Stopped-flow Technique Research Articles

Strategies for overcoming tropical disease by ruthenium complexes with purine analog: Application against Leishmania spp. and Trypanosoma cruzi

Tropical diseases currently constitute a major health problem and thus a challenge in the field of drug discovery. The current treatments show serious disadvantages due to cost, toxicity, long therapy duration and resistance, and the use of metal complexes as chemotherapeutic agents against these ailments appears to be a very attractive alternative. Herein, we describe three newly synthesized ruthenium complexes with a bioactive molecule, the purine analogue 5,6,7-trimethyl-1,2,4-triazolo[1,5-a]pyrimidine (tmtp): cis,fac-[RuCl2(dmso)3(tmtp)] (1), mer-[RuCl3(dmso)(H2O)(tmtp)]·2H2O (2) and fac,cis-[RuCl3(H2O)(tmtp)2] (3). Their structures were characterized using X-ray and spectroscopic methods (IR, NMR or EPR). The stability of the synthesized complexes 1–3 in various buffered solutions (pH=3–7.4) was monitored using conventional and stopped-flow techniques. The in vitro antiproliferative activity of all ruthenium complexes against promastigote forms of Leishmania spp. (L. infantum, L. braziliensis, and L. donovani) and epimastigote forms of Trypanosoma cruzi was investigated. Notably, the results showed that the activity of 1 against L. brasiliensis was more than three-fold higher than that of glucantime, and 1 showed no appreciable toxicity towards J774.2 macrophages. Additionally, 2 displayed even 141-fold lower toxicity against host cells than glucantime, demonstrating significantly higher selectivity than the reference drug. Therefore, 1 and 2 appear to be excellent candidates for further development as potential drugs for the effective treatment of leishmaniasis and Chagas disease. All novel complexes were also shown to be potent inhibitors of Fe-SOD in the studied species, while their effects on human CuZn-SOD were very low.

Separating Proton and Electron Transfer Effects in Three-Component Concerted Proton-Coupled Electron Transfer Reactions.

Multiple-site concerted proton-electron transfer (MS-CPET) reactions were studied in a three-component system. 1-Hydroxy-2,2,6,6-tetramethylpiperidine (TEMPOH) was oxidized to the stable radical TEMPO by electron transfer to ferrocenium oxidants coupled to proton transfer to various pyridine bases. These MS-CPET reactions contrast with the usual reactivity of TEMPOH by hydrogen atom transfer (HAT) to a single e-/H+ acceptor. The three-component reactions proceed by pre-equilibrium formation of a hydrogen-bonded adduct between TEMPOH and the pyridine base, and the adduct is then oxidized by the ferrocenium in a bimolecular MS-CPET step. The second-order rate constants, measured using stopped-flow kinetic techniques, spanned 4 orders of magnitude. An advantage of this system is that the MS-CPET driving force could be independently varied by changing either the pKa of the base or the reduction potential (E°) of the oxidant. Changes in ΔG°MS-CPET from either source had the same effect on the MS-CPET rate constants, and a combined Brønsted plot of ln(kMS-CPET) vs ln(Keq) was linear with a slope of 0.46. These results imply a synchronous concerted mechanism, in which the proton and electron transfer components of the CPET process make equal contributions to the rate constants. The only outliers to the Brønsted correlation are the reactions with sterically hindered pyridines, which apparently hinder the close approach of proton donor and acceptor that facilitates MS-CPET. These three-component reactions are compared with a related HAT reaction of TEMPOH, with the 2,4,6-tri-tert-butylphenoxyl radical. The MS-CPET and HAT oxidations of TEMPOH at the same driving force occurred with similar rate constants. While this is an imperfect comparison, the data suggest that the separation of the proton and electron to different reagents does not significantly inhibit the proton-coupled electron transfer process.

The interaction of organoselenium trans-palladium(II) complexes toward small-biomolecules and CT-DNA

A series of organoselenium trans-palladium(II) complexes 1 (bis(2-(phenylselanylmethyl)oxolane)dichloropalladium(II)), 2 (bis(2-(phenylselanylmethyl)oxane)dichloropalladium(II)) and 3 (bis(2,2-dimethyl-3-(phenylselanyl)oxane)dichloropalladium(II)) have been used to investigate reactivity of this specific type of complexes toward different bio-molecules. This system is of special interest because only little is known about the substitution behavior of organoselenium palladium(II) complexes with trans configuration. The substitution of coordinated chloride by a series of small bio-molecules (l-Met, l-His, l-Cys, GSH and 5′-GMP) was studied under pseudo first-order conditions as a function of nucleophile concentration and temperature using stopped-flow technique. The results for the studied complexes indicate that the complex 1 reacts faster compared to the complexes 2 and 3, which is caused by steric effects. The sulfur-donor bio-molecules have shown better reactivity than nitrogen-donor bio-molecules. The activation parameters indicate that substitution occurs by an associative mechanism which is supported by the significantly negative activation entropies. Furthermore, the interaction of these organoselenium trans-palladium(II) complexes with calf thymus DNA (CT-DNA) was further examined by absorption (UV–Vis) and emission spectral studies (Ethidium bromide displacement studies). Overall, the studied complex exhibited good DNA interaction ability.

Recent Progress on the Performance of Different Rate Promoters in Potassium Carbonate Solvents for CO2 Capture

Over recent years our research group has examined the performance of a range of rate promoters in potassium carbonate solvent for carbon dioxide absorption. Promoters including boric acid, potassium glycinate, potassium prolinate, potassium sarcosinate and a thermally stable carbonic anhydrase enzyme have been studied either with the stopped flow technique, a wetted wall column or in the pilot plant located at The University of Melbourne in Australia. The carbonic anhydrase enzyme was shown to have the best promotion performance. However, improvements are required in further improving thermal stability and recycling options in order for the enzyme to be applied in industrial solvent absorption processes. Amino acid salts including potassium sarcosinate, prolinate and glycinate were proven to have high promotion performance at bench scale. Pilot plant results showed that the promotion performance of boric acid was poor, while potassium glycinate was a good rate promoter with an increase in absorption rate of up to 5–6 times when adding 10 wt. % glycinate into 40–45 wt. % potassium carbonate solvent.

Kinetics of Carbon Dioxide (CO2) with DiethylenetriamineinNon-aqueous Solvents Using Stopped-flow Technique

In this work, the reaction kinetics of carbon dioxide (CO2) with diethylenetriamine (DETA)in methanol and ethanol systems were measured using the stopped flow technique over a temperature range of 293-313K in terms of pseudo first-order rate constant (k0). Concentration in the range of 10 to 50mol/m3 for diethylenetriamine were studied. The experimental data show that the pseudo first-order rate constants (k0) increase with the increase of both amine concentration and temperature. The zwitterion mechanism and the termolecular mechanism were used to representthe data for DETAinmethanol and ethanol systems with excllent ADDs of 3.5%, 2.4%, respectively. In comparison with EDA and AEEA in terms of k2, DETA exhibits a better reaction kinetics performance for capturingCO2. Those results will be useful in finding an efficient method for the removal of CO2 from industrial gases.

Kinetics of Carbon Dioxide Absorption by Nonaqueous Solutions of Promoted Sterically Hindered Amines

The mechanism and kinetics of CO2 capture using piperazine (PZ) promoted nonaqueous solutions of 2-amino-2-methyl-1,3-propanediol (AMPD) and 2-amino-2-ethyl-1,3-propanediol (AEPD) were investigated by stopped-flow technique. The termolecular mechanism was used to model the kinetics of the reactions. AMPD or AEPD like other sterically hindered amines absorbs CO2 in an equimolar ratio that is significantly higher than that of monoethanolamine (MEA). However, the steric hinderance results in decreased reaction rate as in the case of AMPD and AEPD. The reaction can be promoted by addition of small amounts of cyclic polyamines, such as PZ or its derivatives. Our results show clearly that nonaqueous solutions of AMPD and AEPD can achieve reaction rates comparable to commercial systems by the addition of small amounts of PZ.

Affinity switching for lysozyme and dual-responsive microgels by stopped-flow technique: Kinetic control and activity evaluation

The use of proteins as therapeutics in nanomedicine is an emerging research field and has developed rapidly. However, proteins are always vulnerable to renal excretion or digestion by the proteolytic system in vivo, which limits their usage to a large extent. Although biocompatible polymers have been covalently linked to proteins to protect them from recognition by the immune system and prolong their circulation time, the biological activity of them is sometimes decreased. To fill this gap, physical isolation, wrapping, or encapsulation techniques are employed. Up to now, various mature examples were reported, but the whole time scales for guest molecules loading and releasing, especially the initial rapid loading process, were rarely mentioned. Herein, a series of dual-responsive poly(N-isopropylacrylamide-co-methacrylic acid) (P(NIPAM-co-MAA)) microgels were synthesized and employed to investigate the kinetics of in situ complexation and release of lysozyme under external stimuli modulation upon a stopped-flow apparatus, which was suitable for rapid dynamic monitoring. Close inspection of the adsorption kinetics during the early stages (< 50 s) revealed that the initial microgel collapse occurred within ~1 s, with more rapid transitions being observed when higher lysozyme concentrations were targeted. All the dynamic traces could be well fitted with a double exponential function, suggesting a fast (τ 1) and a slow (τ 2) relaxation time, respectively. Then, the kinetics of releasing bound lysozyme from microgels was carried on by utilizing the pH-responsive property, and the evaluation of the activity of released lysozyme was synchronously measured in a Micrococcus lysodeikticus (M. lysodeikticus) cell suspension. The corresponding relaxation time (τ) was also calculated by fitting the recorded dynamic traces. We speculate that this work can provide basic dynamics data and theoretical basis for microgels based nanocarriers to be used for protein delivery, controlled release, and possible chemical separation.

Novel real-time method using the stopped-flow for evaluating bisphenol A degradation kinetics by molecular ozone and radical mechanisms

ABSTRACTA novel real-time method was developed to evaluate the bisphenol A degradation kinetics by molecular ozone and radical pathway using the stopped-flow technique. The second-order kinetics was determined under pseudo-first-order conditions for the molecular pathway by the absolute rate constant method and for the radical pathway by the Rct concept involving the hydroxyl radical and ozone ratio. Bisphenol A degradation by ozone was performed and evaluated at a pH ranging from 2 to 10. At pH < 4 (molecular pathway), the second-order rate constant value was 1.12 × 104 M−1 s−1 and for the radical pathway at pH > 10, the constant was 3.43 × 109 M−1 s−1. To validate the method, ciprofloxacin degradation kinetics was determined at pH 8 by radical pathway, in 4.55 × 109 M−1 s−1. The method permits the determination of kinetic parameters for the design of chemical reactors; avoiding the generation of undesirable reactions and by-products in the degradation of emerging compounds.

Factors controlling the reactivity of divalent metal ions towards pheophytin a

In this study, we evaluate the factors which determine the reactivity of divalent metal ions in the spontaneous formation of metallochlorophylls, using experimental and computational approaches. Kinetic studies were carried out using pheophytin a in reactions with various divalent metal ions combined with non- or weakly-coordinative counter ions in a series of organic solvents. To obtain detailed insights into the solvent effect, the metalations with the whole set of cations were investigated in three solvents and with Zn2+ in seven solvents. The reactions were monitored using electronic absorption spectroscopy and the stopped-flow technique. DFT calculations were employed to shed light on the role of solvent in activating the metal ions towards porphyrinoids. This experimental and computational analysis gives detailed information regarding how the solvent and the counter ion assist/hinder the metalation reaction as activators/inhibitors. The metalation course is dictated to a large extent by the reaction medium, via either the activation or deactivation of the incoming metal ion. The solvent may affect the metalation in several ways, mainly via H-bonding with pyrrolenine nitrogens and the activation/deactivation of the incoming cation. It also seems to affect the activation enthalpy by causing slight conformational changes in the macrocyclic ligand. These new mechanistic insights contribute to a better understanding of the “metal–counterion–solvent” interplay in the metalation of porphyrinoids. In addition, they are highly relevant to the mechanisms of metalation reactions catalyzed by chelatases and explain the differences between the insertion of Mg2+ and other divalent cations.

Mechanistic study of the substitution reactions of [Pt(II)(bis(2-pyridylmethyl)amine)H2O](ClO4)2 and [Pt(II)(bis(2-pyridylmethyl)sulfide)H2O](ClO4)2 with azole Nucleophiles. Crystal structure of [Pt(II)(bis(2-pyridylmethyl)sulfide)Cl]ClO4

The substitution kinetics of the complexes [Pt(II)(bis(2-pyridylmethyl)amine)H2O](ClO4)2, Ptdpa and [Pt(II)(bis(2-pyridylmethyl)sulfide)H2O](ClO4)2, Ptdps, with a series of azole nucleophiles: Imidazole (Im), 1-methylimidazole (MIm), 1,2-Dimethylimidazole (DIm), 1,2,4-triazole (Trz) and pyrazole (Pyz), were studied in an aqueous medium at constant ionic strength (0.1M NaClO4). The substitution of the coordinated water ligand on the Pt(II) complexes by the azoles was studied under pseudo-first order conditions as a function of the incoming nucleophiles concentration and temperature using either stopped-flow techniques or UV-Vis spectroscopy. Ptdps was found to be more reactive (three magnitude higher) than Ptdpa. The second-order rate constant, k2, for all the nucleophiles ranged between 0.087±0.005 and 0.926±0.05M−1s−1 for Ptdpa and between 146±4 and 1458±10M−1s−1 for Ptdps. The rate of substitution of the aqua ligand is dependent on the strength of the σ-donor character and the π-acceptability of the trans atom to the leaving group. The observed reactivity trend for the azoles followed the trend, MIm>Im>DIm>Trz>Pyz. This reactivity trend correlates with the basicity, steric and electrophilic effects of the nucleophiles. The X-ray crystal structure of Ptdps–Cl is reported.

Reaction kinetics of carbon dioxide with aqueous solutions of l-Arginine, Glycine & Sarcosine using the stopped flow technique

The use of amino acids as potential solvents for carbon dioxide (CO2) capture has been considered by a number of researchers. However, very little is known about the kinetics and mechanism of amino acids-CO2 reactions. In this work, we investigate the reactions of three amino acids (l-Arginine, Glycine and Sarcosine) with CO2 in aqueous media using stopped-flow conductivity technique. The experiments were performed at temperatures between 293 and 313K and amino acids concentrations were in the range of 0.05–0.2 molar. The overall rate constants (kov) was found to increase with increased amino acid concentration and solution temperature. Both zwitterion and termolecular mechanisms were used to model and interpret the data. However, the Zwitterion mechanism was found to be the preferred one. From the stopped-flow results at pH around 6, we found that neutral l-Arginine, Glycine and Sarcosine react with CO2(aq) with k(M−1s−1)=2.81×1010exp(−4482.9T(K)), k(M−1s−1)=3.29×1013exp(−8143.7T(K)) and k(M−1s−1)=3.90×1013exp(−7991.0T(K)) respectively. The corresponding activation energies are 37.28kJmol−1, 67.71kJmol−1 And 66.44kJmol−1 respectively. A comparison between the kinetics of the three amino acids showed that Arginine exhibits highest reaction rate with CO2 followed by Sarcosine and then Glycine. The technique and results obtained from this work can be used as strong tools in the development of efficient new solvents for the removal of CO2 from flue and industrial gases.

Cellular Fates of Manganese(II) Pentaazamacrocyclic Superoxide Dismutase (SOD) Mimetics: Fluorescently Labeled MnSOD Mimetics, X-ray Absorption Spectroscopy, and X-ray Fluorescence Microscopy Studies.

Manganese(II) pentaazamacrocyclic complexes (MnPAMs) can act as small-molecule mimics of manganese superoxide dismutase (MnSOD) with potential therapeutic application in conditions linked to oxidative stress. Previously, the in vitro mechanism of action has been determined, their activity has been demonstrated in cells, and some representatives of this class of MnSOD mimetics have entered clinical trials. However, MnPAM uptake, distribution, and metabolism in cells are largely unknown. Therefore, we have used X-ray fluorescence microscopy (XFM) and X-ray absorption spectroscopy (XAS) to study the cellular fate of a number of MnPAMs. We have also synthesized and characterized fluorescently labeled (pyrene and rhodamine) manganese(II) pyane [manganese(II) trans-2,13-dimethyl-3,6,9,12,18-pentaazabicyclo[12.3.1]octadeca-1(18),14,16-triene] derivatives and investigated their utility for cellular imaging of MnPAMs. Their SOD activity was determined via a direct stopped-flow technique. XFM experiments show that treatment with amine-based manganese(II) pyane type pentaazamacrocycles leads to a 10-100-fold increase in the overall cellular manganese levels compared to the physiological levels of manganese in control cells. In treated cells in general, manganese was distributed throughout the cell body, with a couple of notable exceptions. The lipophilicity of the MnPAMs, examined by partitioning in octanol-buffer system, was a good predictor of the relative cellular manganese levels. Analysis of the XAS data of treated cells revealed that some fraction of amine-based MnPAMs taken up by the cells remained intact, with the rest transformed into SOD-active manganese(II) phosphate. Higher phosphate binding constants, determined from the effect of the phosphate concentration on in vitro SOD activity, were associated with more extensive metabolism of the amine-based MnPAMs to manganese(II) phosphate. In contrast, the imine-based manganese(II) pydiene complex that is prone to hydrolysis was entirely decomposed after uptake and free manganese(II) was oxidized to a manganese(III) oxide type species, in cytosolic compartments, possibly mitochondria. Complex stability constants (determined for some of the MnPAMs) are less indicative of the cellular fate of the complexes than the corresponding phosphate binding constants.

Oxidation of inorganic compounds by aqueous permanganate: Kinetics and initial electron transfer steps

Abstract In this study, the kinetics of the reactions between aqueous permanganate (Mn(VII)) and selected inorganic compounds (e.g. KSCN, NaNO2, Na2SO3, Na2S2O3, KI, N2H4, H2O2, and NH2OH) were investigated in aqueous solutions at pH 5–9 with excess pyrophosphate (L) by detecting the decay of Mn(VII) using a stopped-flow technique. With excess L, soluble Mn(III)L rather than MnO2 was generated, which had negligible interference on examining the change of the absorbance of Mn(VII) at 525 nm (∼45 M−1 cm−1 for Mn(III)L vs 2500 M−1 cm−1 for Mn(VII)). The reactions of Mn(VII) with Na2SO3, Na2S2O3, KSCN, NaNO2, and KI (referred to as SA) exhibited second-order kinetics. Comparatively, the reactions of Mn(VII) with N2H4, H2O2, and NH2OH (referred to as SB) showed two-phase kinetics (i.e., an initial lag phase and a secondary fast phase), and the second-order rate constants (k1, M−1 s−1) were calculated in the initial phase. In the second phase, Mn(III)L reacted with SB producing Mn(II), and then Mn(II) reacted fast with Mn(VII) producing Mn(III)L. The kinetics of the reaction between Mn(II) and Mn(VII) with excess L at pH 4–9 were examined and the loss rate of Mn(VII) increased with increasing pH value. The species-specific second-order rate constants (k1i, M−1 s−1) for the reactions of Mn(VII) with inorganic compounds were determined by the experimental k1 values. Furthermore, a linear free energy relationship (LFER) was found (log(k1i) = 2.86 − 1.90E(2)0) between log k1i and 2−e− reduction potentials for inorganic species (i.e., SO32−, S2O32−, SCN−, H2O2, NH2OH, I−, and N2H4) while only NO2− showed 1−e− transfer process, indicating highly active aqueous manganese intermediates (Mn(V) and Mn(VI)) formed in situ upon Mn(VII) reduction.

Enhanced Oxidation of Tetracycline by Permanganate via the Alkali-Induced Alteration of the Highest Occupied Molecular Orbital and the Electrostatic Potential

The reaction between tetracycline and alkaline permanganate was investigated by combining experimental and computational methods. The kinetics was initially studied using a stopped-flow technique and was found to be first-order in tetracycline and permanganate. The second-order rate constant was positive linearly dependent on the concentration of hydroxyl ion (0.01–0.10 M), indicating the presence of base catalysis. By construction of the Eyring plots in the range of 293–308 K, a lower activation barrier ((14.89 ± 0.44) kcal mol–1) was obtained at 298 K for the base-catalyzed pathway compared with that ((17.72 ± 1.84) kcal mol–1) for the uncatalyzed pathway. The effect of ionic strength further suggested the existence of a complex with higher charge and reactivity. It is confirmed by the theoretical analysis that the hydroxyl ion could attract the proton of tetracycline toward itself to form a complex-like structure with a highly reactive phenolate-type moiety. The highest occupied molecular orbital of te...

Restricting the conformational freedom of the neuronal nitric-oxide synthase flavoprotein domain reveals impact on electron transfer and catalysis

The signaling molecule nitric oxide (NO) is synthesized in animals by structurally related NO synthases (NOSs), which contain NADPH/FAD- and FMN-binding domains. During catalysis, NADPH-derived electrons transfer into FAD and then distribute into the FMN domain for further transfer to internal or external heme groups. Conformational freedom of the FMN domain is thought to be essential for the electron transfer (ET) reactions in NOSs. To directly examine this concept, we utilized a "Cys-lite" neuronal NOS flavoprotein domain and substituted Cys for two residues (Glu-816 and Arg-1229) forming a salt bridge between the NADPH/FAD and FMN domains in the conformationally closed structure to allow cross-domain disulfide bond formation or cross-linking by bismaleimides of various lengths. The disulfide bond cross-link caused a ≥95% loss of cytochrome c reductase activity that was reversible with DTT treatment, whereas graded cross-link lengthening gradually increased activity, thus defining the conformational constraints in the catalytic process. We used spectroscopic and stopped-flow techniques to further investigate how the changes in FMN domain conformational freedom impact the following: (i) the NADPH interaction; (ii) kinetics of electron loading (flavin reduction); (iii) stabilization of open versus closed conformational forms in two different flavin redox states; (iv) reactivity of the reduced FMN domain toward cytochrome c; (v) response to calmodulin binding; and (vi) the rates of interflavin ET and the FMN domain conformational dynamics. Together, our findings help explain how the spatial and temporal behaviors of the FMN domain impact catalysis by the NOS flavoprotein domain and how these behaviors are governed to enable electron flow through the enzyme.

Screening Amino Acid Salts as Rate Promoters in Potassium Carbonate Solvent for Carbon Dioxide Absorption

Potassium carbonate shows promise as a solvent for carbon capture due to its low cost and low environmental impact. However, improving the absorption kinetics of potassium carbonate solvent is crucial for reducing the capital cost of absorption equipment required to build the carbon dioxide capture plant. In this study, a series of amino acid salts were screened as reactants with carbon dioxide using the stopped flow technique. The amino acids investigated in this study were 2-piperazinecarboxylic acid, asparagine, aspartic acid, glycine, leucine, lysine, proline, sarcosine, serine, and valine. Furthermore, proline, sarcosine, glycine, leucine, and lysine were tested as rate promoters in potassium carbonate solvent for carbon dioxide absorption using a wetted wall column. Results showed that the amino group in the anions of the amino acid salt is the major species reacting with carbon dioxide. Therefore, the promoting effect of amino acid salts is sensitive to changes in pH values due to changes in specie...

Reaction kinetics of the absorption of carbon dioxide (CO2) in aqueous solutions of sterically hindered secondary alkanolamines using the stopped-flow technique

In this work, pseudo first-order reaction rate constants (k0) for the reaction of CO2 with sterically hindered secondary amines at temperatures of 293–313K and various amine concentrations were studied using the rapid-mixing stopped-flow technique. It was found that the values of k0 increased as the concentration of amine in aqueous solution increased and as solution temperatures were increased. The second order rate constant (k2) at 293–313K was also determined based on the proposed reaction mechanisms. The termolecular mechanism was able to fit the experimental data with predicted CO2 absorption rates of MAE and EAE with an absolute average deviation (AAD) of 2.81% and 14.96%, respectively. However, the base-catalyzed hydration mechanism is more precise in terms of the hindered amines (IPAE and TBAE) in predicting the CO2 absorption rates with AAD of 7.97% and 5.15%. The combined data for the CO2 loading showed that all four amines have good properties for the post-combustion CO2 capture process in comparison with MEA. In comparison with the amine reference MEA (CO2 loading=0.58molCO2/mole of amine), the CO2 loading of the four amines is between 0.66 and 0.93mol of CO2/mole amine. These results show that IPAE and TBAE are good candidates for CO2 capture as alternatives to MEA because of their good CO2 absorption and high reaction rate with CO2.

Binding interaction of sodium-N-dodecanoyl sarcosinate with hemoglobin and myoglobin: Physicochemical and spectroscopic studies with molecular docking analysis

The interaction of an amino acid surfactant, sodium-N-dodecanoyl sarcosinate (SDDS), with two heme proteins, hemoglobin (Hb) and myoglobin (Mb), has been studied employing various physicochemical and spectroscopic techniques like tensiometry, UV–Vis spectroscopy, steady-state fluorometry, time-resolved fluorometry, circular dichroism (CD) spectroscopy, calorimetry and stopped flow kinetics at physiological pH of 7.2 and 298K. Tensiometric and fluorometric analysis suggest that the interaction between SDDS and protein starts with the monomer form of the surfactant which produces small induced micelles. The micelles bind to protein backbone causing denaturation of the protein structure, and finally free micelles are formed on further addition of surfactant. Life-time measurements have been performed to shed light on the binding of surfactant around the tryptophan moieties present in the heme proteins. The changes in protein secondary structure induced by AAS collected from CD measurements reveals that the proteins are quite perturbed upon action of SDDS. The enthalpy change values of each stepwise interaction process have been found from isothermal titration calorimetry (ITC). The kinetics of the protein-surfactant interaction process has been studied using stopped flow technique. Quantum mechanical calculations involving density functional theory (DFT) and molecular docking analysis have been performed to highlight the interactive phenomenon between the surfactant and heme proteins. Thus the entire study shows a total inspection of an amino acid surfactant–heme protein interaction.

Covalently Connected Polymer-Protein Nanostructures Fabricated by a Reactive Self-Assembly Approach.

The synthesis of polymer-protein nanostructures opens up a new avenue for the development of new biomaterials. In this research, covalently connected polymer-protein nanostructures were fabricated through a reactive self-assembly approach. Poly(tert-butyl methacrylate-co-pyridyl disulfide methacrylamide) (PtBMA-co-PPDSMA) was synthesized by reversible addition fragmentation chain transfer (RAFT) polymerization. Covalently connected nanostructures (CCNs) with hydrophobic polymer cores and hydrophilic protein coronae were prepared by adding solutions of PtBMA-co-PPDSMA/DMF to aqueous solutions of bovine serum albumin (BSA). The thiol-disulfide exchange reaction between pyridyl disulfide groups on the polymer chains and thiol groups on the protein molecules plays a key role in the fabrication of CCNs. The self-assembly process was investigated by dynamic light scattering (DLS) and stopped-flow techniques. DLS results indicated that the sizes of the CCNs were determined by the initial polymer concentration, the BSA concentration, and the average number of thiol groups on BSA molecules. TEM and sodium dodecyl sulfate polyacrylamide gel electrophoresis were used to analyze the nanostructures. Far-UV circular dichroism results demonstrated that the original folded conformations of BSA molecules were basically maintained in the reactive self-assembly process. Compared with native BSA, the secondary structure and conformation change of coronal BSA induced by urea or thermal treatment were remarkably suppressed. The cytotoxicity assays demonstrated that the CCNs were essentially nontoxic to Hela and COS-7 cells.

Kinetics and mechanism study of homogeneous reaction of CO2 and blends of diethanolamine and monoethanolamine using the stopped-flow technique

The stopped-flow technique was applied on the measurement of the kinetics of carbon dioxide (CO2) absorption into aqueous blends of monoethanolamine (MEA) and diethanolamine (DEA) over the temperature range of 293–313K. The investigation of blended MEA+DEA with various molar ratios of DEA to MEA revealed that the reaction mechanism between CO2 and blended absorbents could be different at different ratio of DEA to MEA. Consequently, the kinetic data obtained in this work was split into two groups with respect to the different molar ratios of DEA to MEA in order to study the different mechanisms. In group A, the concentration of MEA was in the range of 5–15mol/m3 and the concentration of DEA was low (varied between 5 and 15mol/m3), while in group B, the DEA concentration was high (varied between 140 and 240mol/m3) and the concentration of MEA remained in the range of 5–15mol/m3. Modified models based on the termolecular mechanism were developed for each group and used to interpret the experimental kinetic data. Results showed that the models could explain the data well with an AAD of 4.71% for group A at low concentration of DEA and 3.33% for group B at high concentration of DEA. It is interesting to point out that DEA barely reacts with CO2 at low molar ratios (i.e. group A) whereas at high DEA concentration (i.e. group B), both MEA and DEA reacted with CO2.