Microbial membrane transport proteins and their biotechnological applications.

Because of the hydrophobic nature of the membrane lipid bilayer, the majority of the hydrophilic solutes require special transportation mechanisms for passing through the cell membrane. Integral membrane transport proteins (MTPs), which belong to the Major Intrinsic Protein Family, facilitate the transport of these solutes across cell membranes. MTPs including aquaporins and carrier proteins are transmembrane proteins spanning across the cell membrane. The easy handling of microorganisms enabled the discovery of a remarkable number of transport proteins specific to different substances. It has been realized that these transporters have very important roles in the survival of microorganisms, their pathogenesis, and antimicrobial resistance. Astonishing features related to the solute specificity of these proteins have led to the acceleration of the research on the discovery of their properties and the development of innovative products in which these unique properties are used or imitated. Studies on microbial MTPs range from the discovery and characterization of a novel transporter protein to the mining and screening of them in a large transporter library for particular functions, from simulations and modeling of specific transporters to the preparation of biomimetic synthetic materials for different purposes such as biosensors or filtration membranes. This review presents recent discoveries on microbial membrane transport proteins and focuses especially on formate nitrite transport proteins and aquaporins, and advances in their biotechnological applications.

Comparison of ion selectivities of nitrite channel NirC and water channel aquaporin.

nirC gene coding for the nitrite channel of E. coli K12 was cloned into the pET28a vector and expressed in E. coli BL21 cells. 28.5 kDa NirC monomer was purified from membrane components of E. coli. Selectivity of NirC for different ions including nitrite, nitrate, sulfate, formate, and acetate anions, and a divalent cation, magnesium, was compared with that of bacterial aquaporin from Halomonas elongata. Water and ion permeability values were determined by measuring the light scattering rates of proteoliposomes containing NirC and aquaporins during their water loss and gain. NirC shows a selective permeability to nitrite and is more resistant to the entry of other anions as compared to aquaporin. The single channel permeability of NirC for nitrite is about 10-fold that of a single aquaporin channel. Both aquaporin and NirC channel proteins were impermeable to MgCl2 and (NH4)2SO4 and their permeability to other tested ions was remarkably lower as compared to nitrite ions. The study also presents the 3D model and channel characteristics of NirC. The translocation channel of E. coli NirC is determined to be larger, and its length is shorter than aquaporin channels. Although the NirC channel throat is more hydrophobic than aquaporin, its water permeability is almost equal to that of aquaporin. The hydrophobic nature of the NirC channel might play an important role in the selective permeability of the channel for nitrite ions.

A Rare Cause of Hypophosphatemia: Raine Syndrome Changing Clinical Features with Age.

Raine Syndrome (RS) is caused by biallelic loss-of-function mutations in FAM20C gene and characterized by hypophosphatemia, typical facial and skeletal features. Subperiosteal bone formation and generalized osteosclerosis are the most common radiological findings. Here we present a new case with RS. A 9-month-old male patient on a home-type ventilator was referred for hypophosphatemia. He was born with a weight of 3800 g to non-consanguineous parents. Prenatal ultrasound had demonstrated nasal bone agenesis. A large anterior fontanel, frontal bossing, exophthalmos, hypoplastic nose, high arched palate, low set ears, triangular mouth, and corneal opacification were detected on physical examination. Serial skeletal X-rays revealed diffuse osteosclerosis at birth which was gradually decreased by the age of 5 months with subperiosteal undermineralized bone formation and medullary space of long bone could be distinguishable with bone-within-a-bone appearance. At 9 months of age, hand X-ray revealed cupping of the ulna with loose radial bone margin with minimal fraying and osteopenia. Cranial computed tomography scan showed bilateral periventricular calcification and hydrocephalus in progress. The clinical, laboratory, and radiological examinations were consistent with RS. Molecular analyses revealed a compound heterozygous mutation in FAM20C gene (a known pathogenic mutation, c.1645C > T, p.Arg549Trp; and a novel c.863 + 5 G > C variant). The patient died due to respiratory failure at 17 months of age. This case allowed us to demonstrate natural progression of skeletal features in RS. Furthermore, we have described a novel FAM20C variant causing RS. Previous literature on RS is also reviewed.

Affinity tag effect on the salt rejection potential of Halomonas elongata aquaporin incorporated in thin film nanocomposite membrane.

In this study, effect of affinity tags, Histidine (His) and Glutathione-S-Transferase (GST), on the activity of halophilic aquaporin was analyzed. The gene coding for H. elongata aquaporin was cloned into pET28a vector and expressed in E. coli BL21 successfully. Stopped flow light scattering measurements showed that His-tagged aquaporin is functional. The difference in the filtration parameters caused by affinity tags were determined by using thin film composite nano-filtration (NFC) membranes prepared with the aquaporins. At 100 mM salt concentration, water permeability (L/m2.h) and the % salt rejection of NFC membranes produced with the His-tagged aquaporin was found to be higher than that of the membrane with GST-tagged aquaporin. Salt rejection of His-tagged aquaporin-membrane was found to be 53% with a lower solute permeability value (B). Use of short affinity tag (His tag) for cloning resulted in higher solute rejection ability of TFC membranes prepared with H. elongata aquaporins.

Changing clinical characteristics of non-cystic fibrosis bronchiectasis in children.

BACKGROUND: The prevalence of non-cystic fibrosis (CF) bronchiectasis is increasing in both developed and developing countries in recent years. Although the main features remain similar, etiologies seem to change. Our aim was to evaluate the clinical and laboratory characteristics of our recent non-CF bronchiectasis patients and to compare these with our historical cohort in 2001. METHODS: One hundred four children with non-CF bronchiectasis followed between 2002 and 2019 were enrolled. Age of diagnosis, underlying etiology and microorganisms in sputum culture were recorded. Clinical outcomes were evaluated in terms of lung function tests and annual pulmonary exacerbation rates at presentation and within the last 12 months. RESULTS: Mean FEV1 and FVC %predicted at presentation improved compared to historical cohort (76.6 ± 17.1 vs. 63.3 ± 22.1 and 76.6 ± 15.1 vs. 67.3 ± 23.1, respectively; p <  0.001). There was a significant decrease in pulmonary exacerbation rate from 6.05 ± 2.88 at presentation to 3.23 ± 2.08 during follow-up (p <  0.0001). In 80.8% of patients, an underlying etiology was identified. There was an increase in primary ciliary dyskinesia (PCD) (32.7% vs. 6.3%; p = 0.001), decrease in idiopathic cases (19.2% vs. 37.8%; p = 0.03) with no change in postinfectious and immunodeficiencies as underlying etiology. Sputum cultures were positive in 77.9% of patients which was 46.9% in the historical cohort (p = 0.001). CONCLUSION: Baseline pulmonary function tests were better and distribution of underlying etiology had changed with a remarkable increase in diagnosis of PCD in the recent cohort.

Spatial distribution and temporal trends of VOCs in a highly industrialized town in Turkey.

An extensive monitoring study of volatile organic compounds (VOCs) was conducted at sites across the highly industrialized town of Dilovasi, northern Turkey to determine temporal and spatial trends in pollutant concentrations and relate to the effects of source locations, meteorology, and topography. Two-week passive samplers (Tenax tubes) were deployed at twelve sites from February to December 2012 and analysed using thermal desorption and gas chromatography with mass spectrometric detection (TD-GC-MS). Sampled total VOC (TVOC) levels were highest in the July through October period and were highest at low-altitude sites near industry facilities and vehicle traffic sources (148.3 µg/m(3) at site 11, 154.1 µg/m(3) at site 10) and lowest at high-altitude sites located furthest upwind from industry and traffic sources (78.4 µg/m(3) at site 5 and 78.5 µg/m(3) at site 6). Analysis of "T/B" ratios suggested that contributions to ambient VOC in Dilovasi are dominated by the town's industrial sources. Meteorological conditions and the town's basin topography were also found to significantly influence the city's air quality, with strong winds from the NE observed to correlate with periods of higher sampled TVOC. Compared with other industrialized urban centers, the study revealed that there is significant toluene pollution in Dilovasi and recommended enhanced continuous monitoring at the city's industrial and residential zones.

Heavy metal content of total suspended air particles in the heavily industrialized town of Gebze, Turkey.

Air pollution is a serious environmental problem in industrialized towns, where a significant portion of the residents live in close proximity to factories and major highways with high traffic load. In this study, the ambient air quality in Gebze, an industrial region with an area of 438 km(2) and a population of 300,000, was characterized in terms for total suspended particulate matter and its composition of trace elements, i.e. Cd, Cr, Cu, Fe, Mn, Ni and Pb. Samples were collected using high volume samplers from March to June 2009 at two sites during the day and the night. A significantly higher Cu concentrations during night suggested that Cu emissions were the result of a local source. The known air toxics, Cd and Ni, had average concentrations (34 and 43 ng/m(3), respectively) higher than proposed by the European Union's ambient air quality standards. These results highlight the potential health risks for the local population.

Biological NOx removal by denitrification process in a jet-loop bioreactor: system performance and model development.

Nitrogen monoxide (NO) and nitrogen dioxide referred as NOx are one of the most important air pollutants in the atmosphere. Biological NOx removal technologies have been developing to reach a cost-effective control method for upcoming stringent NOx emission standards. The BioDeNOx system was seen as a promising biological NOx control technology which is composed of two reactors, one for absorbing of NO in an aqueous Fe(II)EDTA2- solution and the other for subsequent reduction to N2 gas in a biological reactor by the denitrification process. In this study, instead of two discrete reactors, only one jet-loop bioreactor (JLBR) was utilized as both absorption and denitrification unit and no chelate-forming chemicals were added. In other words, the advantage of better mass transfer conditions of jet bioreactor was used instead of Fe(II)EDTA2-. The process was named as Jet-BioDeNOx. The JLBR was operated for the removal of NOx from air streams containing 500-3000 ppm NOx and the results showed that the removal efficiency was between 81% and 94%. The air to liquid flow ratio (Q(G)/Q(RAS)) varied in the range of 0.07-0.12. Mathematical modelling of the system demonstrated that the removal efficiency strongly depends on this ratio. The high mass transfer conditions prevailed in the reactor provided a competitive advantage on removing NO gas without any requirement of chelating chemicals.

Implementation of a high fidelity simulation based training program for physicians of children requiring long term invasive home ventilation: a study by ISPAT team.

Introduction: The number of children requiring long-term invasive home ventilation (LTIHV) has increased worldwide in recent decades. The training of physicians caring for these children is crucial since they are at high risk for complications and adverse events. This study aimed to assess the efficacy of a comprehensive high-fidelity simulation-based training program for physicians caring for children on LTIHV. Methods: A multimodal training program for tracheostomy and ventilator management was prepared by ISPAT (IStanbul PAediatric Tracheostomy) team. Participants were subjected to theoretical and practical pre-tests which evaluated their knowledge levels and skills for care, follow-up, and treatment of children on LTIHV. Following the theoretical education and hands-on training session with a simulation model, theoretical and practical post-tests were performed. Results: Forty-three physicians from 7 tertiary pediatric clinics in Istanbul were enrolled in the training program. Seventy percent of them had never received standardized training programs about patients on home ventilation previously. The total number of correct answers from the participants significantly improved after the theoretical training (p < 0.001). The number of participants who performed the steps correctly also significantly increased following the hands-on training session (p < 0.001). All of the 43 participants who responded rated the course overall as good or excellent. Conclusion: The knowledge and skills of clinicians caring for children on LTIHV can be enhanced through a comprehensive training program consisting of theoretical training combined with hands-on training in a simulation laboratory.

Night-to-night variability of polygraphy in children with sleep disordered breathing symptoms.

INTRODUCTION: Polygraphy (PG) can be used as an alternative test for the diagnosis of obstructive sleep apnea syndrome (OSAS) in children. Night-to-night variability of PG in children is not known. Our aim was to determine whether a single night PG was reliable for OSAS diagnosis in children with symptoms of sleep-disordered breathing (SDB). MATERIALS AND METHODS: Otherwise healthy children who had been evaluated for symptoms of SDB were included. Two nocturnal PGs were performed 2-7 days apart. Demographic and clinical characteristics, Pediatric Sleep Questionnaire, and modified Epworth Sleepiness Scale were recorded. OSAS was diagnosed if obstructive apnea-hypopnea index was (oAHI) ⩾ 1/h and classified as mild (oAHI: 1-4.9/h), moderate (oAHI: 5-9.9/h), and severe (oAHI ⩾ 10/h). RESULTS: Forty-eight patients were included (37.5% female, age 10.8 ± 3.9 years) to the study. There were no significant differences in oAHI values and other respiratory parameters between the two PGs (p > 0.05). Thirty-nine children were diagnosed with OSAS if the highest oAHI over any single night was used for diagnosis. Thirty-three of the 39 children (84.6%) were diagnosed with OSAS with the first PG while 35 of 39 (89.7%) children were diagnosed with OSAS with the second PG. There was an agreement for identifying OSAS and its severity between the two PGs in our study even though there were few individual intra-subject differences in oAHI. CONCLUSION: There was no significant first-night effect for PG in this study which suggests that a single night PG is adequate for diagnosis of OSAS in children with SDB- related symptoms.

Optimization of solid-phase microextraction using Taguchi design to quantify trace level polycyclic aromatic hydrocarbons in water.

This article introduces a simple, rapid, and reliable solid-phase microextraction (SPME) method coupled with GC-MS for the quantitative determination of 16 polycyclic aromatic hydrocarbons in water. In this study, the Taguchi experimental design was used to optimize extraction conditions of polycyclic aromatic hydrocarbons using SPME method to obtain highly enriched analytes. Consequently, quantitative determination of polycyclic aromatic hydrocarbons in water was achieved by GC-MS technique. The selected parameters affecting enrichment of polycyclic aromatic hydrocarbons were sample extraction time, stirring speed, temperature, ionic strength, and pH. The study revealed that optimal operating conditions were found to be 90-min extraction time, 1400 rpm stirring speed, and 60°C sample temperature. The effect of ionic strength and pH were shown to be insignificant. Optimized conditions were also reevaluated by placing the 16 polycyclic aromatic hydrocarbons into several subgroups based on their molecular weight. The extraction efficiency of polycyclic aromatic hydrocarbons with low molecular weight was shown to be a function of only the extracting temperature. Satisfactory results were obtained for linearity (0.983-0.999), detection limits (2.67-18.02 ng/L), accuracy (71.2-99.3%), and precision (4.3-13.5%). The optimum conditions reported by other design approaches were evaluated and generalized optimum conditions were suggested.

Nitric oxide and nitrite removal by partial denitrifying hollow-fiber membrane biofilm reactor coupled with nitrous oxide generation as energy recovery.

Nitrogen oxide (NOx) emissions cause significant impacts on the environment and must therefore be controlled even more stringently. This requires the development of cost-effective removal strategies which simultaneously create value-added by-products or energy from the waste. This study aims to treat gaseous nitric oxide (NO) by hollow-fibre membrane biofilm reactor (HFMBfR) in the presence of nitrite (NO2-) and evaluate nitrous oxide (N2O) emissions formed as an intermediate product during the denitrification process. Accumulated N2O can be utilised in methane oxidation as an oxidant to produce energy. In the first stage of the study, the HFMBfR was operated by feeding only gaseous NO as the nitrogen source. During this period, the best performance was achieved with 92% NO removal efficiency (RE). In the second stage, both NO gas and NO2- were supplied to the system, and 91% NO and 99% NO2- reduction were achieved simultaneously with the maximum N2O generation of 386 ± 31 ppm. Lower influent carbon to nitrogen (C/N) ratios, such as 4.5 and 2.0, and higher NO2--N loading rate of 158 mg N day-1 favoured N2O generation. An improved NO removal rate and N2O accumulation were seen with the increasing amount of PO43- in the medium. The 16S rDNA sequencing analysis revealed that Alicycliphilus denitrificans and Pseudomonas putida were the dominant species. The study shows that an HFMBfR can be successfully used to eliminate both NO2- and gaseous NO and simultaneously generate N2O by adjusting the system parameters such as C/N ratio, NO2- and PO43- loading.

Nitrite is reduced by nitrite reductase NirB without small subunit NirD in Escherichia coli.

The assimilatory nitrite reductase enzyme NirB and small subunit NirD genes encoded in nir operon in Escherichia coli were cloned into the pET28a vector, and the recombinant enzyme was characterized for the first time. Docking of NirB with NirD, NADH, NO2-, NO3-, and CHO2- was performed using docking modeling programs. Methyl viologen and sodium dithionite were used as electron couples, and the amount of reduced nitrite was measured to calculate enzyme activity. NirB is the main enzyme and shows high activity with or without NirD. However, the inclusion of NirD into the enzyme solution at a ratio of 1NirD:2NirB resulted in 10% higher nitrite reductase activity. The enzyme tends to aggregate in the absence of ß-mercaptoethanol, which causes the conversion of tetrameric NirB to monomeric form, and the NirB enzyme shows its highest activity in monomeric form. The optimum temperature for enzyme activity was 37 °C and the optimum pH was found to be 7.0. Km and Vmax values of NirB were calculated as 9833 µM and 416.67 µmol NO2- reduced min-1 mg-1. Enzyme activity decreased by 55% and 50% in the presence of 100 mM nitrate and formate, respectively. The presence of 25 mM Cd2+ protected the enzyme at room temperature and the enzyme showed 10% higher activity in the presence of cadmium.

Collaboration between two CF centers; one in USA and one in Turkey before and during CoV2 pandemic.

To address the discrepancy in the quality of care and outcomes between cystic fibrosis centers (CFCs) in high-income countries and limited resources countries (LRCs), a collaboration between our team at the University of Michigan CFC (UMCFC) and a CF center in Turkey (Marmara University CFC [MUCFC], Istanbul) was established. The collaboration included evaluation of all aspects of care and initiation of quality improvement (QI) measures. Teaching and implementing QI tools has led to start of improvement in MUCFC care. Close monitoring and sharing resources like UMCFC algorithms, protocols, and QI processes were done.

COVID-19 disease characteristics in different pediatric age groups.

INTRODUCTION: Little is known about the COVID-19 disease characteristics and differences between different pediatric age groups. This study aimed to investigate the disease characteristics according to age groups. METHODOLOGY: We conducted a retrospective, single-center study of pediatric COVID-19 in a tertiary care hospital in Turkey. The patients were divided into three groups: 15 days-24 months old (Group 1), 25-144 months old (Group 2), and 145-210 months old (Group 3) according to age. RESULTS: A total of 139 pediatric patients with COVID-19 were examined. Twenty-nine patients (20.9%) were in Group 1, 52 (37.4%) were in Group 2, 58 (41.7%) were in Group 3. Thirty-nine patients (28.1%) were hospitalized. The most common symptoms were cough (55.4%) and fever (51.8%). The median chest X-ray (CXR) score of hospitalized patients was 1 (min 0-max 7), and the median CXR score of outpatients was 1 (min 0-max 6). Fever was significantly more frequent in Group 1, and chest pain was more frequent in Group 3. Group 1 had significantly higher WBC, lymphocyte, thrombocyte counts, AST, LDH, D-dimer, and Troponin T levels but lower hemoglobin, total protein, and albumin levels. The treatment included antibiotics, oseltamivir, hydroxychloroquine, and supportive therapy. Only one patient (0.7%) received non-invasive mechanical ventilatory support. CONCLUSIONS: As we know the clinical course of COVID-19 in children is less severe than in adults. We also found significant differences in both clinical and laboratory findings between different pediatric age groups which supports the theory that disease pathogenesis is highly variable according to age.

The Burden of Primary Caregivers of Spinal Muscular Atrophy Patients and Their Needs.

AIM: This study aims to reveal the problems faced by families of children with spinal muscular atrophy (SMA), by evaluating their care burden, needs, and expectations. MATERIALS AND METHODS: The participants were the primary caregivers of 34 children between the ages of 0 and 18 years diagnosed with SMA. Thirteen children were diagnosed with type 1, 13 children with type 2 and 8 children with type 3 SMA. Data on the medical history, functional levels of the participants, and the characteristics of families were collected. The childrens' parents completed the Family Needs Survey and the Zarit Caregiver Burden Scale. RESULTS: According to the results of the Family Needs Survey, it was found that information was the most common requirement, and this was independent of the level of education. According to the Caregiver Burden Scale, it was recorded that 64.7% of the caregivers were under mild/moderate burden. While there was a moderate correlation (r = 0.574; P < .001) between the Caregiver Burden Scale and the Family Needs Survey, it was observed that the functional level of the child was not associated with family needs and caregiver burden. CONCLUSIONS: Our study suggests that the needs of families of SMA patients, especially related to income level, have changed. The caregivers' burden is not directly related to the income level or the functional level of the child. Families' need for information should also be prioritized within the rehabilitation program.

Thermodynamically designed target-specific DNA probe as an electrochemical hybridization biosensor.

Applications of molecular techniques to elucidate identity or function using biomarkers still remain highly empirical and biosensors are no exception. In the present study, target-specific oligonucleotide probes for E. coli K12 were designed thermodynamically and applied in an electrochemical DNA biosensor setup. Biosensor was prepared by immobilization of a stem-loop structured probe, modified with a thiol functional group at its 5' end and a biotin molecule at its 3' end, on a gold electrode through self-assembly. Mercaptopropionic acid (MPA) was used to optimize the surface probe density of the electrode. Hybridization between the immobilized probe and the target DNA was detected via the electrochemical response of streptavidin-horseradish peroxidase in the presence of the substrate. The amperometric response showed a linear relationship with the target DNA concentration, ranging from 10 and 400 nM, with a correlation coefficient of 0.989. High selectivity and good repeatability of the biosensor showed that the thermodynamic approach to oligonucleotide probe design can be used in development of electrochemical DNA biosensors.

Comparison of conventional chest physiotherapy and oscillatory positive expiratory pressure therapy in primary ciliary dyskinesia.

BACKGROUND: Chest physiotherapy (CP) is a recommended treatment modality in primary ciliary dyskinesia (PCD). OBJECTIVE: Primary aim was to compare the efficacy and safety of the conventional chest physiotherapy (CCP) and oscillatory positive expiratory pressure therapy (OPEPT). Secondary aims were to compare the exacerbation rate, time until the first exacerbation, patient compliance and comfort between the two CP methods. METHODS: This is a 6 month randomized, controlled crossover trial. Patients >6 years of age with PCD were randomized into two groups, first group was assigned to OPEPT (Acapella®) for 3 months while second group was assigned to CCP. Groups were crossed over to the other modality after a 15-day washout period. Pulmonary function tests (PFTs) and compliance were monitored by monthly clinic visits. RESULTS: There was a significant increase in FEV1 , FEF25-75 , and PEF values (p = .018, p = .020, and p = .016, respectively) in the OPEPT group and in FVC values (p = .007) in CCP group compared to baseline. However PFT increase at 3rd month was not superior to each other with both physiotherapy methods. Median acute pulmonary exacerbation rate and time period until the first exacerbation were similar in both groups (p = .821, p = .092, respectively). Comfort and effectiveness of OPEPT was higher than CCP according to patients (p = .029 and p = .042, respectively). There were no adverse effects with either therapy. CONCLUSIONS: OPEPT was as effective as CCP in PCD patients. OPEPT was more comfortable and effective than CCP according to patients. OPEPT might be an efficient alternative method for airway cleareance in PCD patients.

Health-related quality of life in patients with bronchiolitis obliterans.

INTRODUCTION: Bronchiolitis obliterans (BO) is mainly caused by infections and hematopoietic stem cell transplantation (HSCT). This study aimed to investigate the health-related quality of life (HRQOL) of children with BO compared to the healthy children and also to assess the HRQOL according to the etiology. METHODS: Postinfectious (group 1) and post-HSCT BO (group 2) patients and healthy children were included in the study. HRQOL was assessed by the Short Form-36 (SF-36) and St George's Respiratory Questionnaire (SGRQ). Correlations between demographic and clinical characteristics, pulmonary function tests, high-resolution chest tomography scores, and HRQOL were assessed. RESULTS: Thirty-seven postinfectious and post-HSCT BO patients and 34 healthy children were included in the study. Mean age was 13.8 ± 0.7 years. Mean forced vital capacity and forced expiratory volume1 were 60.7 ± 2.7% predicted, and 49.8 ± 3.1% predicted, respectively. The SF-36 scores were lower in BO patients compared to healthy children (P < .01). Patients with better lung functions had higher SF-36 scores, but lower SGRQ. The number of inhaled therapies, acute exacerbations, hospitalizations were inversely correlated with SF-36. A positive correlation was found between these parameters and total SGRQ scores (r = .507, P = .02; r = .409, P = .12; r = .326, P = .049, respectively). SF-36 scores were better in group 1 for subscales of physical role functioning and social role functioning compared to group 2. (P = .01, P = .01, respectively). CONCLUSION: The HRQOL of patients with BO measured by SF-36 was low compared to healthy children. SF-36 scores were more affected in post-HSCT BO patients. HRQOL of children with chronic lung disease should be taken into consideration in the management of these patients.

Amperometric nitrate biosensor based on Carbon nanotube/Polypyrrole/Nitrate reductase biofilm electrode.

This study describes the construction and characterization of an amperometric nitrate biosensor based on the Polypyrrole (PPy)/Carbon nanotubes (CNTs) film. Nitrate reductase (NR) was both entrapped into the growing PPy film and chemically immobilized via the carboxyl groups of CNTs to the CNT/PPy film electrode. The optimum amperometric response for nitrate was obtained in 0.1 M phosphate buffer solution (PBS), pH 7.5 including 0.1 M lithium chloride and 7 mM potassium ferricyanide with an applied potential of 0.13 V (vs. Ag/AgCl, 3 M NaCl). Sensitivity was found to be 300 nA/mM in a linear range of 0.44-1.45 mM with a regression coefficient of 0.97. The biosensor response showed a higher linear range in comparison to standard nitrate analysis methods which were tested in this study and NADH based nitrate biosensors. A minimum detectable concentration of 0.17 mM (S/N=3) with a relative standard deviation (RSD) of 5.4% (n=7) was obtained for the biosensor. Phenol and glucose inhibit the electrochemical reaction strictly at a concentration of 1 µg/L and 20 mg/L, respectively. The biosensor response retained 70% of its initial response over 10 day usage period when used everyday.