Malachite green is an organic compound that can be widely used as a dyestuff for various materials; it has also emerged as a controversial agent in aquaculture. Since malachite green is proven to be carcinogenic and mutagenic, it may become a hazard to public health. For this reason, it is urgently required to analyze this controversial dye in more detail. In our current research, the interaction between malachite green and hemoglobin under physiological conditions was investigated by the methods of molecular modeling, fluorescence spectroscopy, circular dichroism (CD) as well as hydrophobic ANS displacement experiments. From the molecular docking, the central cavity of hemoglobin was assigned to possess high-affinity for malachite green, this result was corroborated by time-resolved fluorescence and hydrophobic ANS probe results. The recognition mechanism was found to be of static type, or rather the hemoglobin–malachite green complex formation occurred via noncovalent interactions such as π–π interactions, hydrogen bonds and hydrophobic interactions with an association constant of 104 M−1. Moreover, the results also show that the spatial structure of the biopolymer was changed in the presence of malachite green with a decrease of the α-helix and increase of the β-sheet, turn and random coil suggesting protein damage, as derived from far-UV CD and three-dimensional fluorescence. Results of this work will help to further comprehend the molecular recognition of malachite green by the receptor protein and the possible toxicological profiles of other compounds, which are the metabolites and ramifications of malachite green.

Since the introduction of imidacloprid in the early 1990s, it has become one of the most widely applied insecticides, and currently represents about 20% of the global pesticide market (Tomizawa, M.; Casida, J. E. J. Agric. Food Chem 2011, 59, 2883−2886). In the context of this study, our major aim was to comprehensively scrutinize the nature of imidacloprid with two typical model proteins, lysozyme and albumin, by means of circular dichroism (CD), steady-state and time-resolved fluorescence, and molecular modeling at the molecular level. Far-UV CD verified that the spatial structure of both proteins was altered with a distinct reduction of α helix in the presence of imidacloprid suggesting unfolding of the protein (i.e., protein damage). The data of steady-state and time-resolved fluorescence showed that the conjugation of imidacloprid with lysozyme yielded quenching by a static mechanism (KSV = 3.841 × 104 M–1), while combined static and dynamic properties existed for albumin tryptophan (Trp)-214 fluorescence. Molecular modeling simulations displayed that the imidacloprid binding site was near to the Trp-62 and Trp-63 residues of lysozyme, and it was located at the subdomain IIA (warfarin-azapropazone site) of albumin. Furthermore, the primary forces between protein and imidacloprid are hydrogen bond, hydrophobic, and π–π interactions, but the affinity of lysozyme with imidacloprid is much lower than albumin, probably because the affinity distinctions stem from discrepancy in the three-dimensional structure of the two globular proteins. The results presented here will help to further understand the credible mechanism by which the toxicological implication of neonicotinoid insecticides is palliated by carrier protein.

Phenosafranine is a toxic and recalcitrant compound, whose capacity to intercalate with double stranded DNA has been shown. In this contribution, a biophysical discuss on the conjugation of phenosafranine with two model proteins human serum albumin (HSA) and lysozyme (Lys) has been identified utilizing a combination of molecular modeling, steady state and time-resolved fluorescence and circular dichroism (CD) approaches. The accurate binding domain of phenosafranine in protein has been characterized from molecular modeling, subdomain IIIA of HSA and Trp-62, Trp-63 and Trp-108 residues of Lys was designated to possess high-affinity for this compound, the dominant forces in the protein–phenosafranine adduct are hydrogen bonds and π–π interactions, but hydrophobic interactions between dye and Lys are also not exclude. The data of fluorescence displayed that the complex of phenosafranine with protein produces quenching through static property, this corroborates the time-resolved fluorescence results that the ground state complex formation with a moderate affinity of 104 M−1. Moreover, via synchronous fluorescence, CD and three-dimensional fluorescence we indicated some extent of polypeptide chain of protein partially unfolding upon conjugation with phenosafranine. Through this work, we anticipate it can supply salient clues on the toxicological action of phenosafranine and other azines, which have analogous configuration with phenosafranine.Highlights► Our study supply salient clues on the toxicological action of toxic phenosafranine. ► Phenosafranine is situated within subdomain IIIA, Sudlow's site II on HSA. ► Trp-62, Trp-63 and Trp-108 residues on the Lys molecule are all close to dye. ► Static property of the phenosafranine induced quenching of protein Trp fluorescence. ► Protein spatial structure proved to be disturbed after complex with phenosafranine.

Hesperidin is a flavanone glycoside widely available for dietary intake in citrus fruits or citrus fruit derived products; however, exhaustive and reliable data are scarcely available for biological activity when it exerts protective health effects in humans. The principal intent of this work is to check binding domain and structural changes of human serum albumin (HSA), the primary carrier of flavonoids, in blood plasma association with hesperidin by employing molecular modeling, steady state and time-resolved fluorescence, and circular dichroism (CD) methods. From molecular modeling simulations, subdomains IIA and IIIA, which correspond to Sudlow’s sites I and II, respectively, were earmarked to possess affinity for hesperidin, but the affinity of site I with flavanone is greater than that of site II. This corroborates the site-specific probe and hydrophobic 8-anilino-1-naphthalenesulfonic acid (ANS) displacement results placing the hesperidin at warfarin–azapropazone and indole–benzodiazepine sites. Steady state and time-resolved fluorescence manifested that static type, due to HSA–hesperidin complex formation (1.941 × 104 M–1), is the operative mechanism for the diminution in the tryptophan (Trp)-214 fluorescence. Moreover, via alterations in three-dimensional fluorescence and CD spectral properties, we can securely draw the conclusion that the polypeptide chain of HSA is partially destabilized after conjugation with hesperidin. We anticipate that this study can provide better knowledge of bioavailability such as absorption, biodistribution, and elimination, of hesperidin in vivo, to facilitate the comprehension of the biological responses to physiologically relevant flavanones.

Chloramphenicol is a low cost, broad spectrum, highly active antibiotic, and widely used in the treatment of serious infections, including typhoid fever and other life-threatening infections of the central nervous system and respiratory tract. The purpose of the present study was to examine the conjugation of chloramphenicol with hemoglobin (Hb) and compared with albumin at molecular level, utilizing fluorescence, UV/vis absorption, circular dichroism (CD) as well as molecular modeling. Fluorescence data indicate that drug bind Hb generate quenching via static mechanism, this corroborates UV/vis absorption measurements that the ground state complex formation with an affinity of 104 M−1, and the driving forces in the Hb-drug complex are hydrophilic interactions and hydrogen bonds, as derived from computational model. The accurate binding site of drug has been identified from the analysis of fluorescence and molecular modeling, α1β2 interface of Hb was assigned to possess high-affinity for drug, which located at the β-37 Trp nearby. The structural investigation of the complexed Hb by synchronous fluorescence, UV/vis absorption, and CD observations revealed some degree of Hb structure unfolding upon complexation. Based on molecular modeling, we can draw the conclusion that the binding affinity of drug with albumin is superior, compared with Hb. These phenomena can provide salient information on the absorption, distribution, pharmacology, and toxicity of chloramphenicol and other drugs which have analogous configuration with chloramphenicol.Highlights► Complex formation is dominance for the reduction in the β-37 Trp fluorescence. ► α1β2 interface of Hb is designated to possess high-affinity for chloramphenicol. ► Hydrophilic interactions and hydrogen bonds exist between drug and Hb. ► The Hb structure unfolds upon drug complexation. ► The binding affinity of drug with albumin is superior, compared with Hb.

The purpose of the current work was to examine the complexation of a mammalian protein, hemoglobin (Hb) with a food additive hesperidin at physiological conditions. Molecular modeling, fluorescence, and circular dichroism (CD) methods were exploited to analyze the binding domain, affinity, and the effects of hesperidin conjugation on Hb spatial structure. From molecular modeling, central cavity of Hb was assigned to retain high-affinity for hesperidin, this corroborates the steady state fluorescence and hydrophobic ANS probe results. The association of hesperidin with Hb emerges fluorescence quenching via static type, this phenomenon display that the ground state complex formation with an affinity of 104 M−1, and hypsochromic effect transpires. Additionally, the alterations of synchronous fluorescence, CD, and three-dimensional fluorescence suggest that the polypeptide chain of Hb partially folding after conjugation with hesperidin. The above data suggest that Hb plays a significant role in the plasma distribution and transportation of hesperidin and related dietary flavonoids.Graphical abstractHesperidin situates within the central cavity of Hb, as well as the amino acid residues, such as Tyr-35, Trp-37, Pro-95, Phe-98, Lys-99, Glu-101, and Asp-126, which all keep an appropriate distance involved in making hydrogen bonds, π–π, and electrostatic interactions with hesperidin, this conjugation arouses faint folding of the polypeptide chain of protein, ascends the hydrophobicity and leads to structural changes.Highlights► Central cavity of Hb is designated to possess high-affinity for hesperidin. ► Static type is dominance for the reduction in the β-37 Trp residue fluorescence. ► Hydrogen bond, π–π, and electrostatic interaction exist between Hb and hesperidin. ► Hb structure folds upon complex with hesperidin. ► Our task disclose flavonoids absorption, bioavailability, and biological activity.

Chlorantraniliprole is a novel insecticide belonging to the diamide class of selective ryanodine receptor agonists. A biophysical study on the binding interaction of a novel diamide insecticide, chlorantraniliprole, with staple in vivo transporter, human serum albumin (HSA) has been investigated utilizing a combination of steady-state and time-resolved fluorescence, circular dichroism (CD), and molecular modeling methods. The interaction of chlorantraniliprole with HSA gives rise to fluorescence quenching through static mechanism, this corroborates the fluorescence lifetime outcomes that the ground state complex formation and the predominant forces in the HSA–chlorantraniliprole conjugate are van der Waals forces and hydrogen bonds, as derived from thermodynamic analysis. The definite binding site of chlorantraniliprole in HSA has been identified from the denaturation of protein, competitive ligand binding, and molecular modeling, subdomain IIIA (Sudlow's site II) was designated to possess high-affinity binding site for chlorantraniliprole. Moreover, using synchronous fluorescence, CD, and three-dimensional fluorescence we testified some degree of HSA structure unfolding upon chlorantraniliprole binding.Highlights► Our study highlights for the first time how binding dynamics can predominate for the new diamide insecticide, chlorantraniliprole. ► Chlorantraniliprole is situated within subdomain IIIA, Sudlow's site II, which is the same as that of indole-benzodiazepine site. ► Biophysical and molecular modeling approaches are useful to resolve the ligand interaction with biomacromolecule. ► It serves as a protective device in binding and in inactivating potential toxic compounds to which the body is exposed.

Humic acid, a natural ionic molecule, is rapidly being recognized as one of the crucial elements in our modern diets of the new century. A biophysical protocol utilizing circular dichroism (CD), steady state and time-resolved fluorescence for the investigation of the complexation of the humic acid to the staple in vivo transporter, human serum albumin (HSA), as a model for protein-humic substances, is proclaimed. The alterations of CD and three-dimensional fluorescence suggest that the polypeptide chain of HSA partially folded after complexation with humic acid. The data of fluorescence emission displayed that the binding of humic acid to HSA is the formation of HSA–humic acid complex with an association constant of 104 M−1; this corroborates the fluorescence lifetime measurements that the static mechanism was operated. The precise binding domain of humic acid in HSA has been verified from the denaturation of albumin, hydrophobic ANS displacement, and site-specific ligands; subdomain IIA (Sudlow's site I) was earmarked to possess high-affinity for humic acid. The observations are relevant for other albumin–humic substance systems when the ligands have analogous configuration with humic acid.Highlights► Albumin structure partially folds upon humic acid complexation. ► Static type is dominance for the diminution in the Trp-214 fluorescence.► Subdomain IIA is designate to possess high-affinity site for humic acid.

Binding of the drug phenosafranine to hemoglobin (Hb) in aqueous solutions was investigated by fluorescence, UV/vis and circular dichroism (CD) spectral methods at pH=7.4. The fluorescence data showed that fluorescence quenching of Hb by phenosafranine is the result of formation of a phenosafranine–Hb complex with a 1:1 molar ratio. Thermodynamic analysis implied that hydrophobic, electrostatic and hydrogen bond interactions are all involved in stabilizing the complex. The molecular distance (r=4.29 nm) between the donor (Hb) and acceptor (phenosafranine) was calculated according to Förster’s theory. The features of phenosafranine-induced secondary structure changes of Hb have been studied by synchronous fluorescence, CD and three-dimensional fluorescence spectroscopy. This study improves our knowledge of the interaction dynamics of phenazinium drugs to the physiologically important protein Hb.

Bensulfuron-methyl (BM) is a highly active sulfonylurea herbicide for use on paddy rice. Steady state fluorescence, UV/vis absorption, circular dichroism (CD), time-resolved fluorescence and molecular modeling methods have been exploited to determine the binding affinity and binding site of BM to human serum albumin (HSA). From the synchronous fluorescence, UV/vis, CD and three-dimensional fluorescence spectra, it was evident that the interaction between BM and HSA induced a conformational change in the protein. Steady state and time-resolved fluorescence data illustrates that the fluorescence quenching of HSA by BM was the formation of HSA–BM complex at 1:1 molar ratio. Site marker competitive experiments demonstrated that the binding of BM to HSA primarily took place in subdomain IIIA (Sudlow’s site II), this corroborates the hydrophobic probe ANS displacement and molecular modeling results. Thermodynamic analysis displays hydrophobic, electrostatic and hydrogen bonds interactions are the major acting forces in stabilizing the HSA–BM complex.

The protein–ligand system constituted by human serum albumin (HSA) and sulfometuron-methyl (SM) has been investigated by using tryptophan fluorescence, UV–vis absorption, circular dichroism (CD) and molecular modeling. The Stern–Volmer analysis indicated that the fluorescence quenching of HSA by SM was resulted from static mechanism, and the binding constants (Kb) were 4.785, 3.803, 3.114 and 2.205 × 104 M−1 at 291, 297, 303 and 309 K, respectively. The secondary structure changes of HSA upon SM binding were evaluated by measuring synchronous fluorescence, UV–vis, far-UV CD and three-dimensional fluorescence spectroscopy properties of the HSA–SM complex. Through site marker competitive experiments, subdomain IIA of HSA has been assigned to possess the high-affinity binding site of SM, and corroborates with the hydrophobic probe ANS displacement results and molecular modeling simulations. In addition, thermodynamic analysis implied the roles of hydrophobic forces, van der Waals forces and hydrogen bonds interactions in stabilizing the HSA–SM complex. The binding research presented in this paper enriches our knowledge of the interaction dynamics of sulfonylurea herbicides to the important plasma protein HSA.

Nicosulfuron is a sulfonylurea herbicide developed by DuPont that has been used successfully for weed control in maize. The binding mechanism and binding site identified in human serum albumin (HSA) with the use of fluorescence, circular dichroism (CD) and molecular modeling is the subject of this paper. From the CD, synchronous and three-dimensional fluorescence results, it was apparent that the interaction of nicosulfuron with HSA caused secondary structure changes in the protein. Fluorescence data revealed that the nicosulfuron induced the fluorescence quenching of HSA through a static quenching procedure. Thermodynamic analysis results implied the role of hydrophobic and hydrogen bonds interactions in stabilizing the nicosulfuron–HSA complex. Site marker competitive experiments showed the binding of nicosulfuron to HSA primarily took place in subdomain IIA (Sudlow’s site I), this corroborates the guanidine hydrochloride (GuHCl) induced denaturation of HSA, hydrophobic probe ANS displacement and molecular modeling results. In this work, the presented binding research extends our knowledge of the binding properties of sulfonylurea herbicides to the important plasma protein HSA.

Imidacloprid belongs to a major new class of insecticides, called neonicotinoids, which are accounting for 11–15% of the total insecticide market. The binding characteristics of insecticide imidacloprid with hemoglobin (Hb) have been studied by employing different spectroscopic techniques. The results proved the formation of complex between imidacloprid and Hb. Hydrophobic interaction and hydrogen bond dominated in the association reaction. Hydrophobic probe 8-anilino-1-naphthalenesulfonic acid (ANS) competitive experiments indicated that the binding of imidacloprid to Hb primarily took place in hydrophobic regions. The distance between Hb donor and acceptor imidacloprid was 4.88 nm as derived from Förster’s theory. Alternations of Hb secondary structure in the presence of imidacloprid were confirmed by synchronous fluorescence, circular dichroism (CD) and three-dimensional fluorescence spectra. This study enriches our understanding of toxic effect of imidacloprid to the physiologically important protein Hb.

Pyrazosulfuron-ethyl (PY) is a sulfonylurea herbicide developed by DuPont which has been widely used for weed control in cereals. The determination of PY binding affinity and binding site in human serum albumin (HSA) by spectroscopic methods is the subject of this work. From the fluorescence emission, circular dichroism and three-dimensional fluorescence results, the interaction of PY with HSA caused secondary structure changes in the protein. Fluorescence data demonstrated that the quenching of HSA fluorescence by PY was the result of the formation of HSA–PY complex at 1:1 molar ratio, a static mechanism was confirmed to lead to the fluorescence quenching. Hydrophobic probe 8-anilino-1-naphthalenesulfonic acid (ANS) displacement results show that hydrophobic patches are the major sites for PY binding on HSA. The thermodynamic parameters ΔH° and ΔS° were calculated to be −36.32 kJ mol−1 and −35.91 J mol−1 K−1, which illustrated van der Waals forces and hydrogen bonds interactions were the dominant intermolecular force in stabilizing the complex. Also, site marker competitive experiments showed that the binding of PY to HSA took place primarily in subdomain IIA (Sudlow's site I). What presented in this paper binding research enriches our knowledge of the interaction between sulfonylurea herbicides and the physiologically important protein HSA.

By partial least square regression, simple quantitative structure–activity relationship (QSAR) models were developed for the toxicity of polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs). Quantum chemical descriptors computed by semi-empirical PM3 method were used as predictor variables. Three optimal QSAR models are developed for 25 PCDDs, 35 PCDFs, 25 PCDDs and 35 PCDFs together, respectively. The cross-validated Qcum2 values for the three QSAR models of 25 PCDDs, 35 PCDFs, 25 PCDDs and 35 PCDFs together are 0.816, 0.629 and 0.603, respectively, indicating good predictive capabilities for the biological toxicity of these PCDD/Fs. The present study suggests that quantum chemical descriptors of POPs indeed govern the binding affinity of these chemicals for aryl hydrocarbon receptors. Moreover, different models contain different molecular descriptors to define respective equation, which suggests that the relationship between molecular structure and the binding affinity of these chemicals for aryl hydrocarbon receptors is complex.

Metsulfuron-methyl is a sulfonylurea herbicide widely used for broad-leaved weed control in cereals. The binding interaction between metsulfuron-methyl and human serum albumin was elucidated by fluorescence, circular dichroism and molecular modeling. The results showed that the alterations of albumin secondary structure in the presence of herbicide induced the slight unfolding of the polypeptide chain of albumin. Fluorescence data revealed that the fluorescence quenching of albumin by herbicide was the result of the formation of the albumin–herbicide complex and hydrophobic and hydrogen bonds interactions were the dominant intermolecular force in stabilizing the complex. Fluorescence probes studies implied that the binding of herbicide to albumin primarily took place in subdomain IIA (Sudlow's site I), and this also corroborates with molecular modeling simulations. This study highlights for the first time the binding mechanism, specific binding site and binding region of herbicide on albumin at the first time. Therefore, this investigation enriches our information of the interaction of sulfonylurea herbicide to the physiologically protein albumin.

The interactions between chloramphenicol and lysozyme were studied using fluorescence, UV/vis and circular dichroism spectra. The results proved the mechanism of fluorescence quenching of lysozyme by chloramphenicol is due to the formation of lysozyme–chloramphenicol complex. The thermodynamic parameters, enthalpy change (ΔH) and entropy change (ΔS) for the reaction, were calculated to be −12.41 kJ mol−1 and 37.99 J mol−1 K−1, which indicated that hydrophobic force and hydrogen bond were the dominant intermolecular forces in stabilizing the complex. The distance r = 3.99 nm between donor and acceptor was obtained according to Förster's theory. In addition, the alterations of lysozyme secondary structure in the presence of chloramphenicol were confirmed by the evidences from circular dichroism, synchronous and three-dimensional fluorescence spectroscopy. With the addition of chloramphenicol, the fluorescence intensity of lysozyme decreased regularly suggesting that chloramphenicol could interact with lysozyme and quench its intrinsic fluorescence.

The mechanism of interaction between C.I. Acid Red 2 and human serum albumin was studied using different spectroscopic methods. The binding constants for the formation of a complex between the dye and albumin were 2.557, 2.461 and 2.383 × 105 M−1 at 298, 304 and 310 K, respectively. The associated changes in enthalpy and entropy were −4.512 kJ mol−1 and 88.38 J mol−1 K−1, indicating that hydrophobic interactions as well as H-bonding were the dominant intermolecular forces stabilizing the complex. Site marker competitive experiments revealed that the binding of the dye to albumin occurred in subdomain IIA; the distance between dye and albumin was 3.91 nm according to fluorescence resonance energy transfer theory. Changes in the albumin secondary structure imparted by the dye were confirmed using synchronous fluorescence, electronic absorption, circular dichroism and three-dimensional fluorescence spectroscopy.

The interaction of C.I. Mordant Red 3 and bovine serum albumin was investigated using fluorescence and UV–vis absorption spectroscopy. Fluorescence quenching, from which binding parameters were evaluated, revealed that the quenching of the serum by C.I. Mordant Red 3 resulted from the formation of a dye–serum complex. The enthalpy and entropy changes for the reaction were −50.49 kJ mol−1 and −50.88 J mol−1 K−1 respectively. van der Waals forces and hydrogen bonds were the dominant intermolecular forces that stabilize the complex; the distance between donor and acceptor was 2.79 nm, according to Förster's non-radiative energy transfer theory. The effect of the dye upon the conformation of bovine serum albumin was analyzed using synchronous fluorescence spectrum.

In this paper, the interaction between barbital and bovine serum albumin (BSA) was investigated by the method of fluorescence spectroscopy under simulative physiological conditions. Fluorescence data revealed that the fluorescence quenching of BSA by barbital was the result of the formation of BSA–barbital complex, and the effective quenching constants (Ka) were 1.468×104, 1.445×104 and 1.403×104 M−1 at 297, 303 and 310 K, respectively. The thermodynamic parameters enthalpy change (ΔH) and entropy change (ΔS) for the reaction were calculated to be −2.679 kJ mol−1 and 70.76 J mol−1 K−1, respectively, according to the van’t Hoff equation. The results indicated that hydrophobic and electrostatic interactions were the dominant intermolecular force in stabilizing the complex. The results of synchronous fluorescence spectra showed that binding of barbital with BSA can induce conformational changes in BSA. In addition, the effects of Cu2+ and Zn2+ on the constants of BSA–barbital complex were also discussed.

The interaction between chloramphenicol and human serum albumin (HSA) was studied by fluorescence, UV/vis, circular dichroism (CD) and three-dimensional fluorescence spectroscopy. Fluorescence data revealed that the fluorescence quenching of HSA by chloramphenicol was the result of the formation of drug–HSA complex, and the effective quenching constants (Ka) were 2.852 × 104, 2.765 × 104, 2.638 × 104 and 2.542 × 104 M−1 at 287, 295, 303 and 311 K, respectively. The thermodynamic parameters, enthalpy change (ΔH) and entropy change (ΔS) for the reaction were calculated to be −3.634 kJ mol−1 and 72.66 J mol−1 K−1 according to van’t Hoff equation. The results indicated that the hydrophobic and electrostatic interactions played a major role in the binding of drug to HSA. The distance r between donor and acceptor was obtained to be 3.63 nm according to Förster’s theory. Site marker competitive experiments indicated that the binding of drug to HSA primarily took place in subdomain IIA. The alterations of HSA secondary structure in the presence of chloramphenicol were confirmed by the evidences from synchronous fluorescence, CD and three-dimensional fluorescence spectra. In addition, the effect of common ions on the binding constants of drug–HSA complex was also discussed.

The interaction of a N-methylated diaminotriphenylmethane dye, malachite green, with lysozyme was investigated by fluorescence spectroscopic techniques under physiological conditions. The binding parameters have been evaluated by fluorescence quenching methods. The results revealed that malachite green caused the fluorescence quenching of lysozyme through a static quenching procedure. The thermodynamic parameters like ΔH and ΔS were calculated to be −15.33 kJ mol−1 and 19.47 J mol−1 K−1 according to van’t Hoff equation, respectively, which proves main interaction between malachite green and lysozyme is hydrophobic forces and hydrogen bond contact. The distance r between donor (lysozyme) and acceptor (malachite green) was obtained to be 3.82 nm according to Fӧrster’s theory. The results of synchronous fluorescence, UV/vis and three-dimensional fluorescence spectra showed that binding of malachite green with lysozyme can induce conformational changes in lysozyme. In addition, the effects of common ions on the constants of lysozyme-malachite green complex were also discussed.

Phenosafranine is a toxic and recalcitrant compound, whose capacity to intercalate with double stranded DNA has been shown. In this contribution, a biophysical discuss on the conjugation of phenosafranine with two model proteins human serum albumin (HSA) and lysozyme (Lys) has been identified utilizing a combination of molecular modeling, steady state and time-resolved fluorescence and circular dichroism (CD) approaches. The accurate binding domain of phenosafranine in protein has been characterized from molecular modeling, subdomain IIIA of HSA and Trp-62, Trp-63 and Trp-108 residues of Lys was designated to possess high-affinity for this compound, the dominant forces in the protein–phenosafranine adduct are hydrogen bonds and π–π interactions, but hydrophobic interactions between dye and Lys are also not exclude. The data of fluorescence displayed that the complex of phenosafranine with protein produces quenching through static property, this corroborates the time-resolved fluorescence results that the ground state complex formation with a moderate affinity of 104 M−1. Moreover, via synchronous fluorescence, CD and three-dimensional fluorescence we indicated some extent of polypeptide chain of protein partially unfolding upon conjugation with phenosafranine. Through this work, we anticipate it can supply salient clues on the toxicological action of phenosafranine and other azines, which have analogous configuration with phenosafranine.Highlights► Our study supply salient clues on the toxicological action of toxic phenosafranine. ► Phenosafranine is situated within subdomain IIIA, Sudlow's site II on HSA. ► Trp-62, Trp-63 and Trp-108 residues on the Lys molecule are all close to dye. ► Static property of the phenosafranine induced quenching of protein Trp fluorescence. ► Protein spatial structure proved to be disturbed after complex with phenosafranine.

The immobilization of carboxylesterase from Spodoptera Litura in mesoporus molecular sieves was studied in this paper. Two different types of mesoporous sieves (MCM-41, SBA-15) were initially used as enzyme support to compare the efficiency of immobilization. The outcome showed the maximum enzyme loadings, twice as many in MCM-41, are 0.0267 L/g in SBA-15 support. The most efficient pH value for immobilization was at 6.5 for both types of the support and the optimal immobilizing time was 2 h for SBA-15 and 4 h for MCM-41. Hence, the SBA-15 mesoporous support was determined as the enzyme immobilization matrix to continue the investigation because of its superior properties for enzyme immobilization. Then a series of characters of the free and immobilized Carboxylesterase were compared. As a result, the immobilized enzyme was substantiated to be obviously more stable than the free enzyme under the variant conditions. The experiments of pesticides degradation show that, relative to degradation in natural environment, immobilization enzyme had a higher ability to reduce the organic compounds with ester bonds.