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Objective: The aim of this study was to characterize early urinary gene expression differences between patients with sepsis and patients with sterile inflammation and summarize in terms of a reproducible sepsis probability score. Design: This was a prospective observational cohort study. Setting: The study was conducted in a quaternary care academic hospital. Patients: One hundred eighty-six sepsis patients and 78 systemic inflammatory response syndrome (SIRS) patients enrolled between January 2015 and February 2018. Interventions: Whole-genome transcriptomic analysis of RNA was extracted from urine obtained from sepsis patients within 12 hours of sepsis onset and from patients with surgery-acquired SIRS within 4 hours after major inpatient surgery. Measurements and Main Results: We identified 422 of 23,956 genes (1.7%) that were differentially expressed between sepsis and SIRS patients. Differentially expressed probes were provided to a collection of machine learning feature selection models to identify focused probe sets that differentiate between sepsis and SIRS. These probe sets were combined to find an optimal probe set (UrSepsisModel) and calculate a urinary sepsis score (UrSepsisScore), which is the geometric mean of downregulated genes subtracted from the geometric mean of upregulated genes. This approach summarizes the expression values of all decisive genes as a single sepsis score. The UrSepsisModel and UrSepsisScore achieved area under the receiver operating characteristic curves 0.91 (95% confidence interval, 0.86-0.96) and 0.80 (95% confidence interval, 0.70-0.88) on the validation cohort, respectively. Functional analyses of probes associated with sepsis demonstrated metabolic dysregulation manifest as reduced oxidative phosphorylation, decreased amino acid metabolism, and decreased oxidation of lipids and fatty acids. Conclusions: Whole-genome transcriptomic profiling of urinary cells revealed focused probe panels that can function as an early diagnostic tool for differentiating sepsis from sterile SIRS. Functional analysis of differentially expressed genes demonstrated a distinct metabolic dysregulation signature in sepsis.

Oral squamous cell carcinomas are mostly preceded by precancerous lesions such as leukoplakia and erythroplakia. Our study is aimed at identifying potential biomarker proteins in precancerous lesions of leukoplakia and erythroplakia that can flag their transformation to oral cancer. Four biological replicate samples from clinical phenotypes of healthy control, leukoplakia, erythroplakia, and oral carcinoma were annotated based on clinical screening and histopathological evaluation of buccal mucosa tissue. Differentially expressed proteins were delineated using a label-free quantitative proteomic experiment done on an Orbitrap Fusion Tribrid mass spectrometer in three technical replicate sets of samples. Raw files were processed using MaxQuant version 2.0.1.0, and downstream analysis was done via Perseus version 1.6.15.0. Validation included functional annotation based on biological processes and pathways using the ClueGO plug-in of Cytoscape. Hierarchical clustering and principal component analysis were performed using the ClustVis tool. Across control, leukoplakia, and cancer, L-lactate dehydrogenase A chain, plectin, and WD repeat-containing protein 1 were upregulated, whereas thioredoxin 1 and spectrin alpha chain, nonerythrocytic 1 were downregulated. Across control, erythroplakia, and cancer, L-lactate dehydrogenase A chain was upregulated whereas aldehyde dehydrogenase 2, peroxiredoxin 1, heat shock 70 kDa protein 1B, and spectrin alpha chain, nonerythrocytic 1 were downregulated. We found that proteins involved in leukoplakia were associated with alteration in cytoskeletal disruption and glycolysis, while in erythroplakia, they were associated with alteration in response to oxidative stress and glycolysis across phenotypes. Hierarchical clustering subgrouped half of precancerous samples under the main branch of the control and the remaining half under carcinoma. Similarly, principal component analysis identified segregated clusters of control, precancerous lesions, and cancer, but erythroplakia phenotypes, in particular, overlapped more with the cancer cluster. Qualitative and quantitative protein signatures across control, precancer, and cancer phenotypes explain possible functional outcomes that dictate malignant transformation to oral carcinoma.

Reactions requiring controlled delivery of protons and electrons are important in storage of energy in small molecules. While control over proton transfer can be achieved by installing appropriate chemical functionality in the catalyst, control of electron-transfer (ET) rates can be achieved by utilizing self-assembled monolayers (SAMs) on electrodes. Thus, a deeper understanding of the ET through SAM to an immobilized or covalently attached redox-active species is desirable. Long-range ET across several SAM-covered Au electrodes to covalently attached ferrocene is investigated using protonated and deuterated thiols (R-SH/R-SD). The rate of tunneling is measured using both chronoamperometry and cyclic voltammetry, and it shows a prominent kinetic isotope effect (KIE). The KIE is ∼2 (normal) for medium-chain-length thiols but ∼0.47 (inverse) for long-chain thiols. These results imply substantial contribution from the classical modes at the Au-(H)SR interface, which shifts substantially upon deuteration of the thiols, to the ET process. The underlying H/D KIE of these exchangeable thiol protons should be considered when analyzing solvent isotope effects in catalysis utilizing SAM.

Background: Diabetic retinopathy (DR) is a microvascular complication as a consequence of prolonged hyperglycemia and other risk factors. It is seen in both types of diabetes mellitus (DM) and can lead to progressive vision loss. Many prevalence studies were undertaken in the past, but the eastern part of India was less explored. Aim: (1) The aim of this study is to determine the prevalence of DR among both types of DM patients in an institution-based population of Kolkata. (2) To compare the clinic-biochemical profiles of patients with and without DR. Materials and Methods: From January 2019 to December 2019, 313 patients with a history of DM were screened for DR using slit-lamp bio microscopy with +90D lens after dilating the pupils. Classification of DR was done on the basis of International Clinical classification of DR. Metabolic profile of all participants was noted. Results: The overall prevalence of DR was 16.60% (14.37% nonproliferative and 2.23% proliferative). The prevalence was 80% and 15.58% among Type 1 and 2 DM patients, respectively. Diabetic macular edema was observed in 2.54% of cases. Highest prevalence (38.46%) was observed in patients suffering for more than 20 years of DM. The duration of DM was significantly higher among DR patients (9.46 ± 5.23 years vs. 7.34 ± 5.57; P = 0.012). Patients with DR also displayed significantly higher levels of glycated hemoglobin, creatinine, and albumin-creatinine ratio in comparison to the patients without DR. Conclusion: The prevalence of DR from an institution of Eastern India is comparable to other institutional studies conducted in the various parts of the country. The prevalence is higher among Type 1 DM patients. Nonproliferative DR is the most common type. Comparatively, DR patients had a longer duration of DM. Glycemic control and renal statuses are poor in DR than non-DR patients.

In the absence of a sensitive and specific diagnostic modality capable of detecting all forms of tuberculosis (TB), proteomics may identify specific Mycobacterium tuberculosis (M.tb) proteins in urine, with a potential as biomarkers. To identify candidate biomarkers for TB, proteome profile of urine from pulmonary TB patients was compared with non-disease controls (NDC) and disease controls (DC, Streptococcus pneumonia infected patients) using a combination of two-dimensional difference gel electrophoresis (2D-DIGE) and liquid chromatography tandem mass spectrometry (LCMS/MS). Eleven differentially expressed host proteins and Eighteen high abundant M.tb proteins were identified. Protein-protein interactome (PPI) and functional enrichment analyses like Gene Ontologies, Reactome pathway etc. demonstrated that the human proteins mainly belong to extracellular space and show physiological pathways for immune response and hematological disorders. Whereas, M.tb proteins belong to the cell periphery, plasma membrane and cell wall, and demonstrated catalytic, nucleotide binding and ATPase activities along with other functional processes. The study findings provide valuable inputs about the biomarkers of TB and shed light on the probable disease consequences as an outcome of the bacterial pathogenicity.

OBJECTIVE: This review assimilates and critically evaluates available literature regarding the use of metabolomic profiling in surgical decision-making. SUMMARY BACKGROUND DATA: Metabolomic profiling is performed by nuclear magnetic resonance spectroscopy or mass spectrometry of biofluids and tissues to quantify biomarkers (i.e. sugars, amino acids, and lipids), producing diagnostic and prognostic information that has been applied among patients with cardiovascular disease, inflammatory bowel disease, cancer, and solid organ transplants. METHODS: PubMed was searched from 1995–2019 to identify studies investigating metabolomic profiling of surgical patients. Articles were included and assimilated into relevant categories per PRISMA-ScR guidelines. Results were summarized with descriptive analytical methods. RESULTS: Forty-seven studies were included, most of which were retrospective studies with small sample sizes using various combinations of analytic techniques and types of biofluids and tissues. Results suggest that metabolomic profiling has potential to effectively screen for surgical diseases, suggest diagnoses, and predict outcomes such as postoperative complications and disease recurrence. Major barriers to clinical adoption include a lack of high-level evidence from prospective studies, heterogeneity in study design regarding tissue and biofluid procurement and analytical methods, and the absence of large, multicenter metabolome databases to facilitate systematic investigation of the efficacy, reproducibility, and generalizability of metabolomic profiling diagnoses and prognoses. CONCLUSIONS: Metabolomic profiling research would benefit from standardization of study design and analytic approaches. As technologies improve and knowledge garnered from research accumulates, metabolomic profiling has potential to provide personalized diagnostic and prognostic information to support surgical decision-making from preoperative to post-discharge phases of care.

The complex and heterogeneous pathophysiology of schizophrenia can be deconstructed by integration of large-scale datasets encompassing genes through behavioral phenotypes. Genome-wide datasets are now available for genetic, epigenetic and transcriptomic variations in schizophrenia, which are then analyzed by newly devised systems biology algorithms. A missing piece, however, is the inclusion of information on the proteome and its dynamics in schizophrenia. Proteomics has lagged behind omics of the genome, transcriptome and epigenome since analytic platforms were relatively less robust for proteins. There has been remarkable progress, however, in the instrumentation of liquid chromatography (LC) and mass spectrometry (MS) (LCMS), experimental paradigms and bioinformatics of the proteome. Here, we present a summary of methodological innovations of recent years in MS based proteomics and the power of new generation proteomics, review proteomics studies that have been conducted in schizophrenia to date, and propose how such data can be analyzed and integrated with other omics results. The function of a protein is determined by multiple molecular properties, i.e., subcellular localization, posttranslational modification (PTMs) and protein-protein interactions (PPIs). Incorporation of these properties poses additional challenges in proteomics and their integration with other omics; yet is a critical next step to close the loop of multi-omics integration. In sum, the recent advent of high-throughput proteome characterization technologies and novel mathematical approaches enable us to incorporate functional properties of the proteome to offer a comprehensive multi-omics based understanding of schizophrenia pathophysiology.

The Clock Drawing Test (CDT) is an inexpensive tool to screen for dementia. In this study, we examined if a semi-supervised deep learning (DL) system using Variational Autoencoder (VAE) can extract atypical clock features from a large dataset of unannotated CDTs (n=13,580) and use them to classify dementia (n=18) from non-dementia (n=20) peers. The classification model built with VAE latent space features adequately classified dementia from non-dementia (0.78 Area Under Receiver Operating Characteristics (AUROC)). The VAE-identified atypical clock features were then reviewed by domain experts and compared with existing literature on clock drawing errors. This study shows that a semi-supervised deep learning (DL) analysis of the CDT can extract important clock drawing anomalies that are predictive of dementia.

Supplemental Digital Content is available in the text., Objectives: Identify alterations in gene expression unique to systemic and kidney-specific pathophysiologic processes using whole-genome analyses of RNA isolated from the urinary cells of sepsis patients. Design: Prospective cohort study. Setting: Quaternary care academic hospital. Patients: A total of 266 sepsis and 82 control patients enrolled between January 2015 and February 2018. Interventions: Whole-genome transcriptomic analysis of messenger RNA isolated from the urinary cells of sepsis patients within 12 hours of sepsis onset and from control subjects. Measurements and Main Results: The differentially expressed probes that map to known genes were subjected to feature selection using multiple machine learning techniques to find the best subset of probes that differentiates sepsis from control subjects. Using differential expression augmented with machine learning ensembles, we identified a set of 239 genes in urine, which show excellent effectiveness in classifying septic patients from those with chronic systemic disease in both internal and independent external validation cohorts. Functional analysis indexes disrupted biological pathways in early sepsis and reveal key molecular networks driving its pathogenesis. Conclusions: We identified unique urinary gene expression profile in early sepsis. Future studies need to confirm whether this approach can complement blood transcriptomic approaches for sepsis diagnosis and prognostication.

The extent of heme extraction from myoglobin (Mb) by methylethyl ketone is found to be pH dependent and show three distinct phases. Parallel investigations of the protein using resonance Raman (rR) and circular dichroism (CD) across these pH regions indicate that these phases correspond to three different protonation steps in holoMb as the pH of the solution changed. The first transition occurs between pH5-6 and is due to the protonation of one of the heme propionate groups which disrupts its H-bonding with Arg 45 in the loop. The 2nd phase (pH5-4) likely involves the protonation of the 2nd propionate which H-bonds to Ser 92 in the F-helix. The third phase (pH

Temperature is one of the basic biophysical quantity monitored for various biomedical systems. Moreover, the variation of temperature is also an important parameter which can be used for estimating other measurands, such as respiratory airflow. This paper proposes a simple operational amplifier-based astable multivibrator circuit for linearization of the characteristic of a negative temperature coefficient thermistor constituting one of the timing resistors. The circuit has been combined with a lookup table to get the unknown temperature value from multivibrator output. Moreover, the same system topology can be used as a linearizer for measurement of respiratory airflow. The performance of the composite system has been verified experimentally. A linearity of approximately ±0.75% has been achieved over 30 °C–110 °C in the case of temperature measurement and ±1.2% for airflow of 10–60 LPM. Better results can also be achieved with the introduction of interpolation algorithms, but at a higher computational and component cost. The compactness of the complete system makes it a good candidate for embedded sensing applications in biomedical systems, such as point-of-care monitoring or in sleep study.

Ammonium tetrathiomolybdate (ATM) is found to be a novel electrode material forming self-assembled adlayers on Au electrodes which are robust and stable in aqueous environments. Iron–porphyrin catalysts can be physiadsorbed on this electrode material and these modified electrodes are quite stable during hydrodynamic electrochemical experiments unlike self-assembled monolayers (SAMs) of thiols on Au/Ag. The rate of interfacial charge transfer can be tuned by controlling the height of the ATM assembly by varying the deposition time. These electrodes allow convenient tuning of the electrokinetics of 4e−/4H+ O2 reduction from mass transfer to charge transfer with the same electrode material.

Purpose: To evaluate the efficacy, safety, and steroid-sparing effect of topical cyclosporine A (Cs A) 0.05% in patients with moderate to severe steroid dependent vernal keratoconjunctivitis (VKC). Methods: A prospective, comparative, placebo controlled study was carried out on 68 VKC patients, with 34 patients treated with topical Cs A 0.05% and the remaining 34 with topical carboxymethyl cellulose 0.5% (placebo). Both groups also received topical loteprednol etabonate 0.5%. Symptom (itching, photophobia, tearing, and discharge) score, sign (tarsal and limbal papillae, corneal involvement, and conjunctival hyperemia) score, and drug score (steroid drop usage/day/eye) were recorded at baseline and each follow-up visit. The intraocular pressure (IOP) measurement and evaluation of any ocular side effects were carried out. Results: Significant reduction in symptom score and sign score was seen in both groups. Cs A group significantly showed more reduction in symptom (P < 0.0001 in all follow-up visits) and sign (P < 0.0001 in all follow-up visits) scores compared to the placebo group. At day 7, mean steroid usage reduced from 4 to 3.44 ± 0.5 and 3.79 ± 0.4 in Cs A and placebo groups, respectively (P < 0.0001). Steroid drops completely stopped in 21 patients at day 60 in the Cs A group compared to none in the placebo group. No significant rise in IOP or any side effects were noted in either group. Conclusion: Topical Cs A 0.05% is effective and safe in patients with moderate to severe VKC with good steroid-sparing effect.

Controlled three-dimensional positioning of nanoparticles is achieved by delivering single fluorescent nanoparticles from a nanopipette and capturing them at well-defined regions of an electrified substrate. To control the position of single nanoparticles, the force of the pressure-driven flow from the pipette is balanced by the attractive electrostatic force at the substrate, providing a strategy by which nanoparticle trajectories can be manipulated in real time. To visualize nanoparticle motion, a resistive-pulse electrochemical setup is coupled with an optical microscope, and nanoparticle trajectories are tracked in three dimensions using super-resolution fluorescence imaging to obtain positional information with precision in the tens of nanometers. As the particles approach the substrate, the diffusion kinetics are analyzed and reveal either subdiffusive (hindered) or superdiffusive (directed) motion depending on the electric field at the substrate and the pressure-driven flow from the pipette. By balancing the effects of the forces exerted on the particle by the pressure and electric fields, controlled, real-time manipulation of single nanoparticle trajectories is achieved. The developed approach has implications for a variety of applications such as surface patterning and drug delivery using colloidal nanoparticles.

Introduction: Peripapillary retinal nerve fiber layer (RNFL) thickness analysis is a subjective method of analysis of glaucomatous damage. As almost 50% of retinal ganglion cells are located in the macula, assessment of macular thickness can be an alternative method for diagnosis of glaucoma. Objectives: To evaluate the changes in macular and retinal nerve fiber layer thickness in controls, glaucoma suspects and glaucoma patients using time domain optical coherence tomography (TD-OCT). Materials and methods: Macular and peripapillary RNFL scans were performed in one eye of 70 controls, 35 glaucoma suspects and 70 glaucoma patients by TD-OCT. The discriminating power of each parameter between the groups was determined by area under the receiver operating characteristic (AROC) curve. The correlation of macular thickness and RNFL thickness parameters with global field indices were also performed. P-value of < 0.05 was considered statistically significant. Results: The differences in all the macular thickness parameters between the groups were statistically significant (p< 0.05) except foveal thickness (FT) and nasal inner (NI) quadrant thickness. The temporal outer (TO) macular quadrant produced largest AROC curve of 0.90 between controls and glaucoma patients. The differences in all the RNFL thickness parameters were highly significant between the groups (p

Using a combination of self-assembly and synthesis, bioinspired electrodes having dilute iron porphyrin active sites bound to axial thiolate and imidazole axial ligands are created atop self-assembled monolayers (SAMs). Resonance Raman data indicate that a picket fence architecture results in a high-spin (HS) ground state (GS) in these complexes and a hydrogen-bonding triazole architecture results in a low-spin (LS) ground state. The reorganization energies (λ) of these thiolate- and imidazole-bound iron porphyrin sites for both HS and LS states are experimentally determined. The λ of 5C HS imidazole and thiolate-bound iron porphyrin active sites are 10-16 kJ/mol, which are lower than their 6C LS counterparts. Density functional theory (DFT) calculations reproduce these data and indicate that the presence of significant electronic relaxation from the ligand system lowers the geometric relaxation and results in very low λ in these 5C HS active sites. These calculations indicate that loss of one-half a π bond during redox in a LS thiolate bound active site is responsible for its higher λ relative to a σ-donor ligand-like imidazole. Hydrogen bonding to the axial ligand leads to a significant increase in λ irrespective of the spin state of the iron center. The results suggest that while the hydrogen bonding to the thiolate in the 5C HS thiolate bound active site of cytochrome P450 (cyp450) shifts the potential up, resulting in a negative ΔG, it also increases λ resulting in an overall low barrier for the electron transfer process.

Investigation of the proton coupled electron transfer steps involved in electrocatalytic ORR by hemin modified electrodes allows the determination of pKas of the proximal and distal oxygen atoms of a rate determining FeIII–OOH species.

Purpose: The purpose of this study is to study the pattern of uveitis at a tertiary care hospital in Eastern India. Materials and Methods: This is a prospective study of new cases of uveitis attending our tertiary care hospital in Kolkata, Eastern India, from July 1, 2016, to June 30, 2019. Data regarding patient demographic profile, clinical features, anatomical locations, and etiological diagnosis were evaluated. Results: Among 331 patients, 142 (42.9%) were male and 189 (57.1%) were female (P

A method of tuning the apparent formal potential at the interface between the electrode surface covered with carboxylate thiol self assembled monolayer (SAM) and the solution containing electrolytes is reported. Redox active ferrocene moiety has been covalently attached to the terminal azido groups in the SAM using “click” chemistry. Cyclic voltammetry experiments show that the reduction potential of this ferrocene moiety can be tuned easily from 0.345 V to 0.200 V, i.e. by 145 mV by varying the chain length of the thiol carboxylic acid diluents, changing the pH of the electrolytic solution and by changing the coverage of the surface. Using the (applying the theory of interfacial potential distribution by Smith and White and including the Stern layer effect on the potential of the redox species at the interface proposed by Fawcett) modified Nernst equation the shift of apparent formal potential of redox species at the interface has been analyzed when the chain length of the diluents, pH of the solution and coverage of the surface are changed. The data indicate that the phenomenon is governed by the ionizable polar head groups SAM, i.e. COOH which changes the interfacial microenvironment around the redox active ferrocene centre.

Electrodes bearing thiolate and imidazole coordinated iron porphyrin catalysts are constructed and characterized using resonance Raman spectroscopy, absorption spectroscopy, and electrochemistry. The cyclic voltammetry data and their pH dependences are used to establish the nature of the exchangeable trans ligands in both of these cases. In situ monitoring of partially reduced oxygen species (PROS) produced during O(2) reduction using rotating ring disc electrochemistry (RRDE) experiments provide direct insight into the "push-effect" of the thiolate ligand. The thiolate bound iron porphyrin electrode generates highly oxidizing species on the electrode during electrocatalytic O(2) reductions which are very reactive. These surfaces can utilize these oxidants to catalytically hydroxylate strong C-H bonds using molecular O(2) with turnover numbers as high as 200.

Objectives: To assess and compare the analgesic and anti inflammatory efficacy of nepafenac and prednisolone eye drops used individually and in combination in post-operative period of cataract surgery by estimating anterior chamber cell count(ACCC) and pain in visual analogue scale(VAS). Materials and Methods: A total of 156 subjects after cataract surgery were allocated through lottery to 3 groups (n=52 each). Group A: with topical nepafenac: 1 drop thrice daily for 3 weeks, Group B with topical prednisolone: 1 drop four times daily for 3 weeks. Group C with combination of nepafenac and prednisolone as per the schedule for group A and B respectively. Drugs were continued for 22 consecutive days and there were four follow-up visits on day 4, day 8, day 15 and day 22. Anterior chamber cell count (ACCC) and pain were determined by slit lamp bio microscope and visual analogue scale respectively. Results: End results of group A for ACCC and pain were 5.41 ± 0.44 and 1.27 ± 0.41 respectively. In case of group B, ACCC and pain were 1.09 ± 0.32 and 0. Group C showed the best results where for ACCC and pain were 0.40 ± 0.22 and 0 respectively. Conclusion: Combination of Prednisolone and nepafenac eye drops was the most efficacious in reducing inflammation and pain after cataract surgery.

To compare aqueous angiogenic and inflammatory cytokine concentrations in different patterns of diabetic macular edema (DME) based on optical coherence tomography (OCT). This prospective study was conducted between July 1, 2015, and March 31, 2016, for 9 months. Aqueous samples were obtained from 52 consecutive DME patients and 16 controls. DME patients were divided according to OCT patterns into diffuse retinal thickening (DRT; n = 17), cystoid macular edema (CME; n = 20) and serous retinal detachment (SRD; n = 15) groups. Interleukin (IL)-6, IL-8, vascular endothelial growth factor (VEGF) and tumor necrosis factor alpha (TNF-α) levels were measured by RayBio(R) Human ELISA Kit. IL-6, IL-8 and VEGF levels differed significantly between three DME groups (p

Purpose: To study the demographic profile, severity and causes of visual impairment among registered patients in a tertiary care hospital in north Kolkata, eastern India, and to assess the magnitude of under-registration in that population. Methods: This is a retrospective analytical study. A review of all visually impaired patients registered at our tertiary care hospital during a ten-year period from January 2005 to December 2014, which is entitled for certification of people of north Kolkata, eastern India (with a population denominator of 1.1 million), was performed. Overall, 2472 eyes of 1236 patients were analyzed in terms of demographic characteristics, cause of visual impairment, and percentage of visual disability as per the guidelines established by the government of India. Results: Male patients (844, 68.28%; 95% confidence interval [CI], 65.69-70.87) registered more often than female patients (392; 31.72%, P = 0.0004). The registration rate for visual impairment was 11.24 per 100,000 per annum; this is not the true incidence rate, as both new patients and those visiting for renewal of certification were included in the study. Optic atrophy was the most common cause of visual impairment (384 eyes, 15.53%; 95% CI, 14.1-16.96). Conclusion: Commonest cause of visual impairment was optic atrophy followed by microphthalmos. Under-registration is a prevalent problem as the registration system is voluntary rather than mandatory, and female patients are more likely to be unregistered in this area.

The protonation state of thiols in self-assembled monolayers (SAMs) on Ag and Au surfaces and nanoparticles (NPs) has been an issue of contestation. It has been recently demonstrated that deuterating the thiol proton produces ostentatious changes in the Raman spectra of thiols and can be used to detect the presence of the thiol functional group. Surface enhanced Raman spectroscopy (SERS) of H/D substituted aliphatic thiols on Ag surfaces clearly shows the presence of S-H vibration between 2150-2200 cm(-1) which shifts by 400 cm(-1) upon deuteration and a simultaneous >20 cm(-1) shift in the C-S vibration of thiol deuteration. Large shifts (>15 cm(-1)) in the C-S vibration are also observed for alkyl thiol SAMs on Au surfaces. Alternatively, neither the S-H vibration nor the H/D isotope effect on the C-S vibration is observed for alkyl thiol SAMs on Ag/Au NPs. XPS data on Ag/Au surfaces bearing aliphatic thiol SAMs show the presence of both protonated and deprotonated thiols while on Ag/Au NPs only deprotonated thiols are detected. These data suggest that aliphatic thiol SAMs on Au/Ag surfaces are partially protonated whereas they are totally deprotonated on Au/Ag NPs. Aromatic PhSH SAMs on Ag/Au surfaces and Ag/Au NPs do not show these vibrations or H/D shifts as well indicating that the thiols are deprotonated at these interfaces.

An iron porphyrin with a pre-organized hydrogen bonding (H-Bonding) distal architecture is utilized to avoid the inherent loss of entropy associated with H-Bonding from solvent (water) and mimic the behavior of metallo-enzyme active sites attributed to H-Bonding interactions of active site with the 2nd sphere residues. Resonance Raman (rR) data on these iron porphyrin complexes indicate that H-Bonding to an axial ligand like hydroxide can result in both stronger or weaker Fe(III)-OH bond relative to iron porphyrin complexes. The 6-coordinate (6C) complexes bearing water derived axial ligands, trans to imidazole or thiolate axial ligand with H-Bonding stabilize a low spin (LS) ground state (GS) when a complex without H-Bonding stabilizes a high spin (HS) ground state. DFT calculations reproduce the trend in the experimental data and provide a mechanism of how H-Bonding can indeed lead to stronger metal ligand bonds when the axial ligand donates an H-Bond and lead to weaker metal ligand bonds when the axial ligand accepts an H-Bond. The experimental and computational results explain how a weak Fe(III)-OH bond (due to H-Bonding) can lead to the stabilization of low spin ground state in synthetic mimics and in enzymes containing iron porphyrin active sites. H-Bonding to a water ligand bound to a reduced ferrous active site can only strengthen the Fe(II)-OH2 bond and thus exclusion of water and hydrophilic residues from distal sites of O2 binding/activating heme proteins is necessary to avoid inhibition of O2 binding by water. These results help demonstrate the predominant role played by H-Bonding and subtle changes in its orientation in determining the geometric and electronic structure of iron porphyrin based active sites in nature.