± 0.44 mg*mL^-1*min-1; K_M: 8.07 ± 1.96 mg*mL^-1). The binding of GOX (1GAL) to selected phytocompounds derived from <i>L. multiflora</i> was confirmed by molecular docking. The most stable complexes were obtained for four compounds; <b>8</b> (-10.1±0.0 Kcal/mol), <b>6</b> (-9.5±0.1 Kcal/mol), <b>3</b> (-8.3±0.0 Kcal/mol) and <b>9</b> (-8.2±0.1 Kcal/mol). Among these, compounds <b>8</b> and <b>6</b> formed complexes with the enzyme stabilized by hydrogen bonds, the compound <b>8</b> forms 5 hydrogen bonds (<b>ASN514</b>, <b>ASP424</b>, <b>ARG95</b>, <b>TYP68</b>, <b>LEU65</b>) while compound <b>6</b> forms 2 hydrogen bonds (<b>ASN514</b> and <b>SER422</b>). However, no H-bonding interaction occurs in the complex that involves ligands <b>9</b> and <b>3</b> despite their high binding energy (-8.2±0.1 Kcal/mol and -8.3±0.0 Kcal/mol respectively). Glucose oxidase can serve as a marker enzyme for<i> in vitro</i> antidiabetic activity evaluation of medicinal plants.

Phytoremedial effect of fruit extract of Moringa oleifera on alloxan induced diabetic model in Swiss albino mice

Diabetes is a metabolic disorder and global health issue. It arises because of an absolute or relative insulin deficiency that causes hyperglycemia. The study aimed to assess the antihyperglycemic, hepatoprotective, and renal protective effects of ethanolic fruit extract of Moringa oleifera, on alloxan-induced diabetic mice. Four mice were assigned to each group. Alloxan was injected at the dose of 10mg/kg/body weight intraperitoneally to make the diabetic model in mice. Control and diabetic control mice received drinking water as a placebo, while the diabetic model mice group was administered with ethanolic extract of moringa fruit at the dose of 150mg/kg/bodyweight for 12 weeks. After that, animals were sacrificed, and their blood and tissue samples were collected for biochemical and histopathological examination. The glucose level markedly (p<0.0001) increased many folds in Group I (80.73± 1.24 to 221.5±13.4) and Group II (80.73 ± 1.24 from to 221.9±6.88). The level of insulin markedly (P< 0.0001) decreased in both groups (6.8±0.42 to1.378±0.19) and (6.8±0.42 to1.138±0.08) respectively. Serum hepatic and renal marker enzymes increased in the diabetic group of mice. Glucose level was meaningfully (p<0.0001) decreased in the M. oleifera administered group while serum insulin level significantly (p<0.0001) increased. The level of liver marker enzyme and renal marker also decreased as compared to the diabetic control group. Histopathological study revealed that alloxan treatment caused damage to the liver, kidney, and pancreatic tissues while the M. oleifera administered group exhibited significant improvement in the architecture of the liver, kidney, and pancreas. Hence, M. oleifera has great potential to rejuvenate the damaged tissue and consequently can restore all the serum enzymatic and hormonal parameters.

Changes of adenosine deaminase activity in serum and saliva around parturition in sows with and without postpartum dysgalactia syndrome

Background Postpartum dysgalactia syndrome (PDS) is associated with a significantly higher activation of the inflammatory and stress response at parturition than in the healthy sow. Therefore, reliable and possibly non-invasive biomarkers for substantial increases of inflammation are searched to support the PDS diagnosis. This report studies the possible changes of the inflammatory marker enzyme adenosine deaminase (ADA) in serum and saliva of 38 PDS positive sows (PDS+) and 38 healthy sows (PDS-). Sampling was performed every 24 h from 60 h before to 36 h after parturition. Isoenzyme 1 (ADA1) and isoenzyme 2 (ADA2), as well as total ADA (tADA), were measured and their statistical association with several serum and saliva biomarkers of inflammation and stress was investigated. Results Compared to a baseline (60 to 36h prepartum), salivary activities of ADA1, ADA2 and tADA increased significantly over time in both PDS+ and PDS- sows, reaching their peaks after parturition. In serum from PDS- sows, no changes were observed over time in either ADA1, ADA2 or tADA. In PDS+ sows, serum ADA2 activity decreased temporarily after parturition followed by a significant increase compared to baseline. ADA1, ADA2 and tADA were all significantly associated with several inflammatory biomarkers and ADA1 in serum was associated with serum cortisol. Although serum activity was higher in PDS+ than in PDS- sows, the differences were not statistically significant. Further, no difference was noted between the groups in the analyses of saliva. Conclusions Salivary ADA1 and ADA2 increased in all sows after parturition, potentially as a response to the postpartum inflammation. However, no difference in the activity of ADA1, ADA2 and tADA were found between PDS+ and PDS- sows indicating inability to diagnose PDS under the conditions described in this report.

Microbiological Surveillance of Biogas Plants: Targeting Acetogenic Community

Acetogens play a very important role in anaerobic digestion and are essential in ensuring process stability. Despite this, targeted studies of the acetogenic community in biogas processes remain limited. Some efforts have been made to identify and understand this community, but the lack of a reliable molecular analysis strategy makes the detection of acetogenic bacteria tedious. Recent studies suggest that screening of bacterial genetic material for formyltetrahydrofolate synthetase (FTHFS), a key marker enzyme in the Wood-Ljungdahl pathway, can give a strong indication of the presence of putative acetogens in biogas environments. In this study, we applied an acetogen-targeted analyses strategy developed previously by our research group for microbiological surveillance of commercial biogas plants. The surveillance comprised high-throughput sequencing of FTHFS gene amplicons and unsupervised data analysis with the AcetoScan pipeline. The results showed differences in the acetogenic community structure related to feed substrate and operating parameters. They also indicated that our surveillance method can be helpful in the detection of community changes before observed changes in physico-chemical profiles, and that frequent high-throughput surveillance can assist in management towards stable process operation, thus improving the economic viability of biogas plants. To our knowledge, this is the first study to apply a high-throughput microbiological surveillance approach to visualise the potential acetogenic population in commercial biogas digesters.

Evaluating the Inhibitory Effects of Dichloromethane and Methanol Extracts of Salvia macilenta and Salvia officinalis on the Diabetes Marker Enzyme, Alpha-Glucosidase: An Approach for the Treatment of Diabetes

Background: Diabetes mellitus is believed to be the most serious metabolic disease. One of the treatments for diabetes is to delay glucose uptake by inhibiting carbohydrate-hydrolyzing enzymes. Alpha-glucosidase inhibitors delay glucose uptake. Objectives: The present study was conducted aiming to evaluate the efficacy of Salvia extracts in inhibiting diabetes marker enzymes and their effects on the treatment of diabetes. Methods: This experimental study was performed in vitro. The studied plants included Salvia macilenta and Salvia officinalis. The inhibitory effects of their dichloromethane and methanol extracts were also investigated. After calculating the percentage of inhibition and IC50, Km and Vmax using GraphPad Prism 7 were also calculated. The statistical analysis was performed employing GraphPad Instat 3 software. Results: The results herein showed that the greatest inhibitory effect on alpha-glucosidase belonged to the methanol extract of S. macilenta with IC50 = 8.73 ± 0.26 mg/mL compared to that of acarbose with IC50 = 8.82 ± 0.14 mg/mL as a standard. The IC50 of dichloromethane extract of S. officinalis was 8.95 ± 0.23 mg/mL. Conclusions: The extracts had significant inhibitory effects on alpha-glucosidase. However, methanol extract of S. macilenta and dichloromethane extract of S. officinalis demonstrated the greatest inhibitory effects on alpha-glucosidase compared to acarbose as a standard.

Catechism of protective potential of Areca catechu in Isoprenaline challenged Wistar rats

Areca catechu is an important ancient drug commonly known as Supari in ayurvedic system of medicine. A lot of research work has been done on Areca catechu regarding various cardiovascular disorders such as Hypertension, Arrhythmia but no work has been done to find out its cardioprotective activity. Experimental procedures done on Wistar Albino rats as Normal control group (NC) received 0.5ml of normal saline throughout the study and served as control. Isoprenaline group (ISO) received 0.5ml of normal saline throughout the experimental phase and received Isoprenaline (85mg/kg, s.c.) on 14th and 15th day at a time lapse of 24 hours. Standard group (STD) received Metoprolol (pure) (10mg/kg/day, p.o.) for 13 days and received Isoprenaline (85mg/kg, s.c.) on 14th and 15th day. Test group received Areca catechu extract (100mg/kg/day, p.o.) and (200 mg/kg/day, p.o.) respectively for 13 days and Isoprenaline (85mg/kg, s.c.) on 14th and 15th day. On 16thday animals were sacrificed. The level of marker enzyme in serum as AST, ALT, CK, LDH, Troponin-I have shown significant decrease (P<0.001) in rats pre-treated with Areca catechu when compared to toxic group. Further, histopathological examination showed the reduction of necrosis, edema and inflammation following Areca catechu pre-treatment. Findings revealed that Areca catechu may be a potential preventive and therapeutic agent against the myocardial necrosis associated ischemic heart disease. Thus, the aqueous ethanolic Areca catechu seeds extract could be recommended as a potential cardioprotective drug.

The cryptic gonadotropin-releasing hormone neuronal system of human basal ganglia

Human reproduction is controlled by ~2,000 hypothalamic gonadotropin-releasing hormone (GnRH) neurons. Here we report the discovery and characterization of additional ~150,000-200,000 GnRH-synthesizing cells in the human basal ganglia and basal forebrain. Nearly all extrahypothalamic GnRH neurons expressed the cholinergic marker enzyme choline acetyltransferase. Similarly, hypothalamic GnRH neurons were also cholinergic both in embryonic and adult human brains. Whole-transcriptome analysis of cholinergic interneurons and medium spiny projection neurons laser-microdissected from the human putamen showed selective expression of GNRH1 and GNRHR1 autoreceptors in the cholinergic cell population and uncovered the detailed transcriptome profile and molecular connectome of these two cell types. Higher-order non-reproductive functions regulated by GnRH under physiological conditions in the human basal ganglia and basal forebrain require clarification. The role and changes of GnRH/GnRHR1 signaling in neurodegenerative disorders affecting cholinergic neurocircuitries, including Parkinson's and Alzheimer's diseases, need to be explored.

Immune Response and Bone Marker Enzyme Activities of Broiler Birds Fed Graded Level Taurine-Supplemented-Diets

Taurine, a derived amino acid has been proven to play an important biological roles in enhancing bone strength and immune activities of broiler birds. This research investigated the effect of the different concentrations of graded level dietary taurine supplement on immune response of birds against Newcastle Disease Virus (NCDV) and Infectious Bursal Disease Virus (IBDV), as well as on enzymatic markers of bone metabolism and effect on mineral composition. A total of 300 unsexed day-old arbor acre broiler birds were used for this research. The birds were randomly distributed using a completely randomized design into five dietary treatment of six replicates with 10 birds per replicate. Treatment one (T1) served as the control group with 0% taurine supplement. Treatments 2, 3, 4, and 5 contained dietary taurine supplements at 0.002, 0.004, 0.006, and 0.008%. Antibody titre against NCDV and IBDV were determined according to standard procedures. The study lasted 42 days. Birds on 0.002% taurine had the highest antibody titre (128.38) against NCDV, while birds on the 0.006% taurine-supplemented-diet also portrayed a distinct (p<0.05) titre value (1029) against IBDV. Serum alkaline phosphatase and bone specific alkaline phosphatase (132.74 and 150.66) at the 42nd day were highest (p<0.05) for birds on 0.004 and 0.002% dietary taurine supplement respectively. The activity of serum tartrate resistant acid phosphatase (44.94) was notably highest (p<0.05) for birds on 0.008% taurine. Bone mineral contents showed that birds fed with 0.002% taurinesupplemented- diet had the highest percentage (p<0.05) of phosphorous (9.50), calcium (32.18) and phosphate (21.77) composition. Conclusively, inclusion of taurine as dietary supplement has proven useful not only in enhancing the birds’ immunity against NCDV an IBDV, but also in boosting bone mineral composition of meat type poultry birds.

PHARMACODYNAMIC INTERACTION OF TINOSPORA CORDIFOLIA WITH ATENOLOL AND PROPRANOLOL IN ISOPROTERENOL INDUCED CARDIAC NECROSIS IN RATS

Objective: The present study was carried out to evaluate the combined cardioprotective effect of standardized extract of Tinospora cordifolia extract (TCE) with atenolol (AT) and propranolol (PP) in isoproterenol (ISO) induced cardiac necrosis in rats. Methods: Myocardial infarction (MI) or cardiac necrosis was induced by subcutaneous administration of ISO for two days consecutively at an interval of 24 h. Rats were pre-administered with test drugs for 21 d followed by ISO was administration on 20 and 21st day. 24 h after final ISO administration, mean arterial blood pressure (MAB), Heart rate (HR), electrocardiogram (ECG), heart bio-marker enzyme, and histopathological study of cardiac tissue were evaluated from control and experimental groups and analyzed statistically by one-way ANOVA followed by Tukeys’s test. Results: Rats administered with ISO showed significant (p<0.001) changes in ECG, HR, MAB, heart bio-marker enzyme, antioxidant parameters, and histopathology of the heart. The activities of biomarkers have reduced in serum and there is a significant (p<0.001) increase in antioxidants in the heart tissue of animals treated with drug combination. Similarly, ECG, MAB, and HR were restored to normalcy in drug-treated animals. Conclusion: It may be concluded that the herb-drug combinations i. e TCE (500 mg/kg)+AT (10 mg/kg) and TCE (500 mg/kg)+PP (10 mg/kg) has shown increased cardioprotective activity than they were used alone.