Synthesis, characterization and fabrication of polypyrrole and novel 3-substituted polypyrrole biosensors

This thesis presents the results of the investigations of the use of conducting polymers of unsubstituted and substituted pyrroles for the fabrication of sulfite and catechol biosensors. Part-I of this thesis consists of two chapters. Chapter 1 presents a brief introduction to biosensors, conducting polypyrrole, polypyrrole based biosensors, sulfite and phenolic biosensors, as well as the aims and objectives of this project. The extensive use of polypyrrole (PPy) in the development of electronic noses (ENs) and electronic tongues (ETs) as multi analytical devices was highlighted. A review of the use of PPy based ENs for environmental and industrial analysis is presented in the Chapter 2. The diverse use of PPy in these devices as a transduction material for a wide range of toxic and non-toxic substances, such as ammonia, nitrogen oxides, carbon monoxide, sulphur dioxide, hydrogen sulphide, methane, oxygen, hydrogen, alcohols, phenol, benzene and water vapours, is presented. The development, fabrication and characterization of unsubstituted pyrrole biosensors for sulfite and catechol is presented in Part-II of this thesis. Chapter 3 describes the use of immobilisation of sulfite oxidase (SOx) into a 54 nm thick PPy film for the fabrication of a polypyrrole-based amperometric nanobiosensor for reliable sulfite determination. Optimum immobilisation was accomplished by galvanostatic polymerization in a monomer solution which contained 0.1 M pyrrole and 5 units of SOx with a polymerization period of 120 seconds and an applied current density of 0.2 rnA cm-2• The presence of SOx in the PPy film was confirmed by scanning electron microscopy and amperometric measurement of sulfite. The effect of pH and interferants on sulfite determination with the nanobiosensor was also investigated. A linear concentration range of 0.9 to 400 JlM sulfite was achieved with the nanobiosensor and the minimum detectable sulfite concentration was 0.9 JlM (0.1 ppm). The successful application of the nanobiosensor to the determination of sulfite in wine and beers samples is reported. Chapter 4 further explores the use ofunsubstituted pyrrole for the development of a polypyrrole-tyrosinase (PPy-Tyr) potentiometric biosensor. The entrapment of tyrosinase into the polypyrrole film was carried out by galvanostatic polymerization on a platinum electrode. The optimum conditions for the formation of the PPy-Tyr film include a current density of 0.5 rnA cm·2 , a polymerization period of 150 seconds, 0.1 M pyrrole and 50 U mL-1 tyrosinase. Scanning electron microscopy (SEM), cyclic voltammetry (CV) and potentiometric measurements were used to confirm the presence of tyrosinase in the polymer film. The PPy-Tyr biosensor gave a sensitivity of 10 m VI J..LM for catechol, with a response time of 80 seconds and a linear concentration range of 1-50 J..LM catechol and the minimum detectable concentration was 1.0 J..LM. The PPy-Tyr biosensor was stable for at least 1 month when stored in a buffer at about 4 • C. The use of 3-substituted pyrroles for the fabrication of sulfite and phenol biosensor was explored in Part-III of this thesis by synthesizing 3-ethylpyrrole (EtPy). The final and intermediate products for the synthesis of EtPy were confirmed by fourier transform infrared (FTIR), nuclear magnetic resonance eH NMR and 13C NMR) spectroscopy and mass spectrometry (MS). The synthesized EtPy was employed in Chapter 5 for the fabrication of an amperometric sulfite biosensor. Sulfite oxidase was immobilized by galvanostatic polymerization of an adsorbed EtPy-SOx layer. The optimum conditions for the formation ofpoly(3-ethylpyrrole)-sulifte oxidase (PEtPy-SOx) biosensor include physical adsorption of 0.2 mg EtPy, 0.5 U of SOx, a current density of 0.75 rnA cm·2 and a polymerization period of 10 min. The PEtPy-SOx biosensor can detect as little as 0.1 J..LM sulfite and gave a linear concentration range of0.1-400 J..LM sulfte with a response time of 40 seconds. The electropolymerization of an adsorbed 3-ethylpyrrole-tyrosinase layer on a platinum electrode was investigated in Chapter 6 for the fabrication of a novel poly(3- ethylpyrrole)-tyrosinase (PEtPy-Tyr) based potentiometric catechol biosensor. The optimum conditions for the formation ofthe PEtPy-Tyr biosensor include 0.038 mg of3- ethylpyrrole (EtPy), 15 U of tyrosinase, a current density of 0.7 rnA cm·2 and a polymerization period of 15 min. The PEtPy-Tyr biosensor produced a reliable and sensitive response to catechol with a linear concentration range of 1-350 J!M, a minimum detectable catechol concentration of 0.1 J!M and a response time of 50 seconds. Chapter 7 concludes that both unsubstituted and substituted pyrroles (in this case 3-ethylpyrrole) are useful for fabrication ofbiosensors for sulfite and catechol. However, it was, noted that the use of the substituted pyrrole resulted in a substantial improvement of the sensitivity and minimum detectable concentration achieved for both sulfite and catechol biosensor. Also it resulted in a considerable reduction in the required enzyme concentrations

Synthesis, characterization and fabrication of polypyrrole and novel 3-substituted polypyrrole biosensors

This thesis presents the results of the investigations of the use of conducting polymers of unsubstituted and substituted pyrroles for the fabrication of sulfite and catechol biosensors. Part-I of this thesis consists of two chapters. Chapter 1 presents a brief introduction to biosensors, conducting polypyrrole, polypyrrole based biosensors, sulfite and phenolic biosensors, as well as the aims and objectives of this project. The extensive use of polypyrrole (PPy) in the development of electronic noses (ENs) and electronic tongues (ETs) as multi analytical devices was highlighted. A review of the use of PPy based ENs for environmental and industrial analysis is presented in the Chapter 2. The diverse use of PPy in these devices as a transduction material for a wide range of toxic and non-toxic substances, such as ammonia, nitrogen oxides, carbon monoxide, sulphur dioxide, hydrogen sulphide, methane, oxygen, hydrogen, alcohols, phenol, benzene and water vapours, is presented. The development, fabrication and characterization of unsubstituted pyrrole biosensors for sulfite and catechol is presented in Part-II of this thesis. Chapter 3 describes the use of immobilisation of sulfite oxidase (SOx) into a 54 nm thick PPy film for the fabrication of a polypyrrole-based amperometric nanobiosensor for reliable sulfite determination. Optimum immobilisation was accomplished by galvanostatic polymerization in a monomer solution which contained 0.1 M pyrrole and 5 units of SOx with a polymerization period of 120 seconds and an applied current density of 0.2 rnA cm-2• The presence of SOx in the PPy film was confirmed by scanning electron microscopy and amperometric measurement of sulfite. The effect of pH and interferants on sulfite determination with the nanobiosensor was also investigated. A linear concentration range of 0.9 to 400 JlM sulfite was achieved with the nanobiosensor and the minimum detectable sulfite concentration was 0.9 JlM (0.1 ppm). The successful application of the nanobiosensor to the determination of sulfite in wine and beers samples is reported. Chapter 4 further explores the use ofunsubstituted pyrrole for the development of a polypyrrole-tyrosinase (PPy-Tyr) potentiometric biosensor. The entrapment of tyrosinase into the polypyrrole film was carried out by galvanostatic polymerization on a platinum electrode. The optimum conditions for the formation of the PPy-Tyr film include a current density of 0.5 rnA cm·2 , a polymerization period of 150 seconds, 0.1 M pyrrole and 50 U mL-1 tyrosinase. Scanning electron microscopy (SEM), cyclic voltammetry (CV) and potentiometric measurements were used to confirm the presence of tyrosinase in the polymer film. The PPy-Tyr biosensor gave a sensitivity of 10 m VI J..LM for catechol, with a response time of 80 seconds and a linear concentration range of 1-50 J..LM catechol and the minimum detectable concentration was 1.0 J..LM. The PPy-Tyr biosensor was stable for at least 1 month when stored in a buffer at about 4 • C. The use of 3-substituted pyrroles for the fabrication of sulfite and phenol biosensor was explored in Part-III of this thesis by synthesizing 3-ethylpyrrole (EtPy). The final and intermediate products for the synthesis of EtPy were confirmed by fourier transform infrared (FTIR), nuclear magnetic resonance eH NMR and 13C NMR) spectroscopy and mass spectrometry (MS). The synthesized EtPy was employed in Chapter 5 for the fabrication of an amperometric sulfite biosensor. Sulfite oxidase was immobilized by galvanostatic polymerization of an adsorbed EtPy-SOx layer. The optimum conditions for the formation ofpoly(3-ethylpyrrole)-sulifte oxidase (PEtPy-SOx) biosensor include physical adsorption of 0.2 mg EtPy, 0.5 U of SOx, a current density of 0.75 rnA cm·2 and a polymerization period of 10 min. The PEtPy-SOx biosensor can detect as little as 0.1 J..LM sulfite and gave a linear concentration range of0.1-400 J..LM sulfte with a response time of 40 seconds. The electropolymerization of an adsorbed 3-ethylpyrrole-tyrosinase layer on a platinum electrode was investigated in Chapter 6 for the fabrication of a novel poly(3- ethylpyrrole)-tyrosinase (PEtPy-Tyr) based potentiometric catechol biosensor. The optimum conditions for the formation ofthe PEtPy-Tyr biosensor include 0.038 mg of3- ethylpyrrole (EtPy), 15 U of tyrosinase, a current density of 0.7 rnA cm·2 and a polymerization period of 15 min. The PEtPy-Tyr biosensor produced a reliable and sensitive response to catechol with a linear concentration range of 1-350 J!M, a minimum detectable catechol concentration of 0.1 J!M and a response time of 50 seconds. Chapter 7 concludes that both unsubstituted and substituted pyrroles (in this case 3-ethylpyrrole) are useful for fabrication ofbiosensors for sulfite and catechol. However, it was, noted that the use of the substituted pyrrole resulted in a substantial improvement of the sensitivity and minimum detectable concentration achieved for both sulfite and catechol biosensor. Also it resulted in a considerable reduction in the required enzyme concentrations

Endovascular treatment for acute ischemic stroke patients: implications and interpretation of IMS III, MR RESCUE, and SYNTHESIS EXPANSION trials: A report from the Working Group of International Congress of Interventional Neurology

OBJECTIVE The results of Interventional Management of Stroke (IMS) III, Magnetic Resonance and REcanalization of Stroke Clots Using Embolectomy (MR RESCUE), and SYNTHESIS EXPANSION trials are expected to affect the practice of endovascular treatment for acute ischemic stroke. The purpose of this report is to review the components of the designs and methods of these trials and to describe the influence of those components on the interpretation of trial results. METHODS A critical review of trial design and conduct of IMS III, MR RESCUE, and SYNTHESIS EXPANSION is performed with emphasis on patient selection, shortcomings in procedural aspects, and methodology of data ascertainment and analysis. The influence of each component is estimated based on published literature including multicenter clinical trials reporting on endovascular treatment for acute ischemic stroke and myocardial infarction. RESULTS We critically examined the time interval between symptom onset and treatment and rates of angiographic recanalization to differentiate between "endovascular treatment" and "parameter optimized endovascular treatment" as it relates to the IMS III, MR RESCUE, and SYNTHESIS EXPANSION trials. All the three trials failed to effectively test "parameter optimized endovascular treatment" due to the delay between symptom onset and treatment and less than optimal rates of recanalization. In all the three trials, the magnitude of benefit with endovascular treatment required to reject the null hypothesis was larger than could be expected based on previous studies. The IMS III and SYNTHESIS EXPANSION trials demonstrated that rates of symptomatic intracerebral hemorrhages subsequent to treatment are similar between IV thrombolytics and endovascular treatment in matched acute ischemic stroke patients. The trials also indirectly validated the superiority/equivalence of IV thrombolytics (compared with endovascular treatment) in patients with minor neurological deficits and those without large vessel occlusion on computed tomographic/magnetic resonance angiography. CONCLUSIONS The results do not support a large magnitude benefit of endovascular treatment in subjects randomized in all the three trials. The possibility that benefits of a smaller magnitude exist in certain patient populations cannot be excluded. Large magnitude benefits can be expected with implementation of "parameter optimized endovascular treatment" in patients with ischemic stroke who are candidates for IV thrombolytics

Thrombectomy 6 to 24 Hours after Stroke with a Mismatch between Deficit and Infarct

BACKGROUND: The effect of endovascular thrombectomy that is performed more than 6 hours after the onset of ischemic stroke is uncertain. Patients with a clinical deficit that is disproportionately severe relative to the infarct volume may benefit from late thrombectomy. METHODS: We enrolled patients with occlusion of the intracranial internal carotid artery or proximal middle cerebral artery who had last been known to be well 6 to 24 hours earlier and who had a mismatch between the severity of the clinical deficit and the infarct volume, with mismatch criteria defined according to age (<80 years or ≥80 years). Patients were randomly assigned to thrombectomy plus standard care (the thrombectomy group) or to standard care alone (the control group). The coprimary end points were the mean score for disability on the utility-weighted modified Rankin scale (which ranges from 0 [death] to 10 [no symptoms or disability]) and the rate of functional independence (a score of 0, 1, or 2 on the modified Rankin scale, which ranges from 0 to 6, with higher scores indicating more severe disability) at 90 days. RESULTS: A total of 206 patients were enrolled; 107 were assigned to the thrombectomy group and 99 to the control group. At 31 months, enrollment in the trial was stopped because of the results of a prespecified interim analysis. The mean score on the utility-weighted modified Rankin scale at 90 days was 5.5 in the thrombectomy group as compared with 3.4 in the control group (adjusted difference [Bayesian analysis], 2.0 points; 95% credible interval, 1.1 to 3.0; posterior probability of superiority, >0.999), and the rate of functional independence at 90 days was 49% in the thrombectomy group as compared with 13% in the control group (adjusted difference, 33 percentage points; 95% credible interval, 24 to 44; posterior probability of superiority, >0.999). The rate of symptomatic intracranial hemorrhage did not differ significantly between the two groups (6% in the thrombectomy group and 3% in the control group, P=0.50), nor did 90-day mortality (19% and 18%, respectively; P=1.00). CONCLUSIONS: Among patients with acute stroke who had last been known to be well 6 to 24 hours earlier and who had a mismatch between clinical deficit and infarct, outcomes for disability at 90 days were better with thrombectomy plus standard care than with standard care alone

Risk for Major Bleeding in Patients Receiving Ticagrelor Compared With Aspirin After Transient Ischemic Attack or Acute Ischemic Stroke in the SOCRATES Study (Acute Stroke or Transient Ischemic Attack Treated With Aspirin or Ticagrelor and Patient Outcomes)

Abstract: Background: Patients with minor acute ischemic stroke or transient ischemic attack are at high risk for subsequent stroke, and more potent antiplatelet therapy in the acute setting is needed. However, the potential benefit of more intense antiplatelet therapy must be assessed in relation to the risk for major bleeding. The SOCRATES trial (Acute Stroke or Transient Ischemic Attack Treated With Aspirin or Ticagrelor and Patient Outcomes) was the first trial with ticagrelor in patients with acute ischemic stroke or transient ischemic attack in which the efficacy and safety of ticagrelor were compared with those of aspirin. The main safety objective was assessment of PLATO (Platelet Inhibition and Patient Outcomes)\u2013defined major bleeds on treatment, with special focus on intracranial hemorrhage (ICrH). Methods: An independent adjudication committee blinded to study treatment classified bleeds according to the PLATO, TIMI (Thrombolysis in Myocardial Infarction), and GUSTO (Global Use of Strategies to Open Occluded Coronary Arteries) definitions. The definitions of ICrH and major bleeding excluded cerebral microbleeds and asymptomatic hemorrhagic transformations of cerebral infarctions so that the definitions better discriminated important events in the acute stroke population. Results: A total of 13 130 of 13 199 randomized patients received at least 1 dose of study drug and were included in the safety analysis set. PLATO major bleeds occurred in 31 patients (0.5%) on ticagrelor and 38 patients (0.6%) on aspirin (hazard ratio, 0.83; 95% confidence interval, 0.52\u20131.34). The most common locations of major bleeds were intracranial and gastrointestinal. ICrH was reported in 12 patients (0.2%) on ticagrelor and 18 patients (0.3%) on aspirin. Thirteen of all 30 ICrHs (4 on ticagrelor and 9 on aspirin) were hemorrhagic strokes, and 4 (2 in each group) were symptomatic hemorrhagic transformations of brain infarctions. The ICrHs were spontaneous in 6 and 13, traumatic in 3 and 3, and procedural in 3 and 2 patients on ticagrelor and aspirin, respectively. In total, 9 fatal bleeds occurred on ticagrelor and 4 on aspirin. The composite of ICrH or fatal bleeding included 15 patients on ticagrelor and 18 on aspirin. Independently of bleeding classification, PLATO, TIMI, or GUSTO, the relative difference between treatments for major/severe bleeds was similar. Nonmajor bleeds were more common on ticagrelor. Conclusions: Antiplatelet therapy with ticagrelor in patients with acute ischemic stroke or transient ischemic attack showed a bleeding profile similar to that of aspirin for major bleeds. There were few ICrHs. Clinical Trial Registration: URL: http://www.clinicaltrials.gov. Unique identifier: NCT01994720.Abstract: BACKGROUND: Patients with minor acute ischemic stroke or transient ischemic attack are at high risk for subsequent stroke, and more potent antiplatelet therapy in the acute setting is needed. However, the potential benefit of more intense antiplatelet therapy must be assessed in relation to the risk for major bleeding. The SOCRATES trial (Acute Stroke or Transient Ischemic Attack Treated With Aspirin or Ticagrelor and Patient Outcomes) was the first trial with ticagrelor in patients with acute ischemic stroke or transient ischemic attack in which the efficacy and safety of ticagrelor were compared with those of aspirin. The main safety objective was assessment of PLATO (Platelet Inhibition and Patient Outcomes)-defined major bleeds on treatment, with special focus on intracranial hemorrhage (ICrH). METHODS: An independent adjudication committee blinded to study treatment classified bleeds according to the PLATO, TIMI (Thrombolysis in Myocardial Infarction), and GUSTO (Global Use of Strategies to Open Occluded Coronary Arteries) definitions. The definitions of ICrH and major bleeding excluded cerebral microbleeds and asymptomatic hemorrhagic transformations of cerebral infarctions so that the definitions better discriminated important events in the acute stroke population. RESULTS: A total of 13 130 of 13 199 randomized patients received at least 1 dose of study drug and were included in the safety analysis set. PLATO major bleeds occurred in 31 patients (0.5%) on ticagrelor and 38 patients (0.6%) on aspirin (hazard ratio, 0.83; 95% confidence interval, 0.52-1.34). The most common locations of major bleeds were intracranial and gastrointestinal. ICrH was reported in 12 patients (0.2%) on ticagrelor and 18 patients (0.3%) on aspirin. Thirteen of all 30 ICrHs (4 on ticagrelor and 9 on aspirin) were hemorrhagic strokes, and 4 (2 in each group) were symptomatic hemorrhagic transformations of brain infarctions. The ICrHs were spontaneous in 6 and 13, traumatic in 3 and 3, and procedural in 3 and 2 patients on ticagrelor and aspirin, respectively. In total, 9 fatal bleeds occurred on ticagrelor and 4 on aspirin. The composite of ICrH or fatal bleeding included 15 patients on ticagrelor and 18 on aspirin. Independently of bleeding classification, PLATO, TIMI, or GUSTO, the relative difference between treatments for major/severe bleeds was similar. Nonmajor bleeds were more common on ticagrelor. CONCLUSIONS: Antiplatelet therapy with ticagrelor in patients with acute ischemic stroke or transient ischemic attack showed a bleeding profile similar to that of aspirin for major bleeds. There were few ICrHs

Global Incidence and Risk Factors Associated With Postoperative Urinary Retention Following Elective Inguinal Hernia Repair

Importance Postoperative urinary retention (POUR) is a well-recognized complication of inguinal hernia repair (IHR). A variable incidence of POUR has previously been reported in this context, and contradictory evidence surrounds potential risk factors.Objective To ascertain the incidence of, explore risk factors for, and determine the health service outcomes of POUR following elective IHR.Design, Setting, and Participants The Retention of Urine After Inguinal Hernia Elective Repair (RETAINER I) study, an international, prospective cohort study, recruited participants between March 1 and October 31, 2021. This study was conducted across 209 centers in 32 countries in a consecutive sample of adult patients undergoing elective IHR.Exposure Open or minimally invasive IHR by any surgical technique, under local, neuraxial regional, or general anesthesia.Main Outcomes and Measures The primary outcome was the incidence of POUR following elective IHR. Secondary outcomes were perioperative risk factors, management, clinical consequences, and health service outcomes of POUR. A preoperative International Prostate Symptom Score was measured in male patients.Results In total, 4151 patients (3882 male and 269 female; median [IQR] age, 56 [43-68] years) were studied. Inguinal hernia repair was commenced via an open surgical approach in 82.2% of patients (n = 3414) and minimally invasive surgery in 17.8% (n = 737). The primary form of anesthesia was general in 40.9% of patients (n = 1696), neuraxial regional in 45.8% (n = 1902), and local in 10.7% (n = 446). Postoperative urinary retention occurred in 5.8% of male patients (n = 224), 2.97% of female patients (n = 8), and 9.5% (119 of 1252) of male patients aged 65 years or older. Risk factors for POUR after adjusted analyses included increasing age, anticholinergic medication, history of urinary retention, constipation, out-of-hours surgery, involvement of urinary bladder within the hernia, temporary intraoperative urethral catheterization, and increasing operative duration. Postoperative urinary retention was the primary reason for 27.8% of unplanned day-case surgery admissions (n = 74) and 51.8% of 30-day readmissions (n = 72).Conclusions The findings of this cohort study suggest that 1 in 17 male patients, 1 in 11 male patients aged 65 years or older, and 1 in 34 female patients may develop POUR following IHR. These findings could inform preoperative patient counseling. In addition, awareness of modifiable risk factors may help to identify patients at increased risk of POUR who may benefit from perioperative risk mitigation strategies