The optimization of enzymatic oxidation of levofloxacin, a fluoroquinolone antibiotic for wastetwater treatment.

The global presence of antibiotics in the environment has created concerns about the emergence of antibiotic resistance bacteria and potential hazard to humans and the ecosystem. This work aims to study the removal of levofloxacin, a new generation fluoroquinolone antibiotic from aqueous solutions by enzyme mediated oxidation process and optimization of the conditions thereof by response surface methodology (RSM) using Box-Behnken design (BBD). For this study, experiments were conducted to analyze the effect of independent variables namely, pH, temperature, mediator concentration and antibiotic concentration on the degradation percentage of levofloxacin antibiotic using laccase enzyme derived from Trametes versicolor. The residual levofloxacin concentration was determined using high performance liquid chromatography (HPLC). On applying the quadratic regression analysis, among the main parameters, it was found that the percentage degradation was significantly affected by all the four variables. The predicted values for percentage degradation of levofloxacin were close to the experimental values obtained and the R2 (0.95) indicated that the regression was able to give a good prediction of response for the percentage degradation of levofloxacin in the studied range. The optimal conditions for the maximum degradation (99.9%) as predicted by the BBD were: temperature of 37 °C, pH of 4.5, mediator concentration of 0.1 mM and levofloxacin concentration of 5 µg mL-1. The findings of the study were further extended to study the effect of partially purified enzymes isolated from Pleurotus eryngii, Pleurotus florida and Pleurotus sajor caju on the degradation of levofloxacin at concentrations ranging from as low as 0.1 to as high as 50 µg mL-1 in synthetic wastewater utilizing the optimized conditions generated by BBD. A maximum degradation of 88.8% was achieved with the partially purified enzyme isolated from Pleurotus eryngii at 1 µg mL-1 levofloxacin concentration which was at par with the commercial laccase which showed 89% degradation in synthetic wastewater at the optimized conditions. The biodegradation studies were conducted using only 2 units of laccase. Thus, the expensive commercial laccase can be effectively replaced by crude laccase isolated from indigenous macrofungi such as P. eryngii, P. florida and P. sajor caju as a cost effective alternative to degrade levofloxacin present in contaminated wastewater using as low as 2 units of enzyme for a 72 h treatment period.

Laccase mediated transformation of fluoroquinolone antibiotics: Analyzing degradation pathways and assessing algal toxicity.

Improper waste disposal or inadequate wastewater treatment can result in pharmaceuticals reaching water bodies, posing environmental hazards. In this study, crude extracts containing the laccase enzyme from Pleurotus florida, Pleurotus eryngii, and Pleurotus sajor caju were used to degrade the fluoroquinolone antibiotics (FQs) levofloxacin (LEV), norfloxacin (NOR), ciprofloxacin (CIP), ofloxacin (OFL), and enrofloxacin (ENR) in aqueous solutions. The results for the fungi derived laccase extracts were compared with those obtained using commercially sourced laccase. Proteomics analysis of the crude extracts confirmed the presence of laccase enzyme across all three tested species, with proteins matching those found in Trametes versicolor and Pleurotus ostreatus. In vivo studies were conducted using species pure lines of fungal whole cells. The highest degradation efficiency observed was 77.7% for LEV in the presence of P. sajor caju after 25 days of treatment. Degradation efficiencies ranged from approximately 60-72% for P. florida, 45-76% for P. eryngii, and 47-78% for P. sajor caju. A series of in vitro experiments were also conducted using crude extracts from the three species and outcomes compared with those obtained when commercial laccase was used confirmed laccase as the enzyme responsible for antibiotic removal. The degradation efficiencies in vitro surpassed those measured in vivo, ranging from approximately 91-98% for commercial laccase, 77-92% for P. florida, 76-92% for P. eryngii, and 78-88% for P. sajor caju. Liquid chromatography-high-resolution mass spectrometry (LC-MS/MS) identified the degradation products, indicating a consistent enzymatic degradation pathway targeting the piperazine moiety common to all tested FQs, irrespective of the initial antibiotic structure. Phytoplankton toxicity studies with Dunaliella tertiolecta were performed to aid in understanding the impact of emerging contaminants on ecosystems, and by-products were analysed for ecotoxicity to assess treatment efficacy. Laccase-mediated enzymatic oxidation shows promising results in reducing algal toxicity, notably with Pleurotus eryngii extract achieving a 97.7% decrease for CIP and a 90% decrease for LEV. These findings suggest the potential of these naturally sourced extracts in mitigating antibiotic contamination in aquatic ecosystems.

Plasmonic semi shells derived from simultaneous in situ gold growth and anisotropic acid etching of ZIF-8 for photothermal ablation of metastatic breast tumor.

Open nanoshells such as nanobowls or nanocups collectively described as 'semi shells' have unique plasmonic properties due to their lack of symmetry. So far, their fabrication was based on multistep and laborious methods such as solid state sputter coating or selective deposition/etching using sacrificial templates. In this work, we report a rapid one step colloidal synthetic protocol for PEGylated semi-shell (SS) fabrication by simultaneous facet specific anisotropic chemical etching of rhombic dodecahedral ZIF-8 and heterogenous nucleation & growth of gold. The SS possesses a strong localized surface plasmon resonance in the near-infrared region, which is retained after surface passivation with polyethylene glycol and subsequent cryopreservation for extended shelf-life. Freshly reconstituted PEGylated SS was found to be safe & non-toxic in healthy C57BL/6 mice post intravenous administration. The PEGylated SS displayed significant photothermal efficiency of ~37% with 808 nm laser irradiation. Preclinical assessment of intra-tumoral photothermal efficacy indicated complete remission of primary breast tumor mass with insignificant metastasis to vital organs in 4T1 FL2 tumor bearing CD1 nude mice. Further, PEGylated SS mediated photothermal therapy also yielded morbidity free survivael of 75% for up to 90 days, indicating their potential to significantly improve outcomes in advanced breast tumors.

Profile of micro-organisms in intensive care unit of a level-1 trauma centre: A retrospective study.

BACKGROUND: Patients with trauma usually require highly specialized care in ICU and many times their recovery is complicated by infections. Aim of this study was to determine the profile of pathogens and their impact on outcome among these patients. MATERIALS AND METHODS: The clinical records of 101 consecutive patients who were admitted for more than 48-hrs in ICU during Jun-Dec 2007 were analyzed. RESULTS: Total of 953 samples from blood, urine, BAL or pus/collection were subjected to cultures. From 276 samples, 299 organisms were isolated. Among pathogens Candida Spp. [89 (29%)] were the most common, followed by Acinetobacter Spp. [69 (23%)], Pseudomonas Spp. [63 (21%)], Klebsiella Spp. [31 (10%)], coagulase negative Staphylococcus aureus [16 (5%)], E coli [12 (4%)], Enterobacter Spp. [7 (2%)], S aureus [6 (2%)], Enterococcus Spp. [5 (2%)], Citrobacter Spp. [2 (0.6%)], S maltophila [1 (0.3%)] and Providentia Spp. [1 (0.3%)]. For gram negative pathogens drug-resistance rates were as follows: Fluoroquinolones, 76%; 3(rd) generation cephalosporins, 74%; aminoglycosides, 66%; ß-lactams/ß-lactamase inhibitors combinations, 64%; and carbapenems, 50%. Among these 27% of pathogens were resistant to all 5 classes of drugs. 58% of Staphylococcus aureus were methicillin-resistant whereas 85% of coagulase negative Staphylococcus were methicillin resistant. The mortality was higher among patients in whom pathogens were isolated [Odd's Ratio (OR) 0.185; 95% confidence interval (CI) 0.049-0.640; P = 0.002]. CONCLUSIONS: Isolation of multi-drug resistant pathogens is common among trauma patients admitted in ICU and is associated with increased mortality and could impact on the consumption of hospital resources. The importance of high rate of fungal isolation needs to be studied among these patients.

Serum Vascular Endothelial Gowth Factor Correlates with Hasford Score in Chronic Myeloid Leukemia.

Introduction and Aim Increased angiogenesis in BM is one of the characteristics of chronic myeloid leukemia (CML) implicated in its progression. Vascular endothelial growth factor (VEGF) one of the most potent regulator of angiogenesis is increased in CML. The prognostic impact of serum VEGF in CML is largely unknown with sparse literature from India. So the present study aimed to measure serum VEGF levels in different phases of CML and to assess its prognostic significance using Hasford score. Methods Forty Ph + patients of CML were enrolled in the study. Complete clinical history and physical examination was done. Hemogram was done by Beckman Coulter LH 500. Peripheral smear (Wright's stain) was done by microscopy. Serum VEGF (plain vial) using ELISA was calculated. Statistical analysis was performed using SPSS software version 20. Results The mean serum VEGF levels were significantly higher in patients than in controls (p < 0.0001). The patients in accelerated/blast phase demonstrated significantly higher levels of serum VEGF (mean 151 pg/mL) than those in the chronic phase (mean 90.87 pg/mL) (p = 0.02). Serum VEGF levels showed a significant positive correlation with the overall Hasford prognostic score (p = 0.023). Conclusion Serum VEGF levels can serve as an independent prognostic marker in CML patients irrespective of phase of CML. Also, S. VEGF levels can be used to monitor patients on imatinib therapy and identify those who might benefit from antiangiogenesis therapy. However, larger studies are needed with a larger number of patients in different phases of CML to validate our findings and thus pave the way for future research.

Optimization of growth conditions for enhancing the production of microbial laccase and its application in treating antibiotic contamination in wastewater.

In this work, seven indigenous macrofungal isolates were selected to screen for their laccase production capability. Among them, isolates viz., Pleurotus eryngii, Pleurotus florida, Pleurotus sajor caju and Gandoderma lucidum were found to exhibit high laccase activity in the preliminary studies and were thus selected for the optimization studies with an aim to enhance laccase production. The pH optimization studies were carried out between pH range of 4-6. The laccase activity and biomass were found to be optimum at pH 4, 4.5, 4.5 and 5 for P. eryngii, P. florida, P. sajor caju and G. lucidum, respectively. Optimization studies with chemical inducers namely, tannic acid, 2,6 dimethoxyphenol and copper sulphate at three different concentration levels were conducted and tannic acid at 2 mM concentration was found to increase the laccase activity to about 45% followed by 2,6 dimethoxyphenol (2 mM) with an increase of about 43% and copper sulphate (0.1 mM) showing 21% increase in the yield. Biodegradation studies utilizing laccase isolated from P. eryngii, P. florida and P. sajor caju was carried out for a commonly detected fluoroquinolone antibiotic, levofloxacin, in water and pharmaceutical wastewater. The results indicated that the degradation efficiency of levofloxacin using laccase isolated from P. eryngii (88.9%) was comparable to commercial laccase (89%). When the cost economics of using crude laccase was evaluated against commercial laccase it was evident that the total cost of the treatment could be reduced by 71.7% if commercial grade laccase was replaced by crude enzyme extracted from indigenous macrofungi such Pleurotus eryngii, Pleurotus florida, and Pleurotus sajor caju indicating a promising and cost-effective alternative for wastewater treatment.

Treatment technologies to mitigate the harmful effects of recalcitrant fluoroquinolone antibiotics on the environ- ment and human health.

Antibiotic proliferation in the environment and their persistent nature is an issue of global concern as they induce antibiotic resistance threatening both human health and the ecosystem. Antibiotics have therefore been categorized as emerging pollutants. Fluoroquinolone (FQs) antibiotics are an emerging class of contaminants that are used extensively in human and veterinary medicine. The recalcitrant nature of fluoroquinolones has led to their presence in wastewater, effluents and water bodies. Even at a low concentration, FQs can stimulate antibacterial resistance. The main sources of FQ contamination include waste from pharmaceutical manufacturing industries, hospitals and households that ultimately reaches the wastewater treatment plants (WWTPs). The conventional WWTPs are unable to completely remove FQs due to their chemical stability. Therefore, the development and implementation of more efficient, economical, convenient treatment and removal technologies are needed to adequately address the issue. This review provides an overview of the technologies available for the removal of fluoroquinolone antibiotics from wastewater including adsorptive removal, advanced oxidation processes, removal using non-carbon based nanomaterials, microbial degradation and enzymatic degradation. Each treatment technology is discussed on its merits and limitations and a comparative view is presented on the choice of an advanced treatment process for future studies and implementation. A discussion on the commercialization potential and eco-friendliness of each technology is also included in the review. The importance of metabolite identification and their residual toxicity determination has been emphasized. The last section of the review provides an overview of the policy interventions and regulatory frameworks that aid in retrofitting antibiotics as a central key focus contaminant and thereby defining the discharge limits for antibiotics and establishing safe manufacturing practices.

Glycol chitosan in situ coating on PLGA nanoparticle curtails extraneous paclitaxel precipitates and imparts protein corona independent hemocompatibility.

Poly (lactide-co-glycolide) (PLGA) nanoparticles surface functionalized with water soluble glycol chitosan (GC) and carboxymethyl chitosan (CMC) has been studied for their drug (Paclitaxel and Doxorubicin) loading, yield, cellular uptake, serum protein adsorption and hemocompatibility. It was observed that Paclitaxel (Ptxl) phase out as Extraneous Ptxl Precipitates (EPP) (>25 %) in case of uncoated and CMC coated low molecular weight (LMW) PLGA nanoparticles (PNPs). The EPP formation was significantly reduced to ∼5 % with GC coating as it enhanced LMW PLGA precipitation and yield predominantly spherical polymeric nanoparticles towards better encapsulation of Ptxl and thus uniform intracellular drug distribution. Interestingly, protein corona analysis showed cmcPNPs and gcPNPs to be distinct from each other in associating mainly with serum proteins of molecular weight < 30 kDa and >30 kDa respectively. While CMC functionalization showed >10 % hemolysis, at similar concentration GC coating was found to provide superior hemocompatibility even in the absence of protein corona.

Glycol chitosan assisted in situ reduction of gold on polymeric template for anti-cancer theranostics.

Multifunctional biodegradable nanomaterials that could be used for both imaging and therapy are being researched extensively. A simple technique to synthesize multifunctional nanoparticles without compromising on any of their functionality is a challenge. We have attempted to optimize a two-step procedure of gold coated polymeric template involving 1) Single pot synthesis of PLGA nanoparticles with cationic surface charge using glycol chitosan and 2) in situ gold coating for formation of gold coated PLGA nanoshell (AuPLGA-NS). These gold-coated PLGA nanoparticles were explored for photothermal therapy (PTT) and as X-ray/CT contrast agents. Biocompatibility and photothermal cytotoxicity of AuPLGA-NS were evaluated in-vitro and results confirmed the therapeutic efficacy of these particles resulting in 80% cancer cell death. Besides, it also showed potential X-ray/CT imaging ability with contrast equivalent to that of Iodine. The results demonstrated that these gold-coated PLGA nanoparticles synthesized by a simple approach could be used as a multifunctional nanosystem for cancer theranostics.