Synthesis and analysis of silica nanocarriers for pectinase immobilization: Enhancing enzymatic stability for continuous industrial applications.

Pectinolytic enzymes are among the important group of industrial enzymes that have wide applications in different food industries. In this study, pectinase-based silica nanocarriers were synthesized using co-precipitation and cross-linking techniques. The resulting silica nanoparticles were investigated using scanning electron microscopy (SEM), energy-dispersive electron microscopy (EDEX), and X-ray diffraction (XRD) for determination of its morphology, elemental composition, and crystalline pattern. Under the optimal immobilization conditions like 1.5 % glutaraldehyde, 3000 IU/mg pectinase concentration, 90 min immobilization time and 40 °C immobilization temperature, pectinase showed maximum immobilization yield. The immobilization of pectinase onto the silica nanocarriers led to enhanced catalytic characteristics, displaying higher enzymatic activity across various temperature and pH levels compared to soluble pectinase. Moreover, the immobilization substantially improved the temperature stability of pectinase, exhibiting 100 % of its initial activity even after 120 h of pre-incubation at 50 °C. Additionally, the silica nanocarrier pectinase retained 100 % of its original activity even after being reused 10 times in a single batch of reactions. These findings indicate that the immobilization of silica nanocarriers effectively enhances pectinase's industrial capabilities, making it economically feasible for industrial use and an efficient system for various biotechnological applications.

Salt-Induced Modulation of Ion Transport and PSII Photoprotection Determine the Salinity Tolerance of Amphidiploid Brassicas.

Brassica species show varying levels of resistance to salt stress. To understand the genetics underlying these differential stress tolerance patterns in Brassicas, we exposed two widely cultivated amphidiploid Brassica species having different genomes, Brassica juncea (AABB, n = 18) and Brassica napus (AACC, n = 19), to elevated levels of NaCl concentration (300 mM, half the salinity of seawater). B. juncea produced more biomass, an increased chlorophyll content, and fewer accumulated sodium (Na+) and chloride (Cl-) ions in its photosynthesizing tissues. Chlorophyll fluorescence assays revealed that the reaction centers of PSII of B. juncea were more photoprotected and hence more active than those of B. napus under NaCl stress, which, in turn, resulted in a better PSII quantum efficiency, better utilization of photochemical energy with significantly reduced energy loss, and higher electron transport rates, even under stressful conditions. The expression of key genes responsible for salt tolerance (NHX1 and AVP1, which are nuclear-encoded) and photosynthesis (psbA, psaA, petB, and rbcL, which are chloroplast-encoded) were monitored for their genetic differences underlying stress tolerance. Under NaCl stress, the expression of NHX1, D1, and Rubisco increased several folds in B. juncea plants compared to B. napus, highlighting differences in genetics between these two Brassicas. The higher photosynthetic potential under stress suggests that B. juncea is a promising candidate for genetic modifications and its cultivation on marginal lands.

Development of Pectinase Based Nanocatalyst by Immobilization of Pectinase on Magnetic Iron Oxide Nanoparticles Using Glutaraldehyde as Crosslinking Agent.

To increase its operational stability and ongoing reusability, B. subtilis pectinase was immobilized on iron oxide nanocarrier. Through co-precipitation, magnetic iron oxide nanoparticles were synthesized. Scanning electron microscopy (SEM) and energy dispersive electron microscopy (EDEX) were used to analyze the nanoparticles. Pectinase was immobilized using glutaraldehyde as a crosslinking agent on iron oxide nanocarrier. In comparison to free pectinase, immobilized pectinase demonstrated higher enzymatic activity at a variety of temperatures and pH levels. Immobilization also boosted pectinase's catalytic stability. After 120 h of pre-incubation at 50 °C, immobilized pectinase maintained more than 90% of its initial activity due to the iron oxide nanocarrier, which improved the thermal stability of pectinase at various temperatures. Following 15 repetitions of enzymatic reactions, immobilized pectinase still exhibited 90% of its initial activity. According to the results, pectinase's catalytic capabilities were enhanced by its immobilization on iron oxide nanocarrier, making it economically suitable for industrial use.

Hepatoprotective effect of methanolic extract of Iris florentina L. on paracetamol-induced liver toxicity in rats.

Despite of plethora of research on hepatoprotective potential of medicinal plants, there is still need to discover potential plants with hepatoprotective activity. Iris florentina L. is a medicinal plant with traditional claims but ignored investigation regarding its hepatoprotective effects. The current study is aimed to investigate the hepatoprotective potential of I. florentina L. methanolic extract on paracetamol (PCM)-induced liver injury. The phytochemical and HPLC screening was done which showed the presence of potential constituents including flavonoids and phenols. For investigating the hepatoprotective effect of I. florentina L. methanolic extract, rats were given five different treatments for seven consecutive days. The normal control (group 1) was administered with normal saline, group 2 (Diseased) received paracetamol and group 3 (Standard) was given silymarin as reference drug. In group 4 and 5 (Treated), I. florentina L. methanolic extract (250 and 500mg/kg) were administered. Different serum biomarkers and histopathological studies were performed to assess the recovery caused by PCM in comparison to diseased group. The treatment of I. florentina methanolic extract significantly improve the serum biomarkers and restored the hepatic injury towards normal, indicating the hepatoprotective potential. Thus, we can conclude that I. florentina have significantly reversed the damage caused by paracetamol in hepatotoxic rat model due to their potential phytochemical constituents.

Effects of pH and Ionic Salts on the Emulsifying and Rheological Properties of Acorn Protein Isolate.

This study was designed to evaluate the emulsifying and rheological properties of acorn protein isolate (API) in different pH mediums (pH 3, 7 and 9) and in the presence of ionic salts (1 M NaCl and 1 M CaCl2). API shows higher solubility in distilled water at pH 7, while at the same pH, a decrease in solubility was observed for API in the presence of CaCl2 (61.30%). A lower emulsifying activity index (EAI), lower stability index (ESI), larger droplet sizes and slight flocculation were observed for API in the presence of salts at different pHs. Importantly, CaCl2 treated samples showed relevantly higher EAI (252.67 m2/g) and ESI (152.67 min) values at all pH as compared to NaCl (221.76 m2/g), (111.82 min), respectively. A significant increase in interfacial protein concentration (4.61 mg/m2) was observed for emulsion at pH 9 with CaCl2, while the major fractions of API were observed in an interfacial layer after SDS-PAGE analysis. All of the emulsion shows shear thinning behavior (τc > 0 and n < 1), while the highest viscosity was observed for emulsion prepared with CaCl2 at pH 3 (11.03 ± 1.62). In conclusion, API, in the presence of ionic salts at acidic, neutral and basic pH, can produce natural emulsions, which could be substitutes for synthetic surfactants for such formulations.

Unveiling the catalytic ability of carbonaceous materials in Fenton-like reaction by controlled-release CaO2 nanoparticles for trichloroethylene degradation.

Carbonaceous materials (CMs) have been applied extensively for enhancing the catalytic performance of environmental catalysts, however, the self-catalytic mechanism of CMs for groundwater remediation is rarely investigated. Herein, we unveiled the catalytic ability of various CMs via Fe(III) reduction through polyvinyl alcohol-coated calcium peroxide nanoparticles (PVA@nCP) for trichloroethylene (TCE) removal. Among selected CMs (graphite (G), biochar (BC) and activated carbon (AC)), BC and AC showed enhancement of TCE removal of 89% and 98% via both adsorption and catalytic degradation. BET and SEM analyses showed a higher adsorption capacity of AC (27.8%) than others. The generation of solution-Fe(II) and surface-Fe(II) revealed the reduction of Fe(III) on CMs-surface. The role of O-containing groups was investigated by the FTIR technique and XPS quantified the 52% and 57% surface-Fe(II) in BC and AC systems, respectively. EPR and quenching tests confirmed that both solution and surface-bound species (HO•, O2-• and 1O2) contributed to TCE degradation. Acidic pH condition encouraged TCE removal and the presence of HCO3- negatively affected TCE removal than other inorganic ions. Both schemes (PVA@nCP/Fe(III)/BC and PVA@nCP/Fe(III)/AC) exhibited promising results in the actual groundwater, surfactant-amended solution, and removal of other chlorinated-pollutants, opening a new direction towards green environmental remediation for prolonged benefits.

Thermodynamics, kinetics and optimization of catalytic behavior of polyacrylamide-entrapped carboxymethyl cellulase (CMCase) for prospective industrial use.

In the current study, kinetic and thermodynamic parameters of free and polyacrylamide-immobilized CMCase were analyzed. The maximum immobilization yield of 34 ± 1.7% was achieved at 11% acrylamide. The enthalpy of activation (ΔH) of free and immobilized enzyme was found to be 13.61 and 0.29 kJ mol-1, respectively. Irreversible inactivation energy of free and immobilized CMCase was 96.43 and 99.01 kJ mol-1, respectively. Similarly, the enthalpy of deactivation (ΔHd) values for free and immobilized enzyme were found to be in the range of 93.51-93.76 kJ mol-1 and 96.08-96.33 kJ mol-1, respectively. Michaelis-Menten constant (Km) increased from 1.267 ± 0.06 to 1.5891 ± 0.07 mg ml-1 and the maximum reaction rate (Vmax) value decreased (8319.47 ± 416 to 5643.34 ± 282 U ml-1 min-1) after immobilization. Due to wide pH and temperature stability profile with sufficient reusing efficiency up to three successive cycles, the immobilized CMCase might be useful for various industrial processes.

Plant growth regulators and EDTA improve phytoremediation potential and antioxidant response of Dysphania ambrosioides (L.) Mosyakin & Clemants in a Cd-spiked soil.

Soil pollution due to potentially toxic elements is a worldwide challenge for health and food security. Chelate-assisted phytoextraction along with the application of plant growth regulators (PGRs) could increase the phytoremediation efficiency of metal-contaminated soils. The present study was conducted to investigate the effect of different PGRs [Gibberellic acid (GA3) and indole acetic acid (IAA)] and synthetic chelator (EDTA) on growth parameters and Cd phytoextraction potential of Dysphania ambrosioides (L.) Mosyakin & Clemants grown under Cd-spiked soil. GA3 (10-7 M) and IAA (10-5 M) were applied four times with an interval of 10 days through a foliar spray, while EDTA (40 mg kg-1 soil) was once added to the soil. The results showed that Cd stress significantly decreased fresh biomass, dry biomass, total water contents, and photosynthetic pigments as compared to control. Application of PGRs significantly enhanced plant growth and Cd phytoextraction. The combined application of GA3 and IAA with EDTA significantly increased Cd accumulation (6.72 mg pot-1 dry biomass) and bioconcentration factor (15.21) as compared to C1 (Cd only). The same treatment significantly increased chlorophyll, proline, phenolic contents, and antioxidant activities (CAT, SOD, and POD) while MDA contents were reduced. In roots, Cd accumulation showed a statistically significant and positive correlation with proline, phenolics, fresh biomass, and dry biomass. Similarly, Cd accumulation showed a positive correlation with antioxidant enzyme activities in leaves. D. ambrosioides showed hyperaccumulation potential for Cd, based on bioconcentration factor (BCF) > 1. In conclusion, exogenous application of GA3 and IAA reduces Cd stress while EDTA application enhances Cd phytoextraction and ultimately the phytoremediation potential of D. ambrosioides.

Preparation, characterization and stability studies of cross-linked α-amylase aggregates (CLAAs) for continuous liquefaction of starch.

In current study, α-amylase of fungal origin was immobilized using cross-linking strategy. The influence of precipitant (ammonium sulphate) and cross-linker (glutaraldehyde) concentration revealed that 60% (w/v) precipitant and 1.5% (v/v) cross-linker saturation was required to attain optimum activity. Cross-linked amylase aggregates (CLAAs) were characterized and 10-degree shift in optimum temperature (soluble enzyme: 50 °C; cross-linked: 60 °C) and 1-unit shift in pH (soluble enzyme: pH -6; cross-linked: pH -7) was observed after immobilization. The Vmax for soluble α-amylase and its cross-linked form was 1225 U ml-1 and 3629 U ml-1, respectively. The CLAAs was more thermostable than its soluble form and retained its 30% activity even after 60 min of incubation at 70 °C. Moreover, cross-linked amylase retained its activity after two months while its soluble counterpart lost its complete activity after 10 and 20 days at 30 °C and 4 °C storage, respectively. Reusability test showed that cross-linked amylase could retain 13% of its residual activity after 10 repeated cycles. Therefore, 10 times more glucose was produced after cross-linking than soluble amylase when it was utilized multiple times. This study indicates that amylase aggregates are highly effective for continuous liquefaction of starch, hence have strong potential to be used for different industrial processes.

Encapsulation of pectinase within polyacrylamide gel: characterization of its catalytic properties for continuous industrial uses.

Pectinase as a biocatalyst play a significant role in food and textile industries. In this study, the pectinase was immobilized by encapsulation within polyacrylamide gel to enhance its catalytic properties and ensure the reusability for continuous industrial processes. 9.5% acrylamide and 0.5% N, N'- methylenebisacrylamide concentration gave high percentage of pectinase immobilization yield within gel. The catalytic properties of immobilized pectinase was determined with comparison of soluble pectinase. The immobilization of pectinase within polyacrylamide gel didn't effect catalytic properties of pectinase and both the free and immobilized pectinase showed maximum pectinolytic activity at 45 °C and pH 10. The Michaelis-Menten kinetic behavior of pectinase was slightly changed after immobilization and immobilized pectinase showed somewhat higher Km and lower Vmax value as compared to soluble pectinase. Polyacrylamide gel encapsulation enhanced the thermal stability of pectinase and encapsulated pectinase showed higher thermal stability against various temperature ranging from ranging from 30 °C to 50 °C as compared free pectinase. Furthermore, the surface topography of polyacrylamide gel was analyzed using scanning electron microscopy and it was observed that the surface topography of polyacrylamide gel was changed after encapsulation. The encapsulation of pectinase within polyacrylamide gel enhanced the possibility of reutilization of pectinase in various industries and pectinase retained more than 50% of its initial activity even after seven batch of reactions.

Utilization of different polymers for the improvement of catalytic properties and recycling efficiency of bacterial maltase.

Current study deals with the comparative study related to immobilization of maltase using synthetic (polyacrylamide) and non-synthetic (calcium alginate, agar-agar and agarose) polymers via entrapment technique. Polyacrylamide beads were formed by cross-linking of monomers, agar-agar and agarose through solidification while alginate beads were prepared by simple gelation. Results showed that the efficiency of enzyme significantly improved after immobilization and among all tested supports agar-agar was found to be the most promising and biocompatible for maltase in terms of immobilization yield (82.77%). The catalytic behavior of maltase was slightly shifted in terms of reaction time (free enzyme, agarose and polyacrylamide: 5.0 min; agar-agar and alginate: 10.0 min), pH (free enzyme, alginate and polyacrylamide: 6.5; agar-agar, agarose: 7.0) and temperature (free enzyme: 45 °C; alginate: 50 °C; polyacrylamide: 55 °C; agarose: 60 °C; agar-agar: 65 °C). Stability profile of immobilized maltase also revealed that all the supports utilized have significantly enhanced the activity of maltase at higher temperatures then its free counterpart. However, recycling data showed that agar-agar entrapped maltase retained 20.0% of its initial activity even after 10 cycles followed by agarose (10.0%) while polyacrylamide and alginate showed no activity after 8 and 6 cycles respectively.

High Selectivity to Aromatics by a Mg and Na Co-modified Catalyst in Direct Conversion of Syngas.

The demand for aromatics, especially benzene, toluene, and xylene, has been increased in recent years as the crucial feedstocks of coatings and pharmaceutical industry. In this work, a modified Fischer-Tropsch synthesis (FTS) catalyst FeNaMg was fabricated via a sol-precipitation method and integrated with an HZSM-5 aromatization catalyst for the aromatics synthesis from syngas by a one-step process. Syngas was first converted to lower olefins as intermediates on the active component of the FeNaMg catalyst followed by aromatization on zeolite. Different characterization approaches, such as BET, XRD, XPS, hydrogen temperature-programmed reduction, temperature-programmed desorption of CO, TG, and SEM, revealed that Mg efficiently optimized physicochemical properties of the Fe-based catalyst by generating a MgFe2O4 spinel structure. Further investigation demonstrated that the MgFe2O4 spinel structure could increase the syngas adsorption area, facilitating the reduction and carburization of the Fe phase, while Mg decreased CO2 selectivity (31.26 to21%) by restraining the water-gas shift reaction and improved the utilization efficiency of carbon. At the same time, alkali metal Na changed the surface electronic environment of the FTS catalyst to enhance CO adsorption as an electronic promoter, which suppressed methane formation by restraining over hydrogenation. Therefore, the synergism that existed between Mg and Na during the reaction escalated the CO conversion and aromatics selectivity to 96.19 and 51.38%, respectively.

Biochemical spectrum of parathyroid disorders diagnosed at a tertiary care setting.

OBJECTIVE: To determine the clinical and biochemical pattern of parathyroid disorders in a tertiary care setting.. METHODS: The cross-sectional study was conducted at the Armed Forces Institute of Pathology, Rawalpindi, Pakistan, from September 2017 to February 2018, and comprised patients with suspected parathyroid disorders. A panel of biochemical tests were used for diagnosis of parathyroid disorders, which included parathyroid hormone levels, total calcium, ionized calcium, inorganic phosphorus, alkaline phosphatase, magnesium, total vitamin D and urinary calcium-to-creatinine ratio. SPSS 24 was used for data analysis. RESULTS: Of the 384 subjects, 248(65%) were male and 136(35%) were female. Overall mean age was 48±19years. Of the total, 302(786%) had parathyroid issues, with 244(81%) having secondary hyperparathyroidism. Mean serum total calcium, phosphorus, ionized calcium, magnesium and total vitamin D were 8.98±1.52 mg/dl, 4.0±1.30 mg/dl, 4.65±0.52 mg/dl, 2.11±0.27 mg/dl and 20.5±8.52 ngml respectively. Of the patients diagnosed with secondary hyperparathyroidism, 72.2% patients had chronic kidney disease and 20.2% had isolated vitamin D deficiency. CONCLUSIONS: Parathyroid disorders had significant impact on bone health. Moreover, secondary hyperparathyroidism was seen to be emerging as a major endocrine problem, especially in chronic kidney disease patients and vitamin D-deficient individuals.

Diagnostic accuracy of cannabinoid testing by liquid chromatography-tandem mass spectrometry in human hair.

OBJECTIVE: To determine the diagnostic accuracy of Cannabinoids testing by LC-MS/MS in human hair and compare it with urine in civil heavy vehicle drivers. METHODS: Current study was a diagnostic accuracy study done in "Armed Forces Institute of Pathology Rawalpindi, Pakistan" from February to November 2017. Urine and hair samples were collected by non-probability convenient sampling technique from 151 heavy vehicle drivers from Punjab. Hair and urine samples were collected from each subject. Separation of compounds was performed on Agilent Poroshell and analyzed using 6460 Triple Quadrapole LC-MS along-with software Mass hunter ©. RESULTS: Study population (151 civil heavy vehicle drivers) was divided into three main divisions There were 69 (46%) truck drivers,43 (28.5%) twenty-wheeler drivers and 39 (26%) bus drivers. Mean age of study participants was 36±10.82 years. Paired t-test was applied to check mean difference between the two tests' concentration (i.e urine and hair analysis for cannabis) which showed significant difference at p<0.001. Among the different factors of diagnostic accuracy in hair and urine specimens were: Sensitivity (96% and 62%), Specificity (93% and 95%) Positive Predictive Value (88% and 87%), Negative Predictive Value (97% and 82%) respectively. Overall diagnostic accuracy of Cannabinoids detection in hair was 94% while in urine it was 83%. ROC curve showed area under curve of 0.79 and 0.96 for urine and hair samples respectively. CONCLUSIONS: Current study signified hair as a substitute matrix owing to its non-invasive specimen collection, better diagnostic yield and wider detection period compared to urine.

Isolation and characterization of bacteriophage to control multidrug-resistant Pseudomonas aeruginosa planktonic cells and biofilm.

Pseudomonas aeruginosa is Gram-negative bacterium, one of the leading cause of drug-resistant nosocomial infections in developing countries. This bacterium possesses chromosomally encoded efflux pumps, poor permeability of outer-membrane and high tendency for biofilm formation which are tools to confer resistance. Bacteriophages are regarded as feasible treatment option for control of resistant P. aeruginosa. The aim of the current study was isolate and characterized a bacteriophage against P. aeruginosa with MDR and biofilm ability. A bacteriophage MA-1 with moderate host range was isolated from waste water. The phage was considerable heat and pH stable. Electron microscopy revealed that phage MA-1 belongs to Myoviridae family. Its genome was dsDNA (≈50 kb), coding for eighteen different proteins (ranging from 12 to 250 KDa). P. aeruginosa-2949 log growth phase was significantly reduced by phage MA-1 (2.5 × 103 CFU/ml) as compared to control (without phage). Phage MA-1 also showed significant reductions of 2.0, 2.5 and 3.2 folds in 24, 48, and 74 h old biofilms after 6 h treatment with phage respectively as compared to control. It was concluded from this study that phage MA-1 has capability of killing P. aeruginosa planktonic cells and biofilm, but for complete eradication cocktail will more effective to avoid resistance.

Investigating the In Vitro Regeneration Potential of Commercial Cultivars of Brassica.

In vitro regeneration is a pre-requisite for developing transgenic plants through tissue culture-based genetic engineering approaches. Huge variations among different genotypes of the genus Brassica necessitate the identification of a set of regeneration conditions for a genotype, which can be reliably used in transformation experiments. In this study, we evaluated the morphogenesis potential of four commercial cultivars (Faisal canola, Punjab canola, Aari canola, Nifa Gold) and one model, Westar, from four different explants namely cotyledons, hypocotyls, petioles and roots on three different Brassica regeneration protocols, BRP-I, -II and -III. The regeneration efficiency was observed in the range of 6-73%, 4-79.3%, 0-50.6%, and 0-42.6% from cotyledons, petioles, hypocotyls and roots, respectively, whereas, the regeneration response in terms of average shoots per explant was found to be 0.76-10.9, 0.2-3.2, 0-3.4 and 0-2.7 from these explants. Of the commercial varieties tested, almost all varieties showed poorer regeneration than Westar except Aari canola. In comparison to Westar, its regeneration frequency from cotyledons was up to 7.5-fold higher on BRP-I, while it produced up to 21.9-fold more shoots per explant. Our data show that the explant has strong influence on the regeneration response, ranging from 24% to 92%. While the growth of commercial cultivars was least affected by the regeneration conditions provided, the effect on Westar was twice that of the commercial cultivars. After determining the optimal explant type and regeneration conditions, we also determined the minimum kanamycin concentration levels required to selectively inhibit the growth of untransformed cells for these cultivars. Regenerated shoots of Aari canola could be successfully grown to maturity within 16-18 weeks, with no altered phenotype noted and normal seed yields obtained. Therefore, the commercial variety, Aari canola, could be a good candidate for future genetic transformation studies.

Maltose deterioration approach: Catalytic behavior optimization and stability profile of maltase from Bacillus licheniformis KIBGE-IB4.

Maltase is an economically valuable enzyme that is used to catalyze the hydrolytic process of maltose and yields d-glucose as a product. In this study, the catalytic behavior of maltase was optimized under various physicochemical condition. Results indicated that bacterial maltase exhibited maximum catalytic activity at 45 °C and pH-6.5 after 5.0 min. It presented greater stability within 0.1 M K2HPO4 buffer having pH-6.5 and showed 100 % activity even after 1.0 h. It retained 83.6 % and 45.0 % activity at 40 °C after 1.0 and 3.0 h, respectively. The enzyme retained 90.0 % activity at -20 °C even after 60 days. The molecular weight of enzyme was deduced to be 157.2 kDa as calculated using polyacrylamide gel electrophoresis (PAGE) and zymography. It was concluded that the characterized maltase has notable stability profile with reference to temperature, pH and other reaction conditions which anticipates its utilization in various starch and maltose hydrolyzing processes for the synthesis of glucose.

Comparative evaluation of 30 and 60 minutes cortisol levels during short synacthen test for diagnosis of adrenal insufficiency.

OBJECTIVE: To assess and compare diagnostic value of 30-minute cortisol level over 60-minute level in the diagnosis of adrenal insufficiency. METHODS: The comparative cross-sectional study was conducted at the Armed Forces Institute of Pathology, Rawalpindi, Pakistan, from August 2017 to May 2018, and comprised patients referred to the facility for short synacthen test with suspicion of adrenal insufficiency. Blood samples for serum cortisol were taken at time-0 and then 30 and 60 minutes after the adreno-cortico-tropic hormone injection. Total serum cortisol was measured. Adrenal insufficiency was defined as stimulated cortisol level <500 nmol/l at 30 and 60 minutes post-stimulation. SPSS 24 was used for data analysis. RESULTS: Of the 111 subjects, 56(50.4%) were males and 55(49.5%) were females. Overall mean age was 34±20 years. Mean basal serum cortisol level was 110±98 nmol/l in patients with adrenal insufficiency and it was 294±164 nmol/l in patients with intact adrenal functions. Cortisol level at both 30 and 60 minutes was significant (p<0.001). Receiver Operating Characteristics curve was plotted which showed area under curve of 0.83 and 0.82 for 60 and 30 minutes respectively. CONCLUSIONS: The 30-minute cortisol level post-stimulation carried no diagnostic value . Measuring cortisol level once at 60-minute post-stimulation would be of more value apart from being cost-effective in the diagnosis of adrenal insufficiency.

Improvement of catalytic properties of starch hydrolyzing fungal amyloglucosidase: Utilization of agar-agar as an organic matrix for immobilization.

In this study, amyloglucosidase was immobilized within agar-agar through entrapment technique for the hydrolysis of soluble starch. Enzymatic activities of soluble and entrapped amyloglucosidase were compared using soluble starch as a substrate. Partially purified enzyme was immobilized and maximum immobilization yield (80%) was attained at 40 gL-1 of agar-agar. Enzyme catalysis reaction time shifted from 5.0 min to 10 min after immobilization. Similarly, a five-degree shift in temperature (60 °C-65 °C) and a 0.5 unit increase in pH (pH-5.0 to pH-5.5) were also observed. Substrate saturation kinetics revealed that Km of entrapped amyloglucosidase increased from 1.41 mg ml-1 (soluble enzyme) to 3.39 mg ml-1 (immobilized enzyme) whereas, Vmax decreased from 947 kU mg-1 (soluble enzyme) to 698 kU mg-1 (immobilized enzyme). Entrapped amyloglucosidase also exhibited significant catalytic performance during thermal and storage stability when compared with soluble enzyme. Reusability of entrapped amyloglucosidase for hydrolysis of soluble starch demonstrated its recycling efficiency up to six cycles which is an exceptional characteristic for continuous bioprocessing of soluble starch into glucose.

Xylan deterioration approach: Purification and catalytic behavior optimization of a novel ß-1,4-d-xylanohydrolase from Geobacillus stearothermophilus KIBGE-IB29.

The ß-1,4-d-xylanohydrolase is an industry valuable catalytic protein and used to synthesize xylooligosaccharides and xylose. In the current study, ß-1,4-d-xylanohydrolase from Geobacillus stearothermophilus KIBGE-IB29 was partially purified up to 9.5-fold with a recovery yield of 52%. It exhibited optimal catalytic activity at pH-7.0 and 50 °C within 5 min. Almost 50% activity retained at pH-4.0 to 9.0 however, 70% activity observed within the range of 40 °C to 70 °C. The ß-1,4-d-xylanohydrolase showed a significant hydrolytic pattern with 48.7 kDa molecular mass. It was found that the enzymatic activity improved up to 160% with 1.0 mM ethanol. Moreover, the activity of enzyme drastically increased up to 2.3 and 1.5 fold when incubated with Tween 80 and Triton X-100 (1.0 mM), respectively. The ß-1,4-d-xylanohydrolase also retained 72% activity at -80 °C after 180 days. Such a remarkable biochemical properties of ß-1,4-d-xylanohydrolase make it possible to forecast its potential use in textile and food industries.