PubMed Health⌕ Search

Biomedical subjects

R S Phadke

Publications and source records attributed to R S Phadke.

At least 19 recordsLinked to original sources
PubMed2001

Substance P (free acid) adopts different conformation than native peptide in DMSO, water and DPPC bilayers.

The conformation of substance P (free acid) (SPOH) has been investigated in dimethylsulfoxide (DMSO), water and dipalmitoylphosphotidylcholine (DPPC) bilayers by two-dimensional NMR and restraint molecular dynamics simulations. The observed NOE patterns for SPOH in these media are very much different from each other. Molecular modeling of the conformation of SPOH by incorporating NOEs as distance restraints shows wide differences in its conformation in three media. The main structural features for SPOH in DMSO are y-bends at Pro4 and Phe7 along with a non-specific bend around Lys3-Pro4-Gln5-Gln6, which are stabilized by Lys3CO-->Gln5NH, Gln6CO-->Phe8NH hydrogen bonding. The more flexible conformation of SPOH in water is transformed to an ordered structure after incorporation in DPPC bilayers. The conformation of SPOH in DPPC bilayers is characterized by gamma-bends at Pro4, Gln6 and Phe7, which are stabilized by hydrogen bonding between Lys3CO-->Gln5NH, Gln5CO-->Phe7NH and Gln6CO-->Phe8NH, respectively. The absence of biological activity in SPOH has been attributed to the absence of any helix like structure at the central residues and absence of any interresidue interaction with C-terminal OH group, in DPPC bilayers, a feature shown to be an important prerequisite for SP and SP agonists to bind to the NKI tachykinin receptor.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed2000

A new membrane probing steroidal spin label: synthesis and applications.

The applicability of a new steroidal spin label, 3-oxo-androstan-17 beta-yl-(2",2",6",6"-tetramethyl-N-oxyl) piperidyl butan-1',4'-dioate, in studying the phase transition properties of model membrane L-alpha-dipalmitoyl phosphatidyl choline (DPPC) in the presence and absence of drugs has been explored. Its synthesis and characterization has been described herein. Besides, the localization of this spin label in lipid liposomes has been studied using electron spin resonance (ESR), differential scanning calorimetry (DSC) and 1H and 31P NMR spectroscopic techniques. The label has also been used to study the permeability of epinephrine into membrane. The results show that the spin label has a good potential as a spin probe in the study of biomembranes.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1999

Interaction of 7-hydroxy-8-(phenylazo)1,3-naphthalenedisulfonate with bovine plasma albumin. Spectroscopic studies.

Interaction of Orange G (OG) with bovine plasma albumin (BPA) has been investigated using NMR, UV-visible absorption, CD, and fluorescence techniques. The bound conformation of OG is a compact structure with N9-N10 bond in a non-planar syn conformation. The binding causes a decrease in the 478-nm absorption band of OG. The analysis of the binding isotherm generated from UV-visible absorption measurements gives a dissociation constant of 10 microM and stoichiometry 1:1 for BPA.OG complex. Dissociation constant is invariant in the pH range 5.0-8.0 and is approximately 20 times higher at pH 4.0 than its value at pH 7.0. Near and far UV-CD studies indicate alterations in the helical content and in the tertiary structure of the protein on complexation. The binding induces (-) and (+) CD at 335 nm and 465 nm, respectively. The binding also results into an increase in the steady state fluorescence anisotropy of OG without affecting emission maximum and quantum yield. Fluorescence data indicate that quenching of Trp fluorescence by OG is static in nature and OG selectively binds near Trp-135. Observation of similar rotational correlation time for BPA and BPA.OG complex indicates that the overall globular structure of BPA remains unaltered on binding despite certain internal rearrangement in the protein structure.

Animals↗
PubMed1999

Identification of low-molecular-weight compounds in goat epididymis using multinuclear nuclear magnetic resonance.

Multinuclear nuclear magnetic resonance (NMR) (1H, 13C, and 31P) studies have been performed on aqueous solutions of lyophilysates of cell-free extract, epididymal fluid, and intact cells from caput and cauda regions of epididymis of sacrificed goats. Identification of low-molecular-weight compounds present in different maturation phases of spermatozoa has been carried out. Several low-molecular-weight compounds have been identified by assigning 600 MHz 1H NMR spectra with the help of two-dimensional homonuclear and heteronuclear correlation spectroscopy such as double quantum filtered correlation spectroscopy and heteronuclear single quantum correlation spectroscopy. Homonuclear coupling constants have also been used to get unambiguous assignments of resonances. NMR data were compared with those of standard samples measured at same pH and with those reported in the literature. Identification of several amino acids, carbohydrates, and lipids have been made and their presence has been discussed in relation to their relevance to sperm functions. The presence of beta-alanine and hypotaurine has been reported for the first time in goat epididymis.

Animals↗
PubMed1999

Replacement of phe(8) in substance P by tyr (Tyr(8)-SP) alters the conformation of the peptide in DMSO, water, and lipid bilayers.

The conformation of [Tyr(8)]SP (Y8SP) in dimethylsulfoxide (DMSO), water, and dipalmitoyl phosphatidylcholine (DPPC) bilayers has been investigated by two-dimensional nmr and molecular dynamics simulations. Molecular modeling of the conformation of Y8SP by incorporating nuclear Overhauser effects as distance restraints shows wide differences in its conformation in the three media. In DMSO, the main structural features are gamma-bends along with a nonspecific bend around Gln(6)-Phe(7)-Tyr(8). The random coil structure seen in water is transformed into a beta-turn around the segment Gln(5)-Gln(6)-Phe(7)-Tyr(8) when Y8SP is incorporated into DPPC bilayers. The lower biological activity of Y8SP compared to the native peptide (SP) has been attributed to the absence of any helix like structure at the central residues, a feature shown to be an important prerequisite for SP and SP agonists to bind to the neurokinin 1 tachykinin receptor.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1999

A novel steroidal spin label for membrane structure studies: synthesis and applications.

2,2,6,6-Tetramethyl piperidine-N-oxyl nitroxyls are known to partition between aqueous and lipid phases, thus serving as probes to study membrane dynamics. The synthesis of a novel steroidal spin label, 3alpha-hydroxycholan-24-yl-(2",2",6",6"-tetramethyl-N-oxyl)p iperidyl butan-1',4'-dioate, containing 2,2,6,6-tetramethylpiperidine-N-oxyl moiety covalently bonded to the side chain in 3,24-caprostan-diol has been described. The localization of this spin label in model biomembranes has been studied by using electron spin resonance, differential scanning calorimetry, and 1H and 31P NMR spectroscopic techniques. Its applicability in studying the phase transition properties of model membrane L-alpha-dipalmitoyl phosphatidyl choline in the presence and absence of drugs has been described by using electron spin resonance. The label has also been used to study the permeability of epinephrine into membrane. The results have shown the applicability of the spin label as a potential spin probe in the study of biomembranes.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1999

1-[2-Hydroxy-3-octadecan-1'-oate]propyl-2'',2'',5'',5''-tetramethyl pyrolidine-N-oxyl-3''-carboxylate as a potential spin probe for membrane structure studies.

The synthesis of a new minimum steric perturbing proxyl nitroxide, which is a derivative of glycerol and contains a stearic acid moiety, has been carried out. Its localization in model membrane L-alpha-dipalmitoyl phosphatidyl choline (DPPC) was ascertained with the help of ESR, DSC, 1H and 31P NMR techniques. The nitroxide was used for detecting the changes in the phase transition temperature of the model membranes in the presence and absence of drugs. The permeation of the vasodilating drug epinephrine has also been studied using this spin label. The results prove the potential applicability of the new spin probe in the spin labeling of biomembranes.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1999

Arginine acts as a protective and reversal agent against glycolytic inhibitors in spermatozoa.

It is known that the amino acid arginine stimulates sperm motility and glycolytic activity. We have earlier studied its efficacy as a stimulator of glycolysis in goat spermatozoa under anaerobic conditions. Here, we have assessed the influence of arginine in reversing the impairment caused by glycolytic inhibitors, iodoacetamide and iodoacetic acid. Glycolysis has been monitored by measuring the consumption of 13C labeled glucose and the amount of 13C labeled lactate produced under different experimental conditions, using 13C NMR. It is observed that both L- and D-arginine are able to prevent and reverse the inhibitory action of glycolytic inhibitors. The reversal effect of arginine gives rise to about eight times higher metabolic activity as compared to the inhibited cells while structurally related amino acids such as nitro-arginine, homo-arginine, lysine and ornithine are ineffective. The energetics of spermatozoa as measured by 31P NMR show a reduction in ATP level in cells incubated with iodoacetamide. Treatment of these cells with both L- and D-arginine restores the ATP level. The results may have significance in the treatment of male infertility.

Animals↗
PubMed1998

Arginine activates glycolysis of goat epididymal spermatozoa: an NMR study.

The present study explores the mechanism underlying the action of L-arginine on the metabolic activity of spermatozoa. Goat epididymal spermatozoa were incubated with different concentrations of L-arginine to determine its effect on the utilization of glucose, fructose, and pyruvate. NMR techniques have been applied to elucidate the effect of L-arginine, L-lysine, and L-ornithine on the glycolysis of epididymal goat spermatozoa. Whereas 31P NMR has been used to estimate the change of pH in the presence of different concentrations of L-arginine, 13C NMR has been used to estimate the substrate consumption and lactate production. At optimal concentration of L-arginine, the forward metabolic rates have been found to increase by two to three times over control experiments. Arginine is not consumed in these reactions, but acts as an activator. Longitudinal relaxation time (T1) measurements indicate that the guanidino group of L-arginine plays an active role in binding to cells. The amino acid L-lysine is less effective, and L-ornithine is ineffective.

Animals↗
PubMed1996

Iron coordination by catechol derivative antioxidants.

Iron complexes of nitrocatechols with different substituent groups [1: -CH = CR2; 2: -CH2-CHR2; 3: -CH = CR'(R")] were synthesized and their effects on iron-induced free radical reactions of biological importance investigated. Catechol and nitrocatechol derivatives effectively inhibited iron-induced lipid peroxide-dependent lipid peroxidation. In the Fenton-like reaction, iron-catechol generated hydroxyl radicals more strongly than did iron citrate, and iron-nitrocatechol derivative 2 generated a small amount of hydroxyl radicals. The iron complexes of derivatives 1 and 3 did not generate hydroxyl radicals. Iron-catechol had the highest ratio of reduction to oxidation rate constants and the second was iron-nitrocatechol 2, suggesting that iron chelated by nitrocatechols 1 and 3 may be most difficult to reduce. To elucidate the structure and physical properties of the iron complexes, UV/vis absorption spectroscopic, ESR and 1H NMR studies were performed in aqueous and DMSO solutions. In aqueous solution at pH 7.4, iron complexes of the nitrocatechol derivatives were high-spin tris(nitrocatecholato)ferrate(III) with a characteristic ligand-to-metal charge transfer absorbance (pi -> d pi). The lambda max of iron-nitrocatechol derivative 2 was shorter than those of iron-nitrocatechol derivatives 1 and 3, suggesting that the reduction potential of iron-nitrocatechol 2 is higher than that of iron-nitrocatechols 1 and 3. Nitrocatechol derivatives with a conjugation structure can sequester the chelated iron more effectively than catechol and the derivative without the conjugation against free radical generation by keeping the iron in the ferric state, probably because of the reduction potentials.

Animals↗
PubMed1995

Riboflavin: a potential material for molecular electronics applications.

Riboflavin (Rbf), the simplest manifestation of the isoalloxazine ring, undergoes redox reactions during biochemical processes. The redox reaction can be simulated under electrochemical conditions. The two oxidation states of the isoalloxazine molecule have different optical properties. Rbf embedded in a polyurethane matrix has been shown to undergo colour change on application of an external electric field. Rbf has been modified to form TARbf and TPRbf--the amphiphilic analogues for the convenience of forming ordered two-dimensional organization. The modification has been found to leave native characteristics of Rbf undisturbed. TPRbf forms monolayers at the air-water interface with a collapse pressure of 10 mNm-1.

Electrochemistry↗
PubMed1995

Membrane fluidization by animycotic bifonazole.

Calorimetry, nuclear magnetic resonance and X-ray diffraction techniques have been used to obtain thermodynamic and structural information on dipalmitoyl phosphatidylcholine (DPPC) liposomes doped by the antimycotic drug bifonazole in the range 0 < R < 1, where R = moles of bifonazole/moles of DPPC. The technique of spin labeling electron spin resonance (ESR) has also been used to study permeability and fluidity properties. The decrease of the cooperativity at the gel to liquid crystalline phase transition, as shown by ESR an DSC measurements, indicates that bifonazole imparts higher fluidity to the lipid matrix. Increase in permeability of ascorbate ions, after incorporation of bifonazole in the membrane, has been detected by ESR experiments using spin label 5-SASL. 13C NMR spectra indicate that the drug molecule is highly immobilized. X-ray diffraction and freeze fracture TEM results show that the equilibrated phase at room temperature is lamellar and unidimensional together with the presence of small particles and pits of uniform size. A marked hysteresis is evident in the formation of this phase.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1994

2'-(3 alpha-Benzyloxy-24-norcholan-23-yl)-2',4',4'-trimethyl-4',5'- dihydrooxazoline-N-oxyl as a potential spin probe for model membranes.

A new steroidal doxyl (4,4-dimethyloxazolidine-N-oxyl) nitroxide (SDN) viz. 2'-(3 alpha-benzyloxy-24-norcholan-2'-yl)-2',4',4'-trimethyl-4',5'- dihydrooxazoline-N-oxyl has been synthesized. This is expected to have higher mobility over other spin labels reported earlier. The localization of this spin probe in lipid bilayers has been determined using 1H NMR and 31P NMR techniques. The alterations induced by drugs in the membrane characteristics such as phase transition and permeability have been investigated using electron paramagnetic resonance (EPR) techniques. The results show the applicability of SDN as a potential spin probe in the study of biomembranes.

Cell Membrane Permeability↗
PubMed1993

Proxyl nitroxide of lithocholic acid: a potential spin probe for model membranes.

A new steroidal proxyl (2,2,5,5-tetramethylpyrrolidine-N-oxyl) nitroxide (SPN), with the proxyl nitroxide moiety in the pendant side chain of the steroid, has been synthesized. Its localization in lipid bilayers was ascertained with the help of 1H NMR and 31P NMR experiments. The effects of the nitroxide group in SPN incorporated into the bilayer on 13C relaxation times are interpreted qualitatively in terms of localization of the nitroxide group within the bilayer structure. The nitroxide SPN was used to monitor changes in membrane fluidity and permeability induced by local anaesthetics, mepivacaine and xylocaine and the antikeratinizing agent, azelaic acid. The results conclusively proved the applicability of the new steroidal proxyl nitroxide (SPN) as a potential spin probe for spin labeling studies.

Cyclic N-Oxides↗
PubMed1992

Biosensors and enzyme immobilized electrodes.

A biosensor is a device which consists of a biological sensing element connected to a transducer. The transducer can be electronic, optical, electrical, etc. This emerging technology offers us a powerful tool which is radically altering our approach to analytical methods. It was realised that enzymes are natural sensors on account of their highly selective nature. Much of the impetus to the work has come from medical requirements. Instant analysis of clinical samples has an obvious appeal to physicians and patients alike. Of particular interest is the possibility of continuous 'in-vivo' monitoring of metabolites, drugs and proteins using miniature, portable systems. In recent years, there has been a growing demand for biosensors in the fields of veterinary science, animal husbandry, the food industry and environmental monitoring. However, the possibility of successful application rests upon future developments. Increasing attention will have to be paid to the engineering of both the basic components and the device on the whole. New biochemical reactions will either have to be discovered or engineered through genetic manipulation or chemical techniques. Optimization of response time, selectivity, stability and low costs should receive priority considerations.

Biosensing Techniques↗
PubMed1992

Maturation of rat spermatozoa: ESR spin labeling studies.

The ability of spermatozoa to reduce nitroxide spin--TEMPO has been used as a parameter to understand maturation, capacitation and calcium uptake of sperm obtained from Holstman strain rats. The rate of spin label reduction by sperm follows the trend--caput greater than cauda greater than corpus. With the increase in age, the electron donating capability shows first a gradual increase, for 60- to 85-day-old rats, peaking at 85 days (corresponding to puberty) and leveling off after 92 days. Calcium uptake takes place in two phases which corresponds to accumulation of and activation by calcium. The presence of polyclonal antibody which is known to cause agglutination, does not adversely affect the sperm activity.

Animals↗
PubMed1991

Glucose oxidase immobilized electrode for potentiometric estimation of glucose.

Glucose oxidase has been immobilized onto a thin platinum strip, by co-crosslinking with bovine serum albumin and glutaraldehyde. The retention of redox characteristics of glucose oxidase has been verified by cyclic voltammetry. The activity of the immobilized enzyme reduces to a quarter of its value when the enzyme is in solution but improves when coimmobilized with 1 M urea. The potentiometric response builds up and remains stable after 100 s. It is sensitive to the thickness of the immobilizing matrix. pH and temperature. An improvement in the performance of the electrode has been achieved by co-immobilizing 2 M urea and metal ions such as Mg2+ and Mn2+. The presence of Cu has been proved to be detrimental. The electrode has been calibrated in the 0.1-5.0 mM glucose concentration range. It gives a stable response for more than 50 independent assays and can be stored for 60 days without significant loss of function.

Biosensing Techniques↗