Dihydromyricetin (DMY), a natural flavonoid, was found to effectively inhibit tyrosinase activity in a mixed-type manner with an IC50 value of (3.66 ± 0.14) × 10−5 mol L−1. DMY combined with the dietary vitamin D3 at lower concentrations exhibited a synergistic effect on the inhibition of tyrosinase. The formation of a DMY–tyrosinase complex led to fluorescence quenching and conformational changes of tyrosinase, which was driven mainly by hydrophobic interactions and hydrogen bonds. The molecular simulation further found that DMY inserted into the active pocket of tyrosinase interacted with amino acid residues Tyr78, His85, and Ala323, occupying the catalytic center of tyrosinase to hinder entrance of the substrate, leading to the inhibition of tyrosinase. This study may provide a scientific foundation for screening effective tyrosinase inhibitors.

•Processing had significant effect on soluble conjugated phenolics of brown rice.•Processing decreased antioxidant activity of soluble conjugated phenolics.•Textured rice and rice noodle had more bound phenolics than brown rice.•Textured rice and rice noodle had higher bound antioxidant activities.The phytochemical profiles and antioxidant activity of free, soluble-conjugated, and bound fractions of brown rice and its processed products (textured rice, cooked rice and rice noodle) were studied. Nineteen phenolic acids were identified. Trans-ferulic acid was the most abundant monomeric phenolic acid with trans-trans-8-O-4′ diferulic acid being most abundant diferulic acid. Processing increased the content of free phenolic acids, but decreased the content of soluble-conjugated phenolic acids. The content of bound phenolic acids was increased by improved extrusion cooking technology and cooking, but not affected by rice noodle extrusion. The total phenolic contents and antioxidant activities of free and soluble-conjugated fractions were decreased after processing, whereas those of bound fraction were increased by improved extrusion cooking technology and cooking, but not affected by rice noodle extrusion. Results indicated that whole foods designed for reducing chronic disease risk need to consider the effects of processing on phytochemical profiles and antioxidant activity of whole grains.

•The binding of 8-MOP to trypsin was mainly driven by hydrophobic forces.•8-MOP interacted with the catalytic triplet in the active center of trypsin.•The binding of 8-MOP to trypsin induced the conformational changes of trypsin.•The kinetic process of the interaction was determined by MCR–ALS analysis.8-Methoxypsoralen (8-MOP) is a naturally occurring furanocoumarin with various biological activities. However, there is little information on the binding mechanism of 8-MOP with trypsin. Here, the interaction between 8-MOP and trypsin in vitro was determined by multi-spectroscopic methods combined with the multivariate curve resolution-alternating least squares (MCR–ALS) chemometrics approach. An expanded UV–vis spectral data matrix was analysed by MCR–ALS, the concentration profiles and pure spectra for the three reaction species (trypsin, 8-MOP and 8-MOP–trypsin) were obtained to monitor the interaction between 8-MOP and trypsin. The fluorescence data suggested that a static type of quenching mechanism occurred in the binding of 8-MOP to trypsin. Hydrophobic interaction dominated the formation of the 8-MOP–trypsin complex on account of the positive enthalpy and entropy changes, and trypsin had one high affinity binding site for 8-MOP with a binding constant of 3.81 × 104 L mol− 1 at 298 K. Analysis of three dimensional fluorescence, UV–vis absorption and circular dichroism spectra indicated that the addition of 8-MOP induced the rearrangement of the polypeptides carbonyl hydrogen-bonding network and the conformational changes in trypsin. The molecular docking predicted that 8-MOP interacted with the catalytic residues His57, Asp102 and Ser195 in trypsin. The binding patterns and trypsin conformational changes may result in the inhibition of trypsin activity. This study has provided insights into the binding mechanism of 8-MOP with trypsin.

•The Cu(II)–chrysin complex was synthesized and characterized.•The Cu(II)–chrysin complex reversibly inhibited XO activity in a mixed-type.•The binding of the complex to XO induced the conformational change of enzyme.•The complex inserted into active cavity of XO and blocked the landing of xanthine.Xanthine oxidase (XO) is a key enzyme catalyzing hypoxanthine to xanthine and then uric acid causing hyperuricemia. A Cu(II) complex of chrysin was synthesized and characterized by UV–vis absorption, Fourier transform infrared, nuclear magnetic resonance (1H NMR) and mass spectroscopy studies. The interaction of Cu(II)-complex with XO was investigated by spectroscopic methods and molecular simulation. The Cu(II)-chrysin complex exhibited a better inhibitory ability (IC50 = 0.82 ± 0.034 μM) against XO than its corresponding ligands chrysin and Cu2 + in a mix-competitive manner. The binding affinity of Cu(II)-chrysin complex with XO was much higher than that of chrysin. The hydrogen bonds and van der Waals forces played main roles in the binding. Analysis of circular dichroism spectra indicated that the complex induced the conformational change of XO. The molecular simulation found that the Cu(II)-chrysin complex inserted into the active cavity of XO with Cu acting as a bridge, occupying the catalytic center of the enzyme to avoid entry of the substrate xanthine, leading to the inhibition of XO. This study may provide new insights into the inhibition mechanism of the Cu(II)-chrysin complex as a promising XO inhibitor and its potential application for the treatment of hyperuricemia.Download high-res image (123KB)Download full-size image

Vanillin (VAN) and ethyl vanillin (EVA) are widely used food additives as flavor enhancers, but may have a potential security risk. In this study, the properties of binding of VAN or EVA with calf thymus DNA (ctDNA) were characterized by multi-spectroscopic methods, multivariate curve resolution-alternating least-squares (MCR–ALS) algorithm and molecular simulation. The concentration profiles for the components (VAN or EVA, ctDNA and VAN–ctDNA or EVA–ctDNA complex) by the MCR–ALS analysis showed that VAN or EVA interacted with ctDNA and formed VAN–ctDNA or EVA–ctDNA complex. The groove binding of VAN or EVA to ctDNA was supported by the results from viscosity measurements, melting studies, denaturation experiments, and competitive binding investigations. Analysis of the Fourier transform infrared spectra corroborated the prediction by molecular docking that VAN and EVA preferentially bound to thymine bases region of ctDNA. The circular dichroism and DNA cleavage assays indicated that both VAN and EVA induced conformational change (from B − like DNA structure toward to A − like form), but didn’t lead to a significant damage on DNA. The fluorescence quenching of Hoechst 33,258–ctDNA complex by VAN or EVA was a static quenching, and hydrogen bonding and van der Waals forces were main forces. This study has provided insights into the mechanism of interaction between VAN or EVA with ctDNA, and may also help better understand their potential toxicity with regard to food safety.

•Quercetin could inhibit both monophenolase and diphenolase activities of tyrosinase.•Hydrophobic interaction dominated the binding of quercetin to tyrosinase.•The binding of quercetin to tyrosinase induced conformational changes of tyrosinase.•Catechol structure of quercetin chelated the copper in the active site of tyrosinase.Quercetin, a flavonoid compound, was found to inhibit both monophenolase and diphenolase activities of tyrosinase, and its inhibition against diphenolase activity was in a reversible and competitive manner with an IC50 value of (3.08 ± 0.74) × 10− 5 mol L− 1. Quercetin bound to tyrosinase driven by hydrophobic interaction, thereby resulted in a conformational change of tyrosinase and its intrinsic fluorescence quenching. Tyrosinase had one binding site for quercetin with the binding constant in the order of magnitude of 104 L mol− 1. The molecular docking revealed that quercetin bound to the active site of tyrosinase and chelated a copper with the 3′, 4′-dihydroxy groups. It can be deduced that the chelation may prevent the entrance of substrate and then inhibit the catalytic activity of tyrosinase. These findings may be helpful to understand the inhibition mechanism of quercetin on tyrosinase and functional research of quercetin in the treatment of pigmentation disorders.Download high-res image (119KB)Download full-size image

Inhibition of α-glucosidase activity may suppress postprandial hyperglycemia. The inhibition kinetic analysis showed that apigenin reversibly inhibited α-glucosidase activity with an IC50 value of (10.5 ± 0.05) × 10–6 mol L–1, and the inhibition was in a noncompetitive manner through a monophasic kinetic process. The fluorescence quenching and conformational changes determined by fluorescence and circular dichroism were due to the formation of an α-glucosidase–apigenin complex, and the binding was mainly driven by hydrophobic interactions and hydrogen bonding. The molecular simulation showed that apigenin bound to a site close to the active site of α-glucosidase, which may induce the channel closure to prevent the access of substrate, eventually leading to the inhibition of α-glucosidase. Isobolographic analysis of the interaction between myricetin and apigenin or morin showed that both of them exhibited synergistic effects at low concentrations and tended to exhibit additive or antagonistic interaction at high concentrations.Keywords: apigenin; inhibition mechanism; isobologram; synergy; α-glucosidase;

Daphnetin is an important and naturally occurring coumarin compound with potent biological activities. Herein, the coordination between daphnetin and Cu2+ and the in vitro interaction of the daphnetin–Cu(II) complex with calf thymus DNA (ctDNA) were investigated by multivariate curve resolution-alternating least squares chemometrics combined with multiple spectroscopy, ctDNA thermal denaturation and viscosity studies, and visualized by molecular modeling. The results showed that the daphnetin–Cu(II) complex with a stoichiometry of 1:1 was formed and its fluorescence was quenched by ctDNA. The intercalation mode of the daphnetin–Cu(II) complex to ctDNA was demonstrated by the increases in DNA viscosity and melting temperature, and competitive binding between the daphnetin–Cu(II) complex and the fluorescence probe ethidium bromide with ctDNA. The analysis of Fourier transformed infrared spectra showed that the A–T bases region of ctDNA was the main binding site for the daphnetin–Cu(II) complex, and the interaction induced the reduction of helicity and base stacking, but the B-form of ctDNA was not changed. Moreover, a DNA cleavage assay suggested that the daphnetin–Cu(II) complex caused cleavage of plasmid DNA due to its strong binding to DNA. This study is expected to offer useful information for understanding the interaction mechanism of the daphnetin–Cu(II) complex with ctDNA, and may provide novel insights into the synergistic effect between daphnetin and Cu2+ in exerting their pharmacological function.

Dietary guidelines to promote health are usually based on the patterns’ prediction on disease risk of foods and nutrients. Overactivity of xanthine oxidase (XO) is the underlying cause of gout. Herein, the inhibitory kinetics and mechanism of dietary vitamins D3 and B2 on XO were investigated by multispectroscopic methods and a molecular modeling technique. The results showed that vitamin D3 competitively inhibited XO with an inhibition constant of 26.93 ± 0.42 μM by inserting into the active cavity of XO interacting with the surrounding amino acid residues through hydrogen bond and van der Waals forces. Vitamin D3 bound to XO thereby induced the structural compactness of XO which in turn hindered the binding of substrate xanthine to cause the inhibition on XO. Vitamin B2 exhibited a mixed-type inhibition by binding to the vicinity of the active cavity with an inhibition constant of 37.76 ± 0.87 μM through hydrophobic interactions and a feeble hydrogen bond, and it induced the unfolding of the XO structure and an increase of the flexible loops (β-turns and random coils) which might move to cover the active pocket and reduce the binding of the substrate xanthine, and then lead to a lower catalytic activity of the enzyme. In addition, vitamins D3 and B2 showed a synergistic effect on inhibiting the activity of XO in a certain range of concentration. These findings may provide new insights into the inhibitory mechanism of vitamins D3 and B2 on XO and functional research of the vitamins in the supplementary treatment of gout.

α-Glucosidase is a vital enzyme in carbohydrate metabolism. Over-expression of this enzyme is correlated with hyperglycemia. The inhibitory effect of vitamin D3 on α-glucosidase as well as its mechanism of action was investigated in this work. The results showed that vitamin D3 exhibited stronger inhibition on α-glucosidase than acarbose with the IC50 value of 1.28 × 10−4 mol L−1, and the inhibition was a mixed-type mechanism through a multiphase kinetic process. The inhibition constant was determined to be (5.66 ± 0.03) × 10−5 mol L−1. Vitamin D3 interacted with α-glucosidase by hydrophobic interactions, and molecular docking further verified that the inhibitor inserted into the active site pocket of α-glucosidase and interacted with the amino residues, which induced the rearrangement and conformational changes of α-glucosidase, and might move to cover the active pocket, hindering the binding of the substrate leading to the inhibition of the enzyme activity. Moreover, it was found that vitamin D3 combined with vitamin B1 or vitamin B2 exhibited significant synergistic effects on inhibition of α-glucosidase. This study has provided new insights into the role of vitamin D3 in inhibiting α-glucosidase catalysis and offered useful information on the dietary recommendation of vitamin D3 for the treatment of type 2 diabetes.

•DNA interaction is studied by spectroscopic methods and molecular docking.•The binding mode of TFL or FL to ctDNA is minor groove binding.•Both TFL and FL prefer to bind with A–T rich regions of ctDNA.•Molecular simulation confirms the binding mode and site of TFL or FL with ctDNA.•Both TFL and FL do not cause significant DNA cleavage.Tau-fluvalinate (TFL) and flumethrin (FL), widely used in agriculture and a class of synthetic pyrethroid pesticides with a similar structure, may cause a potential security risk. Herein, the modes of binding in vitro of TFL and FL with calf thymus DNA (ctDNA) were characterized by fluorescence, UV–vis absorption, circular dichroism (CD) and Fourier transform infrared (FT-IR) spectroscopy with the aid of viscosity measurements, melting analyses and molecular docking studies. The fluorescence titration indicated that both TFL and FL bound to ctDNA forming complexes through hydrogen bonding and van der Waals forces. The binding constants of TFL and FL with ctDNA were in the range of 104 L mol− 1, and FL exhibited a higher binding propensity than TFL. The iodide quenching effect, single/double-stranded DNA effects, and ctDNA melting and viscosity measurements demonstrated that the binding of both TFL and FL to ctDNA was groove mode. The FT-IR analyses suggested the A–T region of the minor groove of ctDNA as the preferential binding for TFL and FL, which was confirmed by the displacement assays with Hoechst 33258 probe, and the molecular docking visualized the specific binding. The changes in CD spectra indicated that both FL and TFL induced the perturbation on the base stacking and helicity of B-DNA, but the disturbance caused by FL was more obvious. Gel electrophoresis analyses indicated that both TFL and FL did not cause significant DNA cleavage. This study provides novel insights into the binding properties of TFL/FL with ctDNA and its toxic mechanisms.

The flavonoid family has been reported to possess a high potential for inhibition of xanthine oxidase (XO). This study concerned the structural aspects of inhibitory activities and binding affinities of flavonoids as XO inhibitors. The result indicated that the hydrophobic interaction was important in the binding of flavonoids to XO, and the XO inhibitory ability increased generally with increasing affinities within the class of flavones and flavonols. The planar structure and the C2═C3 double bonds of flavonoids were advantageous for binding to XO and for XO inhibition. Both the hydroxylation on ring B and the substitution at C3 were unfavorable for XO inhibition more profoundly than their XO affinity. The methylation greatly reduced the inhibition (0.75–3.07 times) but hardly affected the affinity. The bulky sugar substitutions of flavonoids decreased the inhibition (1.69–1.99 times) and lowered the affinities (4.20–9.22 times) to different degrees depending on the conjunction site.

Xanthine oxidase (XO), a key enzyme in purine catabolism, is widely distributed in human tissues. It can catalyze xanthine to generate uric acid and cause hyperuricemia and gout. Inhibition kinetics assay showed that kaempferol inhibited XO activity reversibly in a competitive manner. Strong fluorescence quenching and conformational changes of XO were found due to the formation of a kaempferol–XO complex, which was driven mainly by hydrophobic forces. The molecular docking further revealed that kaempferol inserted into the hydrophobic cavity of XO to interact with some amino acid residues. The main inhibition mechanism of kaempferol on XO activity may be due to the insertion of kaempferol into the active site of XO occupying the catalytic center of the enzyme to avoid the entrance of the substrate and inducing conformational changes of XO. In addition, luteolin exhibited a stronger synergistic effect with kaempferol than did morin at the lower concentration.

Resmethrin (RES) is a synthetic pyrethroid insecticide widely used to control pests in agriculture, but it may cause potential hazards to human health. The characteristics of the binding of RES with calf thymus DNA (ctDNA) were investigated in a physiological buffer (pH 7.4) by multiple spectroscopic methods combined with ctDNA melting and viscosity measurements, multivariate curve resolution-alternating least-squares (MCR-ALS) chemometrics and the molecular docking technique. The concentration profiles and the pure spectra for the reactive species (RES, ctDNA and RES–ctDNA complex), obtained through decomposing the augmented UV-vis spectral data matrix by the MCR-ALS approach, indicated that RES could bind to ctDNA and the reaction process could be quantitatively monitored. The RES molecules bound to ctDNA by groove binding, as evidenced by the negligible changes in the ctDNA melting temperature, viscosity and iodide quenching effect and the increase in the single-stranded DNA quenching effect. Fluorescence titration data indicated that the complexation of RES with ctDNA was mainly driven by hydrogen bonds and van der Waals forces, and had strong affinity. The changes in the Fourier transform infrared spectra of ctDNA suggested that RES molecules preferentially bound to the G–C region of ctDNA, which was consistent with the prediction of the molecular docking. The circular dichroism spectral analysis indicated that RES induced a decrease in the right-handed helicity of ctDNA. The DNA cleavage assay showed that RES did not cleave the pUC18 plasmid DNA. This study offers a comprehensive picture of RES–ctDNA interaction, which may provide insights into the toxicological effect of the insecticide.

Dimethyl phthalate (DMP) is widely used as a plasticizer in industrial processes and has been reported to possess potential toxicity to the human body. In this study, the interaction between DMP and trypsin in vitro was investigated. The results of fluorescence, UV–vis, circular dichroism, and Fourier transform infrared spectra along with cyclic voltammetric measurements indicated that the remarkable fluorescence quenching and conformational changes of trypsin resulted from the formation of a DMP–trypsin complex, which was driven mainly by hydrophobic interactions. The molecular docking and trypsin activity assay showed that DMP primarily interacted with the catalytic triad of trypsin and led to the inhibition of trypsin activity. The dimensions of the individual trypsin molecules were found to become larger after binding with DMP by atomic force microscopy imaging. This study offers a comprehensive picture of DMP–trypsin interaction, which is expected to provide insights into the toxicological effect of DMP.

Erythrosine (Ery) is an artificial colorant extensively used in the food industry, but may have a potential safety risk. In this study, the characteristics of the interaction in vitro between Ery and herring sperm DNA (hsDNA) were determined by multi-spectroscopic techniques and docking simulations. The multivariate curve resolution-alternating least squares chemometrics approach was used to process the expanded UV-vis spectral data matrix, and both the equilibrium concentration profiles and the pure spectra for the components (Ery, hsDNA and Ery–hsDNA complex) extracted from the highly overlapping composite response were obtained simultaneously to monitor the Ery–hsDNA interaction process. Partial intercalation as the dominant mode of Ery binding to hsDNA was found based on the hypochromism and red shift effect of Ery, stronger double DNA effect and decrease in hsDNA viscosity. The Fourier transform infrared and nuclear magnetic resonance spectra showed that the benzene ring of Ery most likely inserted into the guanine and cytosine bases of hsDNA, and molecular docking predicted and visualized the specific binding. The circular dichroism and DNA cleavage assays suggested that Ery at high concentrations may perturb B-DNA conformation and cause slight DNA cleavage. This study has provided insights into the binding mechanism of Ery with hsDNA and potential hazards of the colorant.

•TBMP prefers to form the 1:1 complex with Hp-βCD.•The binding mode of TBMP to ctDNA is confirmed to be an intercalative mode.•A specific binding mainly exists between TBMP and the G–C rich region of ctDNA.•MCR-ALS provides quantitative information for binding progress of TBMP with ctDNA.•The formed inclusion complex of TBMP–Hp-βCD decomposes in the presence of ctDNA.The characteristics of binding of 2-tert-butyl-4-methylphenol (TBMP), a synthetic phenolic antioxidant with hydroxypropyl-β-cyclodextrin (Hp-βCD) and calf thymus DNA (ctDNA) were investigated by multi-spectroscopic techniques and molecular simulation. The results indicated that TBMP preferred to form a 1:1 inclusion complex with Hp-βCD, with an inclusion constant of determined to be 7.15 × 103 L mol−1. The intercalative mode of TBMP to ctDNA was supported by ctDNA melting temperature and relative viscosity studies, salt quenching effect, competitive binding with methylene blue and molecular modeling. The changes in Fourier transformed infrared (FT-IR) and circular dichroism (CD) spectra suggested that TBMP mainly bound to the G–C rich region with inducing a significant perturbation in B-like DNA structure. It was also found that Hp-βCD decreased the binding ability of TBMP with ctDNA, but did not affect the interactive mode between TBMP and ctDNA, and the formed inclusion complex of TBMP–Hp-βCD decomposed in the presence of ctDNA. This study may provide insights into the mechanism of binding of TBMP with ctDNA and the role of Hp-βCD in the TBMP–ctDNA interaction.

•Genistein reversibly inhibits xanthine oxidase (XO) in a competitive manner.•The interaction is mainly driven by hydrophobic interactions and hydrogen bonds.•Genistein interacts with several residues located within the active pocket of XO.•Genistein induces the conformational changes of XO.Genistein (Gen), widely distributed in soybean, is proved to be important in homeostasis in the human body. Herein, the inhibitory mechanism of Gen against xanthine oxidase (XO) was studied through multispectroscopic methods and molecular simulation. The inhibition kinetics showed that Gen competitively inhibited XO with an inhibition constant of (1.39 ± 0.11) μM by competing with xanthine for binding to the active site of XO. Fluorescence titration study suggested that the fluorescence quenching mechanism of XO was static, resulting from the formation of a Gen–XO complex at one fold site. The calculated thermodynamic parameters revealed that the interaction process was driven mainly by hydrophobic interactions and hydrogen bonds with affinity of (5.24 ± 0.02) × 104 L mol− 1. Conformational analyses demonstrated that the microenvironment and the secondary structure of XO were changed upon binding of Gen. The molecular docking displayed that Gen bound to the active cavity of XO by interacting with the surrounding amino acid residues (Leu648, Phe649, Glu802, Ser876, Glu879, Arg880, Phe914, Phe1009, Thr1010 and Phe1013). Thus, the inhibition may be attributed to the insertion of Gen into the active site of XO occupying the catalytic center of the enzyme to avoid entry of the substrate and inducing conformational changes of XO (more compact), which was further unfavorable for forming the active cavity and further reduced the landing and oxidation of substrate. This study may offer novel insights into the inhibition mechanism of Gen on XO.

•Psoralen mainly interacts with the catalytic triplet in the active center of trypsin.•The binding of psoralen to trypsin is mainly driven by hydrophobic forces.•Psoralen binding to trypsin induces the conformational changes of the enzyme.•The kinetic processes of the interaction is determined by MCR–ALS analysis.Psoralen (PSO) is a naturally occurring furanocoumarin with a variety of pharmacological activities, however very limited information on the interaction of PSO with trypsin is available. In this study, the binding characteristics between PSO and trypsin at physiological pH were investigated using a combination of fluorescence, UV–vis absorption, circular dichroism (CD), Fourier transform infrared (FT-IR) spectroscopic, chemometric and molecular modeling approaches. It was found that the fluorescence quenching of trypsin by PSO was a static quenching procedure, ascribing the formation of a PSO–trypsin complex. The binding of PSO to trypsin was driven mainly by hydrophobic forces as the positive enthalpy change and entropy change values. The molecular docking showed that PSO inserted into the active site pocket of trypsin to interact with the catalytic residues His57, Asp102 and Ser195 and may cause a decrease in trypsin activity. The results of CD and FT-IR spectra along with the temperature-induced denaturation studies indicated that the addition of PSO to trypsin led to the changes in the secondary structure of the enzyme. The concentration profiles and spectra of the three components (PSO, trypsin, and PSO–trypsin complex) obtained by multivariate curve resolution-alternating least squares analysis exhibited the kinetic processes of PSO–trypsin interaction. This study will be helpful to understand the mechanism of PSO that affects the conformation and activity of trypsin in biological processes.

•Morin reversibly inhibits tyrosinase activity in a competitive manner.•Morin binds to tyrosinase mainly by hydrogen bonds and van der Waals forces.•Morin interacts with several residues located within the active pocket of tyrosinase.•The binding of morin to tyrosinase induces conformational changes of the enzyme.Tyrosinase is a key enzyme in the production of melanin in the human body, excessive accumulation of melanin can lead to skin disorders. Morin is an important bioactive flavonoid compound widely distributed in plants and foods of plant origin. In this study, the inhibitory kinetics of morin on tyrosinase and their binding mechanism were determined using spectroscopic and molecular docking techniques. The results indicate that morin reversibly inhibited tyrosinase in a competitive manner through a multi-phase kinetic process. Morin was found to bind to tyrosinase at a single binding site mainly by hydrogen bonds and van der Waals forces. Analysis of circular dichroism spectra revealed that the binding of morin to tyrosinase induced rearrangement and conformational changes of the enzyme. Moreover, molecular docking results suggested that morin competitively bound to the active site of tyrosinase with the substrate levodopa.

The binding characteristics of sodium saccharin (SSA), an artificial sweetener, with calf thymus DNA (ctDNA) were investigated by multispectroscopic techniques, chemometrics, and molecular simulation. A combined fluorescence and UV–vis spectroscopic data matrix was resolved by the multivariate curve resolution–alternating least-squares (MCR–ALS) chemometrics algorithm. The MCR–ALS analysis extracted simultaneously the concentration profiles and spectra for the three components (SSA, ctDNA, and SSA–ctDNA complex) to quantitatively monitor the SSA–ctDNA interaction, which is difficult to perform by conventional spectroscopic approach. The binding mode of SSA to ctDNA was principally through groove binding as revealed by ctDNA melting temperature studies, viscosity measurements, and iodide and salt quenching effects. Analysis of the Fourier transform infrared and circular dichroism spectra as well as molecular docking indicated that SSA preferentially bound to the guanine base of ctDNA and led to a transformation from B-like DNA structure to A-like conformation. Moreover, gel electrophoresis results suggested that SSA did not induce any significant cleavage in plasmid DNA.

A combination of fluorescence, UV–Vis absorption, circular dichroism (CD), Fourier transform infrared (FT-IR) spectroscopic and molecular modeling approaches was employed to investigate the interaction between toddalolactone (TDT) and human serum albumin (HSA) at physiological buffer conditions (pH 7.4). Fluorescence titration suggests that the mechanism of the fluorescence quenching of HSA is static, resulting from the formation of a TDT–HSA complex. Binding parameters calculated from the modified Stern–Volmer equation show that TDT binds to HSA with high affinity. Negative enthalpy change and positive entropy change values suggest that the binding process is primarily driven by hydrophobic interactions and hydrogen bonds. The binding of TDT to HSA results in an increase in the surface hydrophobicity of HSA. The binding distance between the Trp-214 residue (donor) and TDT (acceptor) was determined to be 4.18 nm based on the Förster theory of non-radioactive energy transfer. Displacement studies of site markers reveal that the binding site of TDT to HSA is located in the subdomain IIA (Sudlow’s site I). Furthermore, the molecular docking results corroborate and illustrate the specific binding mode and binding site. Analysis of UV–Vis absorption, CD and FT-IR spectra demonstrated that TDT induced a small alteration of the protein’s conformation.

Hydrolysis of the pesticide folpet [N-(trichloromethylthio) phthalimide] in aqueous solution in the absence or presence of calf thymus DNA (ctDNA) was investigated using UV–Vis absorption spectroscopy, and the interactions of folpet and its hydrolyzates with ctDNA were determined by fluorescence and circular dichroism spectroscopy, coupled with viscosity and melting temperature measurements. The absorption spectra data was further analyzed by alternate least squares, a chemometrics method, and the concentration profiles of the reacting species (folpet, unstable intermediate, phthalimide and phthalic acid) and their pure component spectra were simultaneously extracted to monitor the hydrolytic process. It was found that the hydrolytic process consists of at least two steps, generation of an unstable intermediate and production of its end hydrolyzates, phthalimide and phthalic acid. Addition of ctDNA significantly affects the hydrolysis of folpet. The results from the competitive binding with intercalator ethidium bromide, ctDNA melting and viscosity measurements, and circular dichroism studies indicate that folpet and the intermediate can intercalate into the double-helix of DNA, phthalic acid is bound to DNA by a partial intercalation, while phthalimide does not show binding to ctDNA. Moreover, the binding of folpet (or the intermediate) and phthalic acid to ctDNA induced structural changes of the DNA.

•The binding mode of BHA to ctDNA is an intercalative binding.•MCR-ALS approach provides quantitative information for the interaction progress.•A specific binding mainly exists between BHA and the G–C rich region of ctDNA.•The molecular docking predicts the binding mode and sites of BHA with ctDNA.•BHA binding to DNA does not induce significantly plasmid DNA damage.The binding properties of food antioxidant butylated hydroxyanisole (BHA) associated with calf thymus DNA (ctDNA) in physiological buffer (pH 7.4) were investigated. Experimental results based on fluorescence, UV–vis absorption, circular dichroism (CD), viscosity measurements and autodocking techniques confirmed the intercalation binding between BHA and ctDNA. The changes in Fourier transform infrared spectra of ctDNA induced by BHA suggested that BHA was more prone to bind to G–C rich region of ctDNA, which was further ascertained with the molecular docking studies. Analysis of the CD spectra indicated that this binding interaction led to a transformation from B-like DNA structure toward A-like conformation. The complexation of BHA with ctDNA was driven mainly by hydrogen bonds and hydrophobic forces. The binding constants of the BHA–ctDNA complex were calculated to be 2.03 × 104, 1.92 × 104 and 1.59 × 104 L mol−1 at 298, 304 and 310 K, respectively. Gel electrophoresis results suggested that intercalated BHA molecules did not significantly affect plasmid DNA. Moreover, the concentration profiles and the spectra for the three reaction components (BHA, ctDNA, and BHA–ctDNA complex) of the system by resolving the augmented UV–vis spectral data matrix with the use of multivariate curve resolution-alternating least squares approach provided quantitative data to estimate the progress of BHA–ctDNA interaction. This study is expected to provide new insights into the mechanism of interaction between BHA and ctDNA.Graphical abstract

The mechanism of interaction between food dye amaranth and human serum albumin (HSA) in physiological buffer (pH 7.4) was investigated by fluorescence, UV–vis absorption, circular dichroism (CD), and Fourier transform infrared (FT-IR) spectroscopy. Results obtained from analysis of fluorescence spectra indicated that amaranth had a strong ability to quench the intrinsic fluorescence of HSA through a static quenching procedure. The negative value of enthalpy change and positive value of entropy change elucidated that the binding of amaranth to HSA was driven mainly by hydrophobic and hydrogen bonding interactions. The surface hydrophobicity of HSA increased after binding with amaranth. The binding distance between HSA and amaranth was estimated to be 3.03 nm and subdomain IIA (Sudlow site I) was the primary binding site for amaranth on HSA. The results of CD and FT-IR spectra showed that binding of amaranth to HSA induced conformational changes of HSA.Highlights► We investigate the interaction of amaranth with HSA by multispectroscopic methods. ► The florescence of HSA is quenched by amaranth through a static quenching mechanism. ► The surface hydrophobicity of HSA increases upon interaction with amaranth. ► Amaranth is located in subdomain IIA (Site I) of HSA. ► The binding of amaranth to HSA can induce changes in the secondary structure of HSA.

The binding of permethrin (PE) with calf thymus DNA (ctDNA) in physiological buffer (pH 7.4) was investigated by ultraviolet–visible (UV–vis) absorption, fluorescence, circular dichroism (CD), and Fourier transform infrared (FT–IR) spectroscopy merging with multivariate curve resolution–alternating least-squares (MCR–ALS) chemometrics approach. The MCR–ALS was applied to resolve the combined spectroscopic data matrix, which was obtained by UV–vis and fluorescence methods. The concentration profiles of PE, ctDNA, and PE–ctDNA complex and their pure spectra were then successfully obtained. The PE molecular was found to be able to intercalate into the base pairs of ctDNA as evidenced by decreases in resonance light-scattering signal and iodide-quenching effect and increase in ctDNA viscosity. The results of FT–IR spectra indicated that PE was prone to bind to G–C base pairs of ctDNA, and the molecular docking studies were used to validate and clarify the specific binding. The observed changes in CD signals revealed that the DNA turned into a more highly wound form of B-conformation. The calculated thermodynamic parameters, enthalpy change (ΔH°) and entropy change (ΔS°), suggested that hydrogen bonds and van der Waals forces played a predominant role in the binding of PE to ctDNA.

The mechanism of paeoniflorin binding to calf thymus DNA in physiological buffer (pH 7.4) was investigated by multispectroscopic methods including UV–vis absorption, fluorescence, circular dichroism (CD) and Fourier transform infrared (FT-IR) spectroscopy, coupled with viscosity measurements and DNA melting techniques. The results suggested that paeoniflorin molecules could bind to DNA via groove binding mode as evidenced by no significant change in iodide quenching effect, increase in single-stranded DNA (ssDNA) quenching effect, and almost unchanged relative viscosity and melting temperature of DNA. The observed changes in CD signals revealed that DNA remains in the B-conformation. Further, the displacement experiments with Hoechst 33258 probe and the results of FT-IR spectra indicated that paeoniflorin mainly binds in the region of rich A–T base pairs of DNA. The thermodynamic parameters, enthalpy change (ΔH°) and entropy change (ΔS°) were calculated to be –30.09±0.18 kJ mol−1 and –14.07±0.61 J mol−1 K−1 by the van't Hoff equation, suggesting that hydrogen bond and van der Waals forces play a predominant role in the binding of paeoniflorin to DNA.Highlights► The binding mode of paeoniflorin to calf thymus DNA is the minor groove binding. ► Paeoniflorin mainly binds in the region of rich A–T base pairs of DNA. ► The binding does not alter the native B-conformation of DNA. ► The binding is driven mainly by hydrogen bonds and van der Waals forces.

•The binding mode of cyanazine to ctDNA is an intercalation binding.•Cyanazine mainly binds to G–C bases of ctDNA.•ALS algorithm is applied to resolve the absorption spectral data array.•The concentration information for cyanazine, EB and ctDNA–EB is provided.•Cyanazine intercalates into ctDNA by substituting for EB in the ctDNA–EB complex.The interaction between cyanazine and calf thymus DNA (ctDNA) in physiological buffer (pH 7.4) was investigated with the use of ethidium bromide (EB) as a spectral probe by UV–vis absorption, fluorescence, circular dichroism (CD) and Fourier transform infrared (FT-IR) spectroscopy, as well as viscosity measurements. The results revealed that intercalation binding should be the interaction mode of cyanazine to ctDNA and the binding constant was obtained to be 2.65 × 104 L mol−1 at 298 K. Analysis of the FT-IR spectra suggested that cyanazine mainly bound to guanine and cytosine of ctDNA bases and led to ctDNA duplex aggregation at higher cyanazine concentrations. The thermodynamic parameters, enthalpy change (ΔH°) and entropy change (ΔS°) suggested that hydrophobic interactions and hydrogen bonds played a predominant role in the binding of cyanazine to ctDNA. Furthermore, a chemometrics approach, the alternate least squares (ALS) algorithm, was applied to resolve the measured absorption spectral data array of the competitive reaction between cyanazine and EB with ctDNA, and the results provided simultaneously the concentration information and corresponding pure spectra for the three reaction components, cyanazine, EB and ctDNA–EB. The results obtained from the ALS analysis indicated that cyanazine intercalated into ctDNA by substituting for EB in the ctDNA–EB complex.

•The binding mode between BHT and ctDNA is an intercalation.•BHT most likely binds to adenine and thymine base pairs of ctDNA.•The binding of BHT to ctDNA does not alter the native B-conformation of ctDNA.•The binding is driven mainly by hydrogen bonds and van der Waals force.The binding properties of butylated hydroxytoluene (BHT) with calf thymus DNA (ctDNA) in simulated physiological buffer (pH 7.4) were investigated using ethidium bromide (EB) dye as a fluorescence probe by various spectroscopic techniques including UV–vis absorption, fluorescence, circular dichroism (CD), and Fourier transform infrared (FT-IR) spectroscopy along with ctDNA melting studies and viscosity measurements. It was found that the binding of BHT to ctDNA could decrease the absorption intensity of ctDNA, significantly increase melting temperature and relative viscosity of ctDNA, and induce the changes in CD spectra. Moreover, the competitive binding studies showed that BHT was able to displace EB from the bound ctDNA–EB complex. All the experimental results indicated that the binding mode between BHT and ctDNA was an intercalation. The association constants between BHT and ctDNA were evaluated to be (4.78 ± 0.04) × 103, (2.86 ± 0.02) × 103 and (1.80 ± 0.04) × 103 L mol−1 at 298, 304, 310 K, respectively. Further, the FT-IR analysis revealed that BHT was more prone to interact with adenine and thymine base pairs, and no significant conformational transition of ctDNA occurred. Thermodynamic analysis of the binding data showed that the binding process was primarily driven by hydrogen bonds and van der Waals forces, as the values of the enthalpy change and the entropy change were calculated to be −62.47 ± 0.07 kJ mol−1 and −139.22 ± 0.22 J mol−1 K−1, respectively.

The interaction between maltol, a food additive, and bovine serum albumin (BSA) under simulated physiological conditions was investigated by fluorescence, UV–Vis absorption, circular dichroism (CD) and Fourier transform infrared (FT-IR) spectroscopy. The results suggested that the fluorescence quenching of BSA by maltol was a static procedure forming a maltol–BSA complex. The positive values of enthalpy change and entropy change indicated that hydrophobic interactions played a predominant role in the interaction of maltol with BSA. The competitive experiments of site markers revealed that the binding of maltol to BSA mainly took place in subdomain IIA (Sudlow site I). The binding distance between maltol and BSA was 3.01 nm based on the Förster theory of non-radioactive energy transfer. Moreover, the results of UV–Vis, synchronous fluorescence, CD and FT-IR spectra demonstrated that the microenvironment and the secondary structure of BSA were changed in the presence of maltol.Highlights► Interaction of maltol and bovine serum albumin (BSA) is investigated by multispectroscopic techniques. ► Maltol can quench the fluorescence of BSA through a static quenching procedure. ► The binding of maltol to BSA is driven mainly by hydrophobic interactions. ► Subdomain IIA (Sudlow site I) of BSA is found to be the main binding site for maltol. ► The secondary structure of BSA has been changed in the presence of maltol.

The interaction of indigo carmine (IC) with calf thymus DNA in physiological buffer (pH 7.4), using ethidium bromide (EB) dye as a fluorescence probe, was investigated by ultraviolet–visible absorption, fluorescence, and circular dichroism (CD) spectroscopy, coupled with viscosity measurements and DNA-melting studies. Hypochromicity of the absorption spectra of IC and enhancement in fluorescence polarization of IC were observed with the addition of DNA. Moreover, the binding of IC to DNA was able to decrease iodide and single-stranded DNA (ssDNA) quenching effects, increase the melting temperature and relative viscosity of DNA, and induce the changes in CD spectra of DNA. All of the evidence indicated that IC interacted with DNA in the mode of intercalative binding. Furthermore, the three-way synchronous fluorescence spectra data obtained from the interaction between IC and DNA–EB were resolved by parallel factor analysis (PARAFAC), and the results provided simultaneously the concentration information and the pure spectra for the three reaction components (IC, EB, and DNA–EB) of the system at equilibrium. This PARAFAC demonstrated that the intercalation of IC molecules into DNA proceeded by substituting for EB in the DNA–EB complex. The calculated thermodynamic parameters, ΔH° and ΔS°, suggested that both hydrophobic interactions and hydrogen bonds played a predominant role in the binding of IC to DNA.

The binding mechanism of molecular interaction between diosmetin and human serum albumin (HSA) in a pH 7.4 phosphate buffer was studied using atomic force microscopy (AFM) and various spectroscopic techniques including fluorescence, resonance light scattering (RLS), UV–vis absorption, circular dichroism (CD), and Fourier transform infrared (FT–IR) spectroscopy. Fluorescence data revealed that the fluorescence quenching of HSA by diosmetin was a static quenching procedure. The binding constants and number of binding sites were evaluated at different temperatures. The RLS spectra and AFM images showed that the dimension of the individual HSA molecules were larger after interaction with diosmetin. The thermodynamic parameters, ΔH° and ΔS° were calculated to be −24.56 kJ mol–1 and 14.67 J mol–1 K–1, respectively, suggesting that the binding of diosmtin to HSA was driven mainly by hydrophobic interactions and hydrogen bonds. The displacement studies and denaturation experiments in the presence of urea indicated site I as the main binding site for diosmetin on HSA. The binding distance between diosmetin and HSA was determined to be 3.54 nm based on the Förster theory. Analysis of CD and FT–IR spectra demonstrated that HSA conformation was slightly altered in the presence of diosmetin.

The interaction between carbaryl and calf thymus DNA (ctDNA) was investigated under simulated physiological conditions (Tris–HCl buffer of pH 7.4) using ethidium bromide (EB) dye as a probe by UV–vis absorption, fluorescence and circular dichroism (CD) spectroscopy, as well as DNA melting studies and viscosity measurements. It can be concluded that carbaryl molecules could intercalate into the base pairs of DNA as evidenced by hyperchromic effect of absorption spectra, decreases in iodide fluorescence quenching effect, induced CD spectral changes, and significant increases in melting temperature and relative viscosity of DNA. The binding constants and thermodynamic parameters of carbaryl with DNA were obtained by the fluorescence quenching method. Furthermore, a chemometrics approach, parallel factor analysis (PARAFAC), was applied to resolve the measured three-way synchronous fluorescence spectral data matrix of the competitive interaction between carbaryl and EB with DNA, and the results provided simultaneously the concentration profiles and corresponding pure spectra for three reaction components (carbaryl, EB and DNA–EB complex) of the kinetic system at equilibrium. This PARAFAC analysis demonstrated the intercalation of carbaryl to the DNA helix by substituting for EB in the DNA–EB complex.Highlights► The interaction between carbaryl and calf thymus DNA is investigated. ► The binding mode of carbaryl to DNA is an intercalation binding. ► The parallel factor analysis confirms the intercalation of carbaryl to the DNA helix. ► The binding is driven mainly by hydrogen bond and van der Waals forces.

The interaction between farrerol and calf thymus DNA in a pH 7.4 Tris-HCl buffer was investigated with the use of neutral red (NR) dye as a spectral probe by UV–vis absorption, fluorescence, and circular dichroism (CD) spectroscopy, as well as viscosity measurements and DNA melting techniques. It was found that farrerol molecules could intercalate into the base pairs of DNA as evidenced by decreases in iodide quenching effect and single-stranded DNA (ssDNA) quenching effect, induced CD spectral changes, and significant increases in relative viscosity and denaturation temperature of DNA. Furthermore, the spectral data matrix of the competitive reaction between farrerol and NR with DNA was resolved with an alternative least-squares (ALS) algorithm, and the concentration profiles in the reaction and the corresponding pure spectra for three species (farrerol, NR, and DNA–NR complex) were obtained. This ALS analysis demonstrated the intercalation of farrerol to the DNA by substituting for NR in the DNA–NR complex. Moreover, the thermodynamic parameters enthalpy change (ΔH°) and entropy change (ΔS°) were calculated to be −16.49 ± 0.51 kJ mol–1 and 32.47 ± 1.02 J mol–1 K–1 via the van’t Hoff equation, which suggested that the binding of farrerol to DNA was driven mainly by hydrophobic interactions and hydrogen bonds.

The interaction of puerarin with human serum albumin (HSA) in pH 7.4 Tris–HCl buffer has been investigated by fluorescence, Fourier transform infrared (FT-IR) and circular dichroism (CD) spectroscopy. The results revealed the presence of static type of quenching mechanism in the binding of puerarin to HSA. The association constants (Ka) between puerarin and HSA were obtained according to Modified Stern–Volmer equation. The calculated thermodynamic parameters indicated that the binding of puerarin to HSA was driven mainly by hydrophobic interaction. The competitive experiments of site markers suggested that the binding site of puerarin to HSA was located in the region of subdomain IIA (sudlow site I). Further, a chemometrics approach, parallel factor analysis (PARAFAC), was applied to resolve the measured three-way synchronous fluorescence spectra data of the competitive interaction between puerarin and warfarin with HSA. The concentration information for the three reaction components, warfarin, puerarin and puerarin−HSA, in the system at equilibrium was obtained simultaneously. The PARAFAC analysis indicated that puerarin in the puerarin–HSA complex was displaced by warfarin, which confirmed the binding site of puerarin to HSA was located in site I. Moreover, the results of CD and FT-IR spectra demonstrated that the secondary structure of HSA was changed in the presence of puerarin.Highlights► Puerarin can quench the fluorescence of human serum albumin (HSA). ► The HSA fluorescence is quenched by puerarin through a static quenching mechanism. ► The binding of puerarin to HSA is driven mainly by hydrophobic interaction. ► The parallel factor analysis confirms that puerarin is located in site I of HSA. ► The binding of puerarin to HSA induces changes in the secondary structure of HSA.

The interaction between vitexin and human serum albumin (HSA) has been studied by using different spectroscopic techniques viz., fluorescence, UV–vis absorption, circular dichroism (CD) and Fourier transform infrared (FT–IR) spectroscopy under simulated physiological conditions. Fluorescence results revealed the presence of static type of quenching mechanism in the binding of vitexin to HSA. The binding constants (Ka) between vitexin and HSA were obtained according to the modified Stern–Volmer equation. The thermodynamic parameters, enthalpy change (ΔH) and entropy change (ΔS) were calculated to be –57.29 kJ mol−1 and –99.01 J mol−1 K−1 via the van't Hoff equation, which indicated that the interaction of vitexin with HSA was driven mainly by hydrogen bond and van der Waals forces. Fluorescence anisotropy data showed that warfarin and vitexin shared a common binding site I corresponding to the subdomain IIA of HSA. The binding distance (r) between the donor (HSA) and the acceptor (vitexin) was 4.16 nm based on the Förster theory of non-radioactive energy transfer. In addition, the results of synchronous fluorescence, CD and FT–IR spectra demonstrated that the microenvironment and the secondary structure of HSA were changed in the presence of vitexin.Research highlights► We investigate the binding mechanism of vitexin to human serum albumin (HSA) by different multi-spectroscopic techniques under simulated physiological conditions. ► Vitexin can strongly quench the fluorescence of HSA through a static quenching mechanism. ► The interaction of vitexin with HSA is driven mainly by hydrogen bond and van der Waals forces. ► The binding distance between HSA and vitexin is 4.16 nm, and vitexin is mainly located in the region of site I (subdomain IIA). ► The binding of vitexin to HSA can induce conformational changes of HSA.

A spectrophotometric method for the simultaneous determination of two herbicides, atrazine and cyanazine, is described for the first time based on their reaction with p-aminoacetophenone in the presence of pyridine in hydrochloric acid medium. The absorption spectra were measured in the wavelength range of 400–600 nm. The optimized method indicated that individual analytes followed Beer's law in the concentration ranges for atrazine and cyanazine were 0.2–3.5 mg L−1 and 0.3–5.0 mg L−1, and the limits of detection for atrazine and cyanazine were 0.099 and 0.15 mg L−1, respectively. The original and first-derivative absorption spectra of the binary mixtures were performed as a pre-treatment on the calibration matrices prior to the application of chemometric models such as classical least squares (CLS), principal component regression (PCR), partial least squares (PLS). The analytical results obtained by using these chemometric methods were evaluated on the basis of percent relative prediction error and recovery. It was found that the application of PCR and PLS models for first-derivative absorbance data gave the satisfactory results. The proposed methods were successfully applied for the simultaneous determination of the two herbicides in several food samples.

The interaction between pirimicarb and calf thymus DNA in physiological buffer (pH 7.4) was investigated with the use of Neutral Red (NR) dye as a spectral probe by UV–vis absorption, fluorescence and circular dichroism (CD) spectroscopy, as well as viscosity measurements and DNA melting techniques. The results revealed that an intercalation binding should be the interaction mode of pirimicarb to DNA. CD spectra indicated that pirimicarb induced conformational changes of DNA. The binding constants of pirimicarb with DNA were obtained by the fluorescence quenching method. The thermodynamic parameters, enthalpy change (ΔHθ) and entropy change (ΔSθ) were calculated to be −52.13 ± 2.04 kJ mol−1 and −108.8 ± 6.72 J mol−1 K−1 according to the van’t Hoff equation, which suggested that hydrogen bonds and van der Waals forces might play a major role in the binding of pirimicarb to DNA. Further, the alternative least squares (ALS) method was applied to resolve a complex two-way array of the absorption spectra data, which provided simultaneously the concentration information for the three reaction components, pirimicarb, NR and DNA–NR. This ALS analysis indicated that the intercalation of pirimicarb into the DNA by substituting for NR in the DNA–NR complex.

The mechanism and conformational changes of farrerol binding to bovine serum albumin (BSA) were studied by spectroscopic methods including fluorescence quenching technique, UV–vis absorption, circular dichroism (CD) spectroscopy and Fourier transform infrared (FT-IR) spectroscopy under simulative physiological conditions. The results of fluorescence titration revealed that farrerol could strongly quench the intrinsic fluorescence of BSA through a static quenching procedure. The thermodynamic parameters enthalpy change and entropy change for the binding were calculated to be −29.92 kJ mol−1 and 5.06 J mol−1 K−1 according to the van’t Hoff equation, which suggested that the both hydrophobic interactions and hydrogen bonds play major role in the binding of farrerol to BSA. The binding distance r deduced from the efficiency of energy transfer was 3.11 nm for farrerol–BSA system. The displacement experiments of site markers and the results of fluorescence anisotropy showed that warfarin and farrerol shared a common binding site I corresponding to the subdomain IIA of BSA. Furthermore, the studies of synchronous fluorescence, CD and FT-IR spectroscopy showed that the binding of farrerol to BSA induced conformational changes in BSA.Graphical abstract. The mechanism and conformational change of farrerol binding to bovine serum, albumin (BSA) were studied by spectroscopic methods. The quenching mechanism, binding constants, thermodynamic parameters, and binding distance were obtained.Highlights►We investigate the mechanism and conformational changes of farrerol binding to BSA. ► The binding constants, binding sites and thermodynamic parameters were calculated. ► Farrerol is located in subdomain IIA (site I) of BSA. ► The binding of farrerol to BSA induced changes in the secondary structure of BSA.

The binding interaction between alpinetin and bovine serum albumin (BSA) in physiological buffer solution (pH 7.4) was investigated by fluorescence, UV–vis spectroscopy and Fourier transform infrared (FT-IR) spectroscopy. It was proved from fluorescence spectra that the fluorescence quenching of BSA by alpinetin was probably a result of the formation of BSA–alpinetin complexes, and the binding constant (Ka) were determined according to the modified Stern–Volmer equation. The thermodynamic parameters, enthalpy change (ΔH) and entropy change (ΔS), were calculated to be 22.10 kJ mol−1 and 166.04 J mol−1 K−1, respectively, which indicated that the interaction between alpinetin and BSA was driven mainly by hydrophobic interaction. Moreover, the competitive experiments of site markers suggested that the binding site of alpinetin to BSA was located in the region of subdomain IIA (sudlow site I). The binding distance (r) between the donor (BSA) and the acceptor (alpinetin) was 3.32 nm based on the Förster theory of non-radioactive energy transfer. In addition, the results of synchronous fluorescence and FT-IR spectra demonstrated that the microenvironment and the secondary structure of BSA were changed in the presence of alpinetin.

The interaction between kaempferol–Eu3+ complex and calf thymus DNA was investigated in physiological buffer (pH 7.4) using the Neutral Red (NR) dye as a spectral probe by UV–vis absorption and fluorescence spectroscopy, as well as viscosity measurements and DNA melting techniques. The results indicated that kaempferol–Eu3+ complex can bind to DNA and the major binding mode is intercalative binding. The binding constants K and number of binding sites n of kaempferol–Eu3+ complex with DNA were obtained by fluorescence quenching method. The thermodynamic parameters (ΔHθ, ΔSθ and ΔGθ) were calculated from the fluorescence data measured at three different temperatures, which showed that the binding of kaempferol–Eu3+ complex to DNA was driven mainly by hydrophobic interaction. Furthermore, the alternative least squares (ALS) method was applied to resolve a complex two-way array of the absorption spectra data, which provided simultaneously the concentration information for the three reaction components, NR, kaempferol–Eu3+ and DNA–NR. This ALS analysis indicated that the intercalation of the kaempferol–Eu3+ complex into the DNA proceeds by exchanging with the NR probe.

The interaction of chrysin with bovine serum albumin (BSA) in physiological buffer solution (pH 7.4) was studied by fluorescence, UV/vis absorption and resonance light scattering (RLS) spectroscopy. The experimental results showed that there was a strong fluorescence quenching of BSA by chrysin. The probable quenching mechanism of fluorescence of BSA by chrysin was a static quenching by forming the BSA–chrysin complex. The addition of increasing chrysin to BSA solution led to the gradual enhancement in RLS intensity, implying the formation of an aggregate in solution. The binding constants K and number of binding sites n of chrysin with BSA were obtained by fluorescence quenching method. The thermodynamic parameters of the interaction of chrysin with BSA were measured according to the van’s Hoff equation. The enthalpy change (ΔHθ) and the entropy change (ΔSθ) were calculated to be 39.19 kJ mol−1, 211.91 J mol−1 K−1 respectively, which indicated that the interaction between chrysin and BSA was driven mainly by hydrophobic interaction. The binding was shown to be spontaneous at the standard state because the changes in standard Gibbs free energy (ΔGθ) values were negative. The binding distance of chrysin from the tryptophan residue in BSA was calculated to be 2.44 nm based on the Förster theory of non-radiation energy transfer. The results of synchronous fluorescence spectra demonstrated that chrysin induced a conformational change of BSA. In addition, the effect of some inorganic ions on the binding constants of chrysin with BSA was also investigated.

The interaction between morin–Eu(III) complex and calf thymus DNA in physiological buffer (pH 7.4) was investigated using UV–vis spectrophotometry, fluorescence spectroscopy, viscosity measurements and DNA melting techniques. Hypochromicity and red shift of the absorption spectra of morin–Eu(III) complex were observed in the presence of DNA, and the fluorescence intensity of morin–Eu(III) complex was greatly enhanced with the addition of DNA. Moreover, fluorescence quenching and blue shift of the emission peak were seen in the DNA–ethidium bromide (EB) system when morin–Eu(III) complex was added. The relative viscosity of DNA increased with the addition of morin–Eu(III) complex, whereas the value of melting temperature of DNA–EB system decreased in the presence of morin–Eu(III) complex. All these results indicated that morin–Eu(III) complex can bind to DNA and the major binding mode is intercalative binding. The 3:1 morin:Eu(III) complex (estimated binding constant = 2.36 × 106 L mol−1) is stabilized by intercalation into the DNA. The calculated binding constants of morin–Eu(III) complex with DNA at 292, 301 and 310 K were 7.47 × 104, 8.89 × 104 and 1.13 × 105 L mol−1, respectively. The thermodynamic parameters were also obtained: ΔHθ was 20.14 kJ mol−1 > 0 and ΔSθ was 161.70 J mol−1 K−1 > 0, suggesting that hydrophobic force plays a major role in the binding of morin–Eu(III) complex to DNA.

The interaction between icariin and human serum albumin (HSA) in physiological buffer (pH 7.4) was investigated by fluorescence and UV–Vis absorption spectroscopy. Results obtained from analysis of fluorescence spectrum and fluorescence intensity indicated that icariin has a strong ability to quench the intrinsic fluorescence of HSA through a static quenching procedure. The thermodynamic parameters, ΔHθ and ΔSθ, were calculated to be 12.29 kJ mol−1 > 0, and 47.08 J mol−1 K−1 > 0, respectively, which suggested that hydrophobic force plays a major role in the reaction of icariin with HSA. The binding constants of icariin with HSA were determined at different temperatures by fluorescence quenching method. The distance r between donor (HSA) and acceptor (icariin) was calculated to be 4.18 nm based on Förster’s non-radiative energy transfer theory. The results of synchronous fluorescence spectra and three-dimensional fluorescence spectra showed that binding of icariin to HSA can induce conformational changes in HSA.

The interaction between irisflorentin (IFR) and bovine serum albumin (BSA) in physiological buffer (pH = 7.4) was investigated by fluorescence quenching technique and UV/vis absorption spectroscopy. The results of fluorescence titration revealed that IFR could strongly quench the intrinsic fluorescence of BSA through a dynamic quenching procedure. The apparent binding constants KA and number of binding sites n of IFR with BSA were obtained by fluorescence quenching method. The thermodynamic parameters, enthalpy change (ΔHθ) and entropy change (ΔSθ), were calculated to be 18.45 kJ mol−1 >0 and 149.72 J mol−1 K−1 >0, respectively, which indicated that the interaction of IFR with BSA was driven mainly by hydrophobic forces. The process of binding was a spontaneous process in which Gibbs free energy change was negative. The distance r between donor (BSA) and acceptor (IFR) was calculated to be 3.88 nm based on Förster’s non-radiative energy transfer theory. The results of synchronous fluorescence spectra showed that binding of IFR with BSA can induce conformational changes in BSA.

The interaction between aminocarb and calf thymus DNA in physiological buffer (pH 7.4) was investigated by UV–vis spectrophotometry, fluorescence spectroscopy, viscosity measurements and DNA melting techniques. The absorption spectra of aminocarb with DNA showed a slight blue shift and hypochromic effect. Using ethidium bromide (EB) as a fluorescence probe, fluorescence quenching of the emission peak was seen in the DNA–EB system when aminocarb was added. The fluorescence polarization was gradually increased with increasing amounts of DNA. The value of melting temperature of DNA increased in the presence of aminocarb. Moreover, the relative viscosity of DNA increased with the addition of aminocarb. All the evidences indicated that the binding mode of aminocarb with DNA was an intercalative binding. The binding constants of aminocarb with DNA were determined. The calculated thermodynamic parameters suggested that the binding of aminocarb to DNA was driven mainly by hydrogen bond and van der Waals.

•Chrysin inhibits xanthine oxidase (XO) activity reversibly in a competitive type.•Chrysin interacts with several residues located within the active cavity of XO.•Chrysin competes with allopurinol for the same biding site of XO.•The binding of chrysin to XO induces conformational changes of the enzyme.•Chrysin and apigenin showed an additive effect on inhibition of XO.Chrysin, a bioactive flavonoid, was investigated for its potential to inhibit the activity of xanthine oxidase (XO), a key enzyme catalyzing xanthine to uric acid and finally causing gout. The kinetic analysis showed that chrysin possessed a strong inhibition on XO ability in a reversible competitive manner with IC50 value of (1.26 ± 0.04) × 10−6 mol L−1. The results of fluorescence titrations indicated that chrysin bound to XO with high affinity, and the interaction was predominately driven by hydrogen bonds and van der Waals forces. Analysis of circular dichroism demonstrated that chrysin induced the conformational change of XO with increases in α-helix and β-sheet and reductions in β-turn and random coil structures. Molecular simulation revealed that chrysin interacted with the amino acid residues Leu648, Phe649, Glu802, Leu873, Ser876, Glu879, Arg880, Phe1009, Thr1010, Val1011 and Phe1013 located within the active cavity of XO. The mechanism of chrysin on XO activity may be the insertion of chrysin into the active site occupying the catalytic center of XO to avoid the entrance of xanthine and causing conformational changes in XO. Furthermore, the interaction assays indicated that chrysin and its structural analog apigenin exhibited an additive effect on inhibition of XO.

ObjectiveTo characterize the pharmacokinetics and distribution profiles of deltamethrin in miniature pig tissues by gas chromatography-mass spectrometry (GC-MS).MethodsPharmacokinetics and distribution of deltamethrin in blood and tissues of 30 miniature pigs were studied by GC-MS after oral administration of deltamethrin (5 mg/kg bw). Data were processed by 3P97 software.ResultsThe serum deltamethrin level was significantly lower in tissues than in blood of miniature pigs. The AUC0-72 h, Cmax, of deltamethrin were 555.330±316.987 ng h/mL and 17.861±11.129 ng/mL, respectively. The Tmax, of deltamethrin was 6.004±3.131 h.ConclusionThe metabolism of deltamethrin in miniature pigs is fit for a one-compartment model with a weighting function of 1/C2. Deltamethrin is rapidly hydrolyzed and accumulated in miniature pig tissues.

•The fluorescence quenching of HSA by prometryn was a static quenching.•The binding of prometryn to HSA mainly took place in subdomain IIA (site I).•Molecular modeling studies confirmed the binding site and binding mode.•The binding of prometryn to HSA induced the conformational alteration of HSA.Prometryn possesses much potential hazard to environment because of its chemical stability and biological toxicity. Here, the binding properties of prometryn with human serum albumin (HSA) and the protein structural changes were determined under simulative physiological conditions (pH 7.4) by multispectroscopic methods including fluorescence, UV–vis absorption, Fourier transform infrared (FT-IR) and circular dichroism (CD) spectroscopy, coupled with molecular modeling technique. The result of fluorescence titration suggested that the fluorescence quenching of HSA by prometryn was considered as a static quenching procedure. The negative enthalpy change (ΔH○) and positive entropy change (ΔS○) values indicated that the binding process was governed mainly by hydrophobic interactions and hydrogen bonds. The site marker displacement experiments suggested the location of prometryn binding to HSA was Sudlow’s site I in subdomain IIA. Furthermore, molecular docking studies revealed prometryn can bind in the large hydrophobic activity of subdomain IIA. Analysis of UV–vis absorption, synchronous fluorescence, CD and FT-IR spectra demonstrated that the addition of prometryn resulted in rearrangement and conformational alteration of HSA with reduction in α-helix and increases in β-sheet, β-turn and random coil structures. This work provided reasonable model helping us further understand the transportation, distribution and toxicity effect of prometryn when it spreads into human blood serum.Download full-size image

The interaction of puerarin with human serum albumin (HSA) in pH 7.4 Tris–HCl buffer has been investigated by fluorescence, Fourier transform infrared (FT-IR) and circular dichroism (CD) spectroscopy. The results revealed the presence of static type of quenching mechanism in the binding of puerarin to HSA. The association constants (Ka) between puerarin and HSA were obtained according to Modified Stern–Volmer equation. The calculated thermodynamic parameters indicated that the binding of puerarin to HSA was driven mainly by hydrophobic interaction. The competitive experiments of site markers suggested that the binding site of puerarin to HSA was located in the region of subdomain IIA (sudlow site I). Further, a chemometrics approach, parallel factor analysis (PARAFAC), was applied to resolve the measured three-way synchronous fluorescence spectra data of the competitive interaction between puerarin and warfarin with HSA. The concentration information for the three reaction components, warfarin, puerarin and puerarin−HSA, in the system at equilibrium was obtained simultaneously. The PARAFAC analysis indicated that puerarin in the puerarin–HSA complex was displaced by warfarin, which confirmed the binding site of puerarin to HSA was located in site I. Moreover, the results of CD and FT-IR spectra demonstrated that the secondary structure of HSA was changed in the presence of puerarin.Highlights► Puerarin can quench the fluorescence of human serum albumin (HSA). ► The HSA fluorescence is quenched by puerarin through a static quenching mechanism. ► The binding of puerarin to HSA is driven mainly by hydrophobic interaction. ► The parallel factor analysis confirms that puerarin is located in site I of HSA. ► The binding of puerarin to HSA induces changes in the secondary structure of HSA.

•The binding mode between DEHP and ctDNA was groove binding.•DEHP most likely bound to the A–T rich region of ctDNA.•Molecular docking illustrated the specific binding.•The interaction process of DEHP with ctDNA was monitored by MCR–ALS approach.In this study, the interaction between di-(2-ethylhexyl) phthalate (DEHP) and calf thymus DNA (ctDNA) was investigated by a combination of multispectroscopic methods, chemometrics algorithm, cyclic voltammetry and molecular simulation. The concentration profiles of the components obtained from resolving the UV–vis absorption data by multivariate curve resolution–alternating least-squares (MCR–ALS) provided a basic evidence for the formation of DEHP–ctDNA complex. Furthermore, the groove binding of DEHP to ctDNA was evidenced by the results from iodide quenching effect, single-stranded DNA quenching effect, melting studies, viscosity measurements and cyclic voltammetry. The binding constant of the complex was in the order of magnitudes of 104 L mol−1, and hydrophobic forces were inferred to drive the binding process. Analysis of Fourier transform infrared spectra suggested that DEHP preferentially bound to A–T rich region of ctDNA in the minor groove, and these results further confirmed by molecular docking. The circular dichroism spectra indicated that DEHP induced a decrease in base stacking degree and an increase in right-handed helicity of ctDNA, but did not cause a significant damage in DNA. This study may improve the understanding of interaction between DEHP and ctDNA and help evaluate the toxicological effect of DEHP.

A microwave-assisted enzymatic extraction (MAEE) method was developed and optimized to enhance the polyphenols extraction yield from waste peanut shells. The optimum conditions were as follows: irradiation time 2.6 min, amount of cellulase 0.81 wt.%, a pH of 5.5, and incubation at 66 °C for 2.0 h. Under these conditions, the extraction yield of total polyphenols could reach 1.75 ± 0.06%, which was higher than other extraction methods including heat-refluxing extraction, ultrasonic-assisted extraction and enzyme-assisted extraction. The structural changes of the plant material after different extractions observed by scanning electron microscopy provided visual evidence of the disruption effect. Moreover, the crude extract was then purified by NKA-9 resin, the polyphenols content in the purified extract increased to 62.73%. The antioxidant activities of the crude and purified polyphenols extract were evaluated by DPPH and hydroxyl radicals, reducing power and β-carotene bleaching test. The antibacterial activities of purified extract were also tested using Oxford cup method. The results indicated that the MAEE method was efficient and environment-friendly, and the polyphenols have significant antioxidant and antibacterial activities, which can be used as a source of potential antioxidant and preservative.Graphical abstractDownload high-res image (296KB)Download full-size imageHighlights► The polyphenols of peanut shell are extracted by microwave-assisted enzymatic method. ► The response surface methodology is used to optimize the polyphenols extraction. ► The extraction yield of polyphenols by different methods is compared. ► Microwave irradiation cause the disruptions of cell walls of the plant material. ► The MAEE extract exhibits potential antioxidant and antibacterial activities.