Research Journal of Biotechnology

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Vol. 13(5) May 2018

Effect of Punica granatum (Pomegranate) and Lepidium sativum (Garden Cress) on Herpes Simplex Virus Type 1 Infectivity

Zakiyah Ibrahim Y. Fallata, Ayman I. Elkady, Aly Fahmy Mohamed and Mohamed Morsi M. Ahmed

Herpes simplex virus type 1 (HSV-1) infection is a health concern worldwide, it is responsible for the orolabial lesions, ulceration and several mild and severe diseases. The current study was carried out to investigate the in vitro antiviral activity of Punica granatum, Pomegranate peel extracts (PPE) and Lepidium sativum (Garden Cress) seed extract (GCSE) on Herpes Simplex Virus Type 1. The IC50 values for PEE C4H10O and CHCl3 were 23.72 µg/ ml, 191.71 µg/ ml respectively whilst the IC50 for GCSE was 53.37 µg/ml. The antiviral activity of each extract was estimated using end point assay and infectivity titer was calculated according to Reed and Muench equation. Data recorded revealed that HSV-1 seed stock was 108 /ml, The IC50 values of tested extracts were 23.72 µg/ml, 53.37 µg/ml and 191.71 µg/ml for C4H10O, GCSE and CHCl3 respectively. The antiviral assay showed variable potentials relative to the test extract as the infectivity titer reduction arranged in the order of Diethyl ether (C4H10O) recording 1.5 log (10) /ml (21.4%), Chloroform (CHCl3) recording 0.67 log (10) /ml (9.7%) and GCSE recording 0.25 log (10) /ml (3.6 %). taken together.

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Wheat bran fermentation for the production of cellulase and xylanase by Aspergillus niger NFCCI 4113

- Bharti Amit Kumar, Kumar Amit, Kumar Alok and Dutt Dharm

Agricultural residues are the low-cost lignocellulosic materials that have potential to employ as the substrate for enzyme production under solid-state fermentation. Total eight solid substrates were examined for enzyme production; wheat bran was found to induce maximum endoglucanase (26.33 IU/gds), FPase (2.11 IU/gds) and β-glucosidase (34.30 IU/gds by Aspergillus niger NFCCI 4113). After optimization of cultural parameters, Aspergillus niger NFCCI 4113 was found to produce maximum endoglucanase (55.26 IU/gds), FPase (4.64 IU/gds), xylanase (1175.76 IU/gds) and β-glucosidase (71.30 IU/gds) at temperature (30°C), incubation time (6th day), pH (7.0) and moisture content (70%), using peptone (1.0%, w/v) as the nitrogen source and Tween-80 (0.10%, w/v) as the surfactant.

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Cognitive-Enhancing Activities of Spirulina platensis Extract from High-Pressure Homogenization Process in Scopolamine-Induced Amnesic Mice

Lee Hyeon Yong

This study demonstrates that an extract of Spirulina platensis from a high-pressure homogenization extraction process has cognitive-enhancing abilities mediated by stimulating the expression of important Neuroprotective factors, Cyclic AMP Responsive Element Binding protein (CREB) and Brain-derived Neurotrophic Factor (BDNF) in memory-deficient mice. Interestingly, the expression level of BDNF showed higher increase following the treatment with the extract, indicating that the extract could protect against neurotoxicity possibly associated with strong antioxidant activities because this extract contains 9.92 mg/g of chlorophyll a, a strong antioxidant. This study also proved that chlorophyll a has the ability to improve cognitive functions by preventing neuronal damage caused by oxidative stress. In in vivo experiments, the extract was more effective at preventing long-term memory deficiency than enhancing memory-learning ability which corresponds to the observed suppression of Acetylcholinesterase (AChE) activity and the increased expression levels of Extracellular Signal-Regulated Kinase (ERK), too. In general, the extract was found to be more effective than chlorophyll a when administered at the same concentrations possibly due to the synergistic effects of chlorophyll a and other factors in the extract. It was also interesting that there was 200 mg of the extract/kg. b.w. of saturation concentration for the maintenance of efficacy. Therefore, the results of this study could recommend the use of crude extracts in the development of functional foods for the management of memory-deficient diseases.

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Equine infectious anaemia virus dynamics and stability analysis: The role of agar gel immunodiffusion test and enzyme immunoabsorbent assay

Geethamalini S. and Balamuralitharan S.

Equine Infectious Anaemia Virus (EIAV) is a retrovirus which is transmitted to equids and can be used as an HIV model. We examined a mathematical model of EIAV infection. The disease-free state and the basic reproduction number ( ) have been figured for this framework. In this model, there are always two cases: R0 < 1 (disease-free state) and R0 > 1 (epidemic existing state). Reproductions of the framework, which demonstrate the varieties of the populace in various circumstances, have been exhibited for both these cases. We discuss a mathematical model for investigating between equilibrium and stability analysis. Additionally, we talk about the testing methods. Method 1 is the agar gel immunodiffusion test (AGID) or Coggins test. This evaluation is fairly accurate. Method 2 is the focused catalyst connected immunoadsorbent examine test (C-ELISA). This test shows a better likelihood of finding false-positive outcomes which indicates it shows a lesser degree of correction. A numerical simulation was carried out by MATLAB to illustrate the results.

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Lipid peroxidation and antioxidant 1 enzyme response to freezing stress on Lamiophlomis rotata leaves material

Zhang Y.J., Chen Y. and Guo F.X.

Electrolyte leakage, lipid peroxidation and antioxidant enzyme activity were measured in Lamiophlomis rotata (Benth) Kudo (L. rotate) leaf material subjected to low temperature treatments at -7°C and 8°C respectively throughout 84 hours experimental period. Results showed that electrolyte leakage maintained a low level for the first 54 hours after the treatment was applied before increasing and reaching a peak at the 84th hour when the L. rotate leaf material was exposed to -7°C. Malondialdehyde (MDA) content increased during the first 12 hours before decreasing and remaining at a relatively low level as the freezing treatment progressed. Superoxide dismutase (SOD), peroxidase (POD), dehydroascorbate reductase (DHAR) and ascorbate peroxidase (APX) activity were enhanced during the whole freezing process. Catalase (CAT) activity notably increased during the first six hours and fluctuated then after. GR activity was higher than the control group activity. All results suggest that L. rotate possesses a strong recovery capacity when exposed to freezing conditions. Its strong antioxidant system keeps the cell membrane intact and plays an important role in protecting the plant from freezing damage.

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Cloning and expression of human peroxiredoxin 2 and its interaction studies with curcumin

Raja N. Sella, Adithya Achuthan K.V. and Eric N.E. Manoharan

Peroxiredoxins (PRDXs) belong to the family of antioxidant proteins. Human peroxiredoxin 2 (PRDX2) is a typical 2-Cys class protein and acts as an antioxidant. It is abundant in erythrocytes and mammalian neuron and plays important role in protecting them against oxidative stress. It also has a role in enhancing the proliferative effect of cancer cells by reduction of H2O2. We successfully cloned and expressed the recombinant PRDX2 in E. coli BL21. The purification of the recombinant protein was achieved by using nickel-nitrilotriacetic acid chelating affinity chromatography. The molecular weight of the expressed protein with the His-tag was 22700 Da. This protein was used for the interactive studies with curcumin. The changes in the fluorescence of PRDX2 at the varying concentrations of curcumin were used to determine the binding constant (Kb = 3.1 × 10–4 M). Circular dichroism spectroscopic analysis indicated small perturbations in the secondary structure of the protein in the presence of curcumin. However, there was no noticeable change in the overall protein structure. This study establishes the direct binding of curcumin, a potential supplement in the cancer therapy with another antioxidant protein PRDX2.

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Isolation, identification and biological activities of endophytic fungus Chaetomium sp. from Euphorbia hirta

Uma Anitha K.P.G. and Mythili S.

The fungal endophyte was isolated from the medicinal plant E. hirta was identified as Chaetomium sp. and the gene sequence has been submitted in NCBI with an accession number MF192757. The butanolic extract, dichloromethane and the methanolic extract of Chaetomium sp. showed significant antioxidant activity using DPPH radical scavanging method, and potential hepatoprotective activity compared with the standard silymarin. Hepatoprotectivity of methanolic, dichloromethane and butanolic extracts of Chaetomium sp. was evaluated by MTT assay in HepG2 cells with CCl4 induced toxicity and subsequently evaluated for cell viability. Antioxidant activity was demonstrated by DPPH radical scavenging assay. Quantification of total phenolic content was assessed by spectroscopic methods. The hepatoprotective activity of the butanolic, dichloromethane and methanolic extract was 61.93±1.477%, 62.19±0.9333% and 70.49±1.661% of cell viability at a concentration of 150µg/mL whereas the standard silymarin showed 93.26 ± 0.784%. The antioxidant activity of butanolic, dichloromethane and methanolic extracts of Chaetomium sp. is ranging from 50.39±1.060 to 69.63±0.452 with 41.29±0.769 mgGAE/g total phenolics, 55.14±2.135 to 55.14±2.135 with the total phenolic content of 69.78±0.615 mgGAE/g and 62.95±0.756 to 83.65±1.972 with the total phenolic content of 98.31±0.905 mg GAE/g. Ascorbic acid, BHT (butylated hydroxyl toluene) Gallic acid and Pyrogallol were used as standards which showed 98.37±0.763; 97.08±0.636; 94.89±1.103 and 96.98±0.098 reducing potential respectively.

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Evaluation of cytotoxic and genotoxic effects of PEG 1500 MS in Allium cepa and modulatory action by vitamin C

Tomar Arpita, Mohapatra Rashmi and Geetanjali Swapna

PEG 1500 Mono Stearate (PEG 1500 MS) is a polyethylene glycol ester surfactant. PEG 1500 MS is used as an emulsifier, as a co-emulsifier in oil/water (O/W) emulsions, as a viscosity builder in detergent and as a lubricant in silicone-based formulations. This study was carried out to determine the cytotoxicity and genotoxicity of PEG 1500 MS in the root tip of Allium Cepa. The mitotic index was calculated and chromosomal aberrations were observed after treating the roots with three concentrations (0.4%, 0.6% and 0.8%) of PEG 1500 MS and this damage was modulated by vitamin C. In the cytotoxic assay, roots treated with 0.8% PEG 1500 showed lower mitotic indices compared to control roots. PEG 1500 MS treatment resulted in inhibition of certain cell cycle phases and various chromosomal aberrations like c-mitosis, anaphase fragmentation and stick chromosomes, for all three PEG 1500 MS concentrations. To study the modulatory effect of vitamin C in the management of mitosis in Allium cepa, root tips treated with 0.8% PEG 1500 MS were exposed to vitamin C (1mg/L, 1.5mg/L and 2mg/L) to reinitiate mitotic growth. Vitamin C significantly decreased the cytotoxic and genotoxic effects caused by 0.8% PEG 1500 MS.

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Effects of processing on residue levels of neonicotinoid insecticides in apple

Lu Hai-bo, Huang Zhi-hong, Gao Bao-jia and Wang Su-li

The effect of processing of apple can, jam, wine and vinegar on neonicotinoid residues was studied and the processing factors of all apple products and by-products were also determined using high performance liquid chromatography (HPLC). The results showed that during the processing of apple can, apple jam and apple wine, the processing factors (PF) were all less than 1.00. The PF of nitenpyram, thiamethoxam and imidacloprid in apple can were 0.36, 0.36 and 0.36 respectively; the PFs in apple can juice were 0.72, 0.80, 0.61. The residues of 3 pesticides were mainly transferred into the can juice (transfer rate all higher than 50%). During the progressing of apple jam, the PFs of 3 pesticides were 0.55, 0.63 and 0.54 respectively. During the processing of apple wine and apple vinegar, the PFs of 3 pesticides in apple wine were 0.01, 0.03 and 0.11, respectively. The PFs in apple vinegar were 0.01, 0.03 and 0.04 respectively. The results indicated that there was no processing risk during the process of apple can, apple jam, apple wine and apple vinegar.

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Synthesis of Silver Nanoparticles from Schinus Molle extract and its larvicidal activity against Aedes aegypti (L.), the vector of dengue fever in kingdom of Saudi Arabia

Ghramh Hamed A., Mahyoub Jazem A. and Alhag Sadeq K.

The aim of this study was to investigate the larvicidal potential of the ethanol leaf extracts of chinus Molle and the synthesized silver nanoparticle (AgNPs) against the fourth-life larvae of Aedes aegypti mosquito, the dengue vector in Saudi Arabia. A series of different concentrations was tested. The effective concentrations of crude extract and AgNPs ranged from 100 to 500 and 10 to 30 ppm respectively and the death percentages corresponding to these concentrations ranged from 21 to 91 and 15 to 96% respectively. According to the LC50 values of treated larvae, AgNPs (13.894 ppm) are more effective against A. aegypti mosquito larvae than the crude extract (228.345 ppm) at about 16.4 times. The mixing of the plant extract with the silver nitrate has led to potentiation. This is due to the synergy that occurs between the extract and the silver particles during the reduction process. The compounds in the extract are related to the surface of the particles, increasing the strength of their effects. It is recommended to separate the active elements in the Schinus Molle plant and its preparation in the form of nanoparticles as a promising compound in mosquito control programs.

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In-vitro immunomodulation of fish macrophages by macrophage activating factors derived from head kidney leukocytes of Labeo rohita

Awasthi Abhishek, Rathore Gaurav, Verma Dev Kumar, Sood Neeraj, Khan M.Y. and Lakra W.S.

Head kidney leucocytes of rohu, Labeo rohita, were co-stimulated with two mitogens i.e. Concanavalin A (Con A) and Phorbol myristate acetate (PMA) to trigger the release of macrophage activation factor (MAF). Effect of different concentrations of MAF on important functional properties of rohu head kidney macrophages (HKM) was studied individually and also in combination with varying concentrations of bacterial lipopolysaccharide (LPS), a known immunomodulator of macrophages. Our results revealed that co-stimulation of rohu macrophages with varying dilution of MAF (1:2 to 1:64) and different concentrations of LPS (5µg-40µg/ml) inhibited reactive oxygen species (ROS) production. Maximum production of ROS was observed with 1:8 dilution of MAF alone. Synergistic effect of MAF and LPS combination (1:4 dilution of MAF and 10 μg ml-1LPS) was seen on nitric oxide production by HKM of rohu. Similarly, lysozyme activity was also enhanced after co-stimulation with the MAF and LPS as compared to MAF or LPS alone. These data suggest that leucocytes of rohu could be stimulated to produce MAF which can be used for functional activation of HKM.

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Direct In Vitro Regeneration in Fig (Ficus carcia L.) Cv. Brown Turkey

Guranna Prabhuling and Hoolageri Huchesh

The fig (Ficus carcia L.) cv. brown turkey is one of the important exotic fruit crop originated from Asia. Its nutracutical importance, drought tolerance and remunerable economy of the crop have made faster worldwide expansion. Micropropagation of fig has provided many advantages over the conventional methods of vegetative propagation. It ensures true to type of plants, uniform quality and rapid mass production of disease free planting materials with seasonal independency. The present study was carried out with the objective to standardize efficient and reproducible protocol for direct regeneration in fig cv. “Brown Turkey”. The current season twigs were collected from healthy and vigorously growing mother plant. The nodal explants of length 4-6 cm were isolated and subjected for surface sterilization. The effective surface sterilization was achieved with HgCl2 0.10 % for 10 minute which resulted in significantly higher aseptic culture establishment (71 %) and lowest contamination (19 %) and lesser intensity of browning (10 %) on to the initiation MS medium supplemented with BAP 0.50 mg/l + activated charcoal 500 mg/l. Further the in vitro established aseptic cultures were transferred to the MS medium with various concentration and combination of BAP, NAA, Kinetin for induction of shoots. The best shoot proliferation was observed with MS medium supplemented with BAP 1 mg/l + NAA 0.10 mg/l with 3.50 shoot/explants, 4.10 cm shoot length and 5 leaves/shoot. The microshoots of length 6-8 cm were transferred on to the M.S medium supplemented with various concentrations of IBA and NAA. Best rooting was observed with MS medium supplemented with IBA 0.50 mg/l + activated charcoal 500 mg/l with 58 % rooting, 4.30 primary roots/per shoot and 7.90 cm root length. The rooted plantlets were successfully hardened under polytunnels with 100 % survival.

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Purification and characterization of β-amylase from sweet potato (Baizhengshu 2) tuberous roots

Liang Xinhong, Zhang Wanli, Wang Yuling, Sun Junliang, Zhao Ruixiang, Ran Junjian and He Hongju

Purification of β-amylase (EC from the tuberous roots of sweet potato (Baizhengshu 2) was conducted by anion exchange and gel filtration chromatography. Results showed that the enzyme is monomeric with a molecular weight of 57862.3 Da by SDS-PAGE and MALDI-TOF. The apparent Km value in the enzyme for sweet potato starch, potato starch, maize starch, soluble starch, amylose and amylopectin was 1.87±0.032, 2.16±0.031, 2.56±0.053, 1.96±0.029, 2.32±0.037 and 2.37±0.045 mg/mL respectively. According to the value of Km, Vmax and Ea, the order of the affinity between β-amylase and substrates was (1) sweet potato starch, (2) soluble starch, (3) potato starch, (4) amylose, (5) amylopectin and (6) maize starch. The optimization conditions for activation of sweet potato β-amylase were 600C, pH 5.5. The enzyme was stable ranging between a temperature of 45-650C and the pH value ranging between 5.0-7.0. The enzyme could be activated by metal ions such as Mn2+, K+, Zn2+ and Ca2+, but Mg2+ slightly reduced the activity; Cu2+, NH4+, Fe3+, Al3+, Ba2+, EDTA, Cd2+, Hg+ and Ag+ inhibited the activity. This enzyme could be important during the producing of high maltose syrup and maltodextrins from starch judging from the kinetic data and the properties indicated in this study.

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Medicinal Plants and their Phyto-Constituents with Potential Antidiabetic Activity – A Review

Krishnamoorthy Vijayalakshmi and Chinnadurai Immanuel Selvaraj

Diabetes mellitus is a metabolic disorder caused by oxidative strain or the loss of glucose homeostasis with the disorder of carbohydrate, fat and protein metabolism. It consequently affects insulin function and secretion. The growth and usage of synthetic drugs may cause side effects like mental illness or the other physiological imbalances. A new approach for treating diabetic complications is possible by using alternative medications from medicinal plants which possess many active bio-compounds. The plant sources consist of important phyto-constituents such as polyphenols, flavonoids, alkaloids and other components which acts against defective cellular metabolism and regulate its functional property. The variance in the cellular products helps to boost the body or cells against immune-complications caused by diabetes. The active compounds from the plants could be recorded and systematically validated to increase the plant-based medicines with fewer side effects or without side effects. The prolonged usage of synthetic drug can be replaced by natural form of supplements as it regulates mechanism against diabetes mellitus.

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Mitochondrial DNA D-Loop mutation in malignancy

Jayaramayya Kaavya and Santhy K.S.

Studies have shown the presence of mitochondrial (mt) deoxyribonucelic acid (mtDNA) mutations in primary cancers. mtDNA is constantly subject to reactive oxygen species and does not have any protective proteins or introns unlike nuclear DNA. Hence, mtDNA is more prone to mutations and alterations. Furthermore, wild type and mutated mitochondrial DNA can coexist in a single cell. This phenomenon is known as heteroplasmy. Mutated DNA may have a replicative advantage, resulting in the increased proliferation of cells which can lead to cancer. mtDNA consists of a non-coding, triplex structure known as the displacement loop (D-loop) region. Mutations in the D-loop region have been associated with tumours. Studies have indicated that the D-loop region plays a significant role in cancer progression. The exact role of specific D-loop mutations in different malignancies is yet to be discovered. This study provides a comprehensive summary of D-loop region mutations associated with cancer. This review offers a perspective on D-loop region mutations, their effects on tumour progression and the association of the D-loop mutations with patient prognosis.

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