Elucidating the Interaction of Indole-3-Propionic Acid and Calf Thymus DNA: Multispectroscopic and Computational Modeling Approaches.

Indole-3-propionic acid (IPA) is a plant growth regulator with good specificity and long action. IPA may be harmful to human health because of its accumulation in vegetables and fruits. Therefore, in this study, the properties of the interaction between calf thymus DNA (ctDNA) and IPA were systematically explored using multispectroscopic and computational modeling approaches. Analysis of fluorescence spectra showed that IPA binding to ctDNA to spontaneously form a complex was mainly driven by hydrogen bonds and hydrophobic interaction. DNA melting analysis, viscosity analysis, DNA cleavage study, and circular dichroism measurement revealed the groove binding of IPA to ctDNA and showed that the binding did not significantly change ctDNA confirmation. Furthermore, molecular docking found that IPA attached in the A-T rich minor groove region of the DNA. Molecular dynamics simulation showed that DNA and IPA formed a stable complex and IPA caused slight fluctuations for the residues at the binding site. Gel electrophoresis experiments showed that IPA did not significantly disrupt the DNA structure. These findings may provide useful information on the potential toxicological effects and environmental risk assessments of IPA residue in food at the molecular level.

Effect of narrative videos and mini-VR games in nursing smoking cessation training on empathy and self-efficacy of smoking cessation counseling: A randomized controlled trial.

BACKGROUND: Empathy and self-efficacy for smoking cessation counseling can be enhanced through smoking cessation training. Narrative videos and virtual reality (VR) games have been applied in medical education, but their application in smoking cessation training is limited and understudied. OBJECTIVES: To evaluate the effect of smokers' narrative videos and mini-VR games on nursing students' empathy towards smokers (State Empathy Scale), confidence in practicing empathy, self-efficacy in smoking cessation counseling, and learning satisfaction. DESIGN: An open-labeled randomized controlled trial, registration number: NCT05440877 (ClinicalTrials.gov). SETTING AND PARTICIPANTS: Master of nursing students from the University in Hong Kong who enrolled in a smoking cessation course. METHODS: All students attended a tutorial including case-based discussions and role-play. The intervention group could additionally access the narrative videos and mini-VR games of smokers' cases. Linear mixed models and Cohen's d were used to evaluate the intervention effect on the self-reported learning outcomes after the intervention. RESULTS: 26 students enrolled in this trial, with 13 in each trial arm. All completed the trial. Post-test state empathy scores were significantly higher in the control group than in the intervention group (Cohen's d = 0.814, p = 0.049). No significant group differences were observed in the change of confidence in practicing empathy (ß = -11.154, p = 0.073), self-efficacy (ß = 4.846, p = 0.096), and students' learning satisfaction (Cohen's d = 0.041, p = 0.917). Both groups showed a significant increase in self-efficacy post-test (p < 0.001). CONCLUSIONS: Narrative videos showing smoking scenes and reluctance to quit, and our mini-VR games may weaken nursing students' empathy towards smokers. Smoking cessation training involving narrative videos should be modified. Debriefing and guidance to understand smokers' difficulties and express empathy are needed. VR games for smoking cessation training can involve more frame stories and challenging tasks to increase engagement.

Clinical characteristics and risk factors for bacterial co-infections in COVID-19 patients: A retrospective study.

OBJECTIVE: This study aimed to analyse the bacterial composition, distribution, drug sensitivity, and clinical characteristics of patients with coronavirus disease 2019 (COVID-19) who develop bacterial co-infections. METHODS: We conducted a retrospective study of 184 patients with COVID-19 admitted between December 2022 and January 2023. Data on gender, age, length of hospital stay, pneumonia classification, underlying diseases, invasive surgery, hormone therapy, inflammation indicators, and other relevant information were collected. Samples of sputum, bronchoscopy sputum, alveolar lavage fluid, middle urine, puncture fluid, wound secretions, and blood were collected for pathogen isolation, identification, and drug sensitivity testing. RESULTS: The majority of patients with COVID-19 with bacterial co-infection were elderly and had underlying diseases. Invasive surgery and hormone therapy were identified as risk factors for co-infections. Laboratory analysis showed reduced lymphocyte counts and elevated levels of C-reactive protein and procalcitonin. The most common pathogens in co-infections were Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae, Acinetobacter baumannii, and Pseudomonas aeruginosa. The detection rate of drug-resistant strains, including methicillin-resistant S. aureus, carbapenem-resistant K. pneumoniae, carbapenem-resistant A. baumannii, carbapenem-resistant P. aeruginosa, and carbapenem-resistant E. coli, increased with the severity of pneumonia. CONCLUSION: Respiratory tract infections were the most common site of bacterial co-infection in patients with COVID-19. Severe cases were more susceptible to multidrug-resistant pathogens, leading to a higher mortality rate. Timely control and prevention of co-infection are crucial for improving the prognosis of patients with COVID-19.

Baicalein glycymicelle ophthalmic solution: Preparation, in vitro antimicrobial activities, and antimicrobial mechanism evaluations.

The purpose of this study was to develop a novel baicalein (BAI) loaded glycymicelle ophthalmic solution with small molecule phytochemical glycyrrhizin as nanocarriers and to explore this solution's potential as an antimicrobial agent against ocular infections. The optimized BAI glycymicelles had a high encapsulation efficiency (98.76 ±â€¯1.25 %), a small particle size (54.38 ±â€¯2.41 nm), a uniform size distribution (polydispersity index = 0.293 ±â€¯0.083), and a zeta potential of -28.3 ±â€¯1.17 mV. The BAI glycymicelle ophthalmic solution exhibited an excellent short-term storage stability. BAI glycymicelles significantly increased the apparent solubility and in vitro release capability of BAI. The BAI glycymicelle ophthalmic solution exhibited no hen's egg-chorioallantoic membrane' irritation and strong in vivo ocular tolerance in rabbits. The BAI glycymicelles noticeably enhanced the in vivo corneal permeation. The BAI glycymicelles also precipitated increased in vitro antioxidant activity and significantly improved in vitro antipathogen activities. Various antimicrobial mechanisms, including the destruction of the bacterial cell wall, damage to the bacterial cell membranes, interruptions to the biofilm structure, and the apoptosis of bacteria, were inflicted on BAI glycymicelles. These findings provided useful knowledge regarding the development of a novel ophthalmic solution and formulation of BAI.

The mediating role of psychological capital on the relationship between authentic leadership and innovative behaviour among Chinese nurses.

AIM: To explore the mediating role of psychological capital between authentic leadership and innovative behaviour among Chinese nurses. DESIGN: A cross-sectional study. METHODS: In December 2021, online surveys were delivered among nurses from 37 hospitals in Anhui Province by convenience sampling approach. The data was collected using the Chinese version of the Authentic Leadership Questionnaire, Nurse Psychological Capital Questionnaire and Nurse Innovative Behaviour Scale. The structural equation model and bootstrap test examined the mediating role of psychological capital between authentic leadership and innovative behaviour. RESULTS: 3495 Chinese nurses from 37 Anhui Province hospitals participated in this study. The mean item score of authentic leadership was 3.25 (SD 0.83), psychological capital was 4.85 (SD 0.89), and innovative behaviour was 2.82 (SD 0.84). Authentic leadership perceived by nurses, psychological capital, and innovative behaviour were statistically significant and positively correlated with each other (r = 0.524 ~ 0.806, p < 0.01). Psychological capital significantly mediated the association between authentic leadership and innovative behaviour (ß = 0.449, p < 0.001), and its mediating effect accounted for 76.75% of the total effect. CONCLUSION: Results suggest the importance of developing nurse managers' authentic leadership to foster nurses' psychological capital and innovation behaviour.

The learning outcomes of smoking cessation training in undergraduate nursing students: A systematic review.

AIM: To systematically evaluate the effect of smoking cessation training on nursing students' learning outcomes. DESIGN: This systematic review followed the PRISMA guidelines and was registered with PROSPERO (CRD42022373280). METHODS: Ten electronic English and Chinese databases were searched to identify articles on nursing students' smoking cessation training from inception to October 2022. Medical Education Research Study Quality Instrument was used to assess the methodological quality of eligible studies. RESULTS: Twelve moderate methodological quality studies (three randomized controlled trials and nine quasi-experimental studies) with 2114 undergraduate nursing students were included. Teaching strategies included didactic lectures, collaborative learning, blended learning and the service-learning approach. Eight studies showed that the training significantly increased nursing students' knowledge of smoking cessation (p < 0.05) and three of them showed a large effect size (Cohen's d: 0.83-1.31). Seven studies showed that training enhanced students' attitudes/motivation towards smoking cessation interventions significantly (p < 0.05) and two of them showed a large effect size (Cohen's d: 1.11-1.84). Nine studies reported that students' self-efficacy improved significantly after training (p < 0.05) and six of them showed a large effect size (Cohen's d: 0.98-2.18). Two one-group pre-post studies showed training significantly improved students' practice of 5 A's behavior (p < 0.05), with a small effect size (Cohen's d < 0.50). CONCLUSIONS: This review showed that smoking cessation training had a positive and large effect on undergraduate nursing students' knowledge, attitudes/motivation and self-efficacy towards smoking cessation intervention, but the effect of changing the practice of 5 A's was modest. We noted that very few RCTs were done and most studies only used subjective measures. More robust experimental studies with long-term follow-up are warranted in evaluating nursing students' practice/behavior of smoking cessation intervention and patients' quitting outcomes. Blended and collaborative learning are recommended in smoking cessation education.

Hesperitin-Copper(II) Complex Regulates the NLRP3 Pathway and Attenuates Hyperuricemia and Renal Inflammation.

BACKGROUND: Hyperuricaemia (HUA) is a disorder of purine metabolism in the body. We previously synthesized a hesperitin (Hsp)-Cu(II) complex and found that the complex possessed strong uric acid (UA)-reducing activity in vitro. In this study we further explored the complex's UA-lowering and nephroprotective effects in vivo. METHODS: A mouse with HUA was used to investigate the complex's hypouricemic and nephroprotective effects via biochemical analysis, RT-PCR, and Western blot. RESULTS: Hsp-Cu(II) complex markedly decreased the serum UA level and restored kidney tissue damage to normal in HUA mice. Meanwhile, the complex inhibited liver adenosine deaminase (ADA) and xanthine oxidase (XO) activities to reduce UA synthesis and modulated the protein expression of urate transporters to promote UA excretion. Hsp-Cu(II) treatment significantly suppressed oxidative stress and inflammatory in the kidney, reduced the contents of cytokines and inhibited the activation of the nucleotide-binding oligomerization domain (NOD)-like receptor thermal protein domain associated protein 3 (NLRP3) inflammatory pathway. CONCLUSIONS: Hsp-Cu(II) complex reduced serum UA and protected kidneys from renal inflammatory damage and oxidative stress by modulating the NLRP3 pathway. Hsp-Cu(II) complex may be a promising dietary supplement or nutraceutical for the therapy of hyperuricemia.

Characterization of a novel recombinant D-mannose isomerase from Bifidobacterium bifidum and its catalytic mechanism.

Due to the increasing demand for health-conscious and environmentally friendly products, D-mannose has gained significant attention as a natural, low-calorie sweetener. The use of D-mannose isomerases (D-MIases) for D-mannose production has emerged as a prominent area of research, offering superior advantages compared with conventional methods such as plant extraction and chemical synthesis. In this study, a gene encoding D-MIase was cloned from Bifidobacterium and expressed in E. coli BL21 (DE3). The heterologously expressed enzyme, Bifi-mannose, formed a trimer with a molecular weight of 146.3 kDa and a melting temperature (Tm) of 63.39 ± 1.3 °C. Bifi-mannose exhibited optimal catalytic activity at pH 7.5 and 55 °C, and retained more than 80% of its activity after a 3-hour incubation at 55 °C, demonstrating excellent thermal stability. The Km, Vmax, and kcat/Km values of Bifi-mannose for D-fructose isomerization were determined as 538.7 ± 62.5 mM, 11.7 ± 0.9 µmol·mg1·s1, and 1.02 ± 0.3 mM1·s1, respectively. Notably, under optimized conditions, catalytic yields of 29.4, 87.1, and 148.5 mg·mL1 were achieved when using 100, 300, and 500 mg·mL1 of D-fructose as substrates, resulting in a high conversion rate (29%). Furthermore, kinetic parameters and molecular docking studies revealed that His387 residue primarily participates in the opening of the pyranose ring, while His253 acts as a basic catalyst in the isomerization process.

External factors affecting the linear and nonlinear rheological behavior of oleogel-based emulsions.

This study reported oleogel-based emulsions (OGEs, W/O) stabilized by carnauba wax. The effects of different external factors (heating temperature, crystallization temperature, and shear application during crystallization) on the microstructure and linear/nonlinear rheological properties of OGEs were investigated. Microstructural observation suggested that the OGEs had a uniform droplet distribution, and the carnauba wax crystals trapped oil in the continuous phase. The gelatinized oil phase allowed the OGEs to have a solid appearance and typical yielding behavior. The small amplitude oscillation shear analysis showed that lower heating temperature, higher crystallization temperature, and suitable shear application resulted in a stronger, more stable, and tighter packed network structure and better resistance to deformation of the OGEs. For nonlinear behavior, the elastic dominant behavior of OGEs transformed into viscous dominant behavior at large strain amplitudes, accompanied by more energy dissipation, strain stiffening, and a transition from shear thickening to shear thinning.

The mediating role of psychological capital on the relationship between authentic leadership and nurses' caring behavior: a cross-sectional study.

BACKGROUND: Caring behavior among nurses would have an impact on patient outcomes. External organizational job resources and personal internal psychological resources are correlated to nurses' caring behavior. Authentic leadership and psychological capital were shown to be correlated with nurses' caring behavior in previous studies. However, the relationships among the three are nevertheless unclear. This study aimed to examine if psychological capital could act as a mediator between nursing managers' authentic leadership and nurses' caring behavior. METHODS: In December 2021, a total of 3,662 nurses were recruited from 37 hospitals in Anhui Province, China. They filled out online surveys, including general demographic information, the Authentic Leadership Questionnaire, the Psychological Capital Questionnaire, and the Caring Behavior Inventory. Structural Equation Modeling and the bootstrapping procedure were used to examine the mediating role of psychological capital. RESULTS: The scores of authentic leadership, psychological capital, and caring behavior of 3,495 nurses were 52.04 ± 13.24, 96.89 ± 17.78, and 104.28 ± 17.01, respectively. Psychological capital significantly mediated the relationship between authentic leadership and nurses' caring behavior (ß = 0.378, p < 0.001, 95% confidence interval: 0.350 ~ 0.402), which made up 78.75% of the total impact (0.480). CONCLUSION: The findings of this study suggested that nursing managers should develop an authentic leadership style, which can effectively improve nurses' caring behaviors toward patients in clinical practice. Meanwhile, nursing leaders should strengthen nurses' psychological evaluation and training, and promote nurses' caring behavior in clinical settings.

Inhibition of toll-like receptor 4 activation by apigenin and chrysin via competition for sites and conformational changes.

The activation of toll-like receptor 4 (TLR4) signaling is crucial for initiating and coordinating the immune response against infections, and is proved as a vital target for inflammatory diseases. Herein, TLR4 with sufficient amount and functional activity was generated by heterologous expression and used to investigate the mechanism of apigenin (Api)/chrysin (Chr) inhibition of TLR4 activation. The results demonstrated that Api/Chr exhibited a strong fluorescence quenching effect through a static quenching and a high binding affinity (Ka > 105 L·mol-1) with TLR4, indicating the potential of Api/Chr as a TLR4 inhibitor. Additionally, the binding of Api/Chr induced a loose and unstable conformation of TLR4 with evidence like the decreased hydrophobicity of the tryptophan microenvironment, decreased α-helix content and increased free sulfhydryl content, resulting in reduced stability of the TLR4. The computer simulations revealed that Api/Chr occupied the myeloid differentiation factor 2 (MD-2) binding region, preventing MD-2 from binding to TLR4. Furthermore, the accuracy of the binding site between Api/Chr and TLR4 was confirmed through genetic mutations. Overall, the mechanism by which Api/Chr inhibited TLR4 activation was elucidated at the macroscopic and molecular levels, providing the worthful information concerning the future therapeutic application of Api/Chr as a natural TLR4 inhibitor.

Effects of Processing Conditions and Simulated Digestion In Vitro on the Antioxidant Activity, Inhibition of Xanthine Oxidase and Bioaccessibility of Epicatechin Gallate.

The bioactivity and gastrointestinal stability of epicatechin gallate (ECG) may be affected by processing conditions. Results showed that the antioxidant ability and inhibitory activity on xanthine oxidase (XO) of ECG were higher at low pH values. Appropriate microwave and heating treatments improved the antioxidant (the scavenging rate increased from 71.75% to 92.71% and 80.88% under the microwave and heating treatments) and XO inhibitory activity (the inhibitory rate increased from 47.11% to 56.89% and 51.85% at the microwave and heating treatments) of ECG. The treated ECG led to a more compact structure of XO. Moreover, there may be synergistic antioxidant and inhibitory effects between ECG and its degradation products. The bioaccessibility of ECG after simulated digestion was untreated > microwave > heating, and the microwave-treated ECG still had good XO inhibitory activity after digestion. These findings may provide some significant information for the development of functional foods enriched in catechins.

Protocatechuic acid and gallic acid improve the emulsion and thermal stability of whey protein by covalent binding.

This study aimed to explore the impacts of gallic acid (GA)/protocatechuic acid (PA) on the structural and functional characteristics of whey proteins (WP) through covalent binding. To this purpose, the covalent complexes of WP-PA and WP-GA at different concentration gradients were prepared by the alkaline method. SDS-PAGE indicated that PA/GA was cross-linked by covalent bonds. The decreased contents of free amino and sulfhydryl groups suggested that WP formed covalent bonds with PA/GA by amino and sulfhydryl groups, and the structure of WP became slightly looser after covalent modification by PA/GA. When the concentration of GA was added up to 10 mM, the structure of WP was slightly loosened with a reduction of α-helix content by 2.3% and an increase in random coil content by 3.0%. The emulsion stability index of WP increased by 14.9 min after interaction with GA. Moreover, the binding of WP and 2-10 mM PA/GA increased the denaturation temperature by 1.95 to 19.87 °C, indicating the improved thermal stability of the PA/GA-WP covalent complex. Additionally, the antioxidant capacity of WP was increased with increasing GA/PA concentration. This work may offer worthful information for enhancing the functional properties of WP and the application of the PA/GA-WP covalent complexes in food emulsifiers.

Amelioration of ovalbumin gel properties by EGCG via protein aggregation, hydrogen, and van der Waals force.

The mechanism of epigallocatechin gallate (EGCG)-modified ovalbumin gel (EMOG) was investigated. Results indicated that, with the increase of EGCG concentration from 0% to 0.05%, the opacity, hardness, and cohesiveness of EMOG increased significantly from 0.058 to 0.133, 321.0 g to 377.6 g, and 0.879 to 0.951, respectively, while the soluble protein, surface hydrophobicity, and free sulfhydryl decreased significantly by 41.74%, 28.26%, and 39.36%, respectively. Moreover, EGCG promoted the formation of dense and stable microstructures of EMOG, changed the expansion rate, and improved the stability of EMOG. Moreover, the results of silico simulation showed that EGCG would insert into ovalbumin and interact with the amino acids through van der Waals force and hydrogen bonds, leading to a compact and stable protein structure. In this paper, the mechanism of modification of ovalbumin by EGCG was elucidated at the macro and micro levels, providing insights into the action of polyphenols and proteins.

Improving the gel properties of salted egg white/cooked soybean protein isolate composite gels by ultrasound treatment: Study on the gelling properties and structure.

In this study, the effects of ultrasound treatment on the texture, physicochemical properties and protein structure of composite gels prepared by salted egg white (SEW) and cooked soybean protein isolate (CSPI) at different ratios were investigated. With the increased SEW addition, the ζ-potential absolute values, soluble protein content, surface hydrophobicity and swelling ratio of composite gels showed overall declining trends (P < 0.05), while the free sulfhydryl (SH) contents and hardness of exhibited overall increasing trends (P < 0.05). Microstructural results revealed that composite gels exhibited denser structure with the increased SEW addition. After ultrasound treatment, the particle size of composite protein solutions significantly decreased (P < 0.05), and the free SH contents of ultrasound-treated composite gels were lower than that of untreated composite gels. Moreover, ultrasound treatment enhanced the hardness of composite gels, and promoted the conversion of free water into non-flowable water. However, when ultrasonic power exceeded 150 W, the hardness of composite gels could not be further enhanced. FTIR results indicated that ultrasound treatment facilitated the composite protein aggregates to form a more stable gel structure. The improvement of ultrasound treatment on the properties of composite gels was mainly by promoting the dissociation of protein aggregates, and the dissociated protein particles further interacted to form denser aggregates through disulfide bond, thus facilitating the crosslinking and reaggregation of protein aggregates to form denser gel structure. Overall, ultrasound treatment is an effective approach to improve the properties of SEW-CSPI composite gels, which can improve the potential utilization of SEW and SPI in food processing.

Novel Reactive Distillation Process for Cyclohexyl Acetate Production: Design, Optimization, and Control.

A side-reactor column (SRC) configuration, comprising a vacuum column coupled with atmospheric side reactors, is proposed to overcome the thermodynamic restriction in the esterification of cyclohexene with acetic acid to produce cyclohexyl acetate. Meantime, this configuration can avoid the utilization of the high-pressure steam and provide enough zone for catalyst loading. In order to obtain the minimum total annual cost (TAC), the process is optimized by a mixed-integer nonlinear programming optimization method based on the improved bat algorithm. The results indicate that the optimized SRC configuration saves about 44.81% of the TAC compared to the reactive distillation process. Based on the optimized SRC process, dynamic control is carried out. The dual-point temperature and temperature-composition control structures are proposed to reject throughput and feed composition disturbances. The dynamic performances demonstrate that the temperature-composition control structure is better in maintaining product purity.

Exploring inhibitory effect and mechanism of hesperetin-Cu (II) complex against protein glycation.

Inhibition of advanced glycation end products (AGEs) formed in protein glycosylation is crucial for minimizing diabetic complications. Herein, the anti-glycation potential of hesperetin-Cu (II) complex was investigated. Hesperetin-Cu (II) complex strongly inhibited three stages glycosylation products in bovine serum albumin (BSA)-fructose model, especially for the inhibition of AGEs (88.45%), which was stronger than hesperetin (51.76%) and aminoguanidine (22.89%). Meanwhile, hesperetin-Cu (II) complex decreased the levels of BSA carbonylation and oxidation products. 182.50 µg/mL of hesperetin-Cu (II) complex inhibited 66.71% ß-crosslinking structures of BSA, and scavenged 59.80% superoxide anions and 79.76% hydroxyl radicals. Moreover, after incubating with methylglyoxal for 24 h, hesperetin-Cu (II) complex removed 85.70% methylglyoxal. The mechanisms of protein antiglycation by hesperetin-Cu (II) complex may be through protecting structure, trapping methylglyoxal, scavenging free radicals and interacting with BSA. This study may contribute to the development of hesperetin-Cu (II) complex as a functional food additive against protein glycation.

Research trends in the mental health and multimorbidity of older people from 2002 to 2022: A bibliometric analysis via CiteSpace.

Purpose: Using bibliometric analysis, the objective of this study was to identify research hotspots and trends on multimorbidity and mental health in older adults. This could help guide future research on this topic. Methods: We searched the Web of Science Core Collection for eligible studies. Type of publications were not restricted and the timeframe was set between 2002 and 2022. Knowledge maps were created using CiteSpace to visualize publications, nations, journals, institutions, authors, cited references, and keywords. Microsoft Excel displayed pertinent tables. Results: A total of 216 studies were collected for analysis. The annual publication over the past 20 years demonstrated an upward trend. The main contributors in publications were in North America, Europe, Asia, and Oceania, of which aging was a primary issue. However, collaboration between countries, institutions, and authors were relatively sparce. Cluster analysis and co-citation analysis of references and keywords revealed that the research field could be subdivided into four themes: social psychology as the fundamental disciplinary base, Prevalence of mental disorders and multimorbidity in older adults, pertinent health conditions, and effective interventions. Research trends at present lies in health status, risk factors of prognoses, and effective interventions for prevention and management. Conclusion: The results revealed a reciprocal risk relationship between mental health and multimorbidity. Mental disorders such as depression and anxiety in older adults with multimorbidity garnered considerable interest in the defined population, and further exploration is still promising. Substantial studies on evidence-based prevention and treatment strategies are warranted for improved prognoses.

New Insights into the Inhibition of Hesperetin on Polyphenol Oxidase: Inhibitory Kinetics, Binding Characteristics, Conformational Change and Computational Simulation.

The inhibitory activity of hesperetin on polyphenol oxidase (PPO) and their interaction characteristics were investigated using multiple spectroscopic methods and computational simulation. Hesperetin, a mixed inhibitor, reversibly inhibited PPO activity, and its half-maximum inhibitory concentration (IC50) values on monophenolase and diphenolase were 80.8 ± 1.4 µM and 776.0 ± 15.5 µM, respectively. Multivariate curve resolution-alternate least squares (MCR-ALS) analysis suggested PPO interacted with hesperetin and formed PPO-hesperetin complex. Hesperetin statically quenched PPO's endogenous fluorescence, and hydrophobic interactions mainly drove their binding. Hesperetin affected the polarity of the microenvironment around the Trp residues in PPO, but had no effect on that around Tyr residues. Circular dichroism (CD) results showed that hesperetin increased α-helix content and decreased ß-fold and random coil contents, thus tightening PPO's structure. Molecular docking showed that hesperetin entered the hydrophobic cavity of PPO, bound near the dinuclear copper active center, interacted with Val283, Phe264, His85, Asn260, Val248, and His263 via hydrophobic interactions, formed hydrogen bonds with Met280, His89, and His259 residues and also interacted with Phe292, His61, Phe90, Glu256, His244, Asn260, Phe264, and Gly281 via van der Waals forces. The molecular dynamics simulation results also demonstrated that the addition of hesperetin reduced the stability and hydrophobicity of PPO and increased PPO's structural denseness. Thus, the inhibition of hesperetin on PPO may be because hesperetin bound near the active center of PPO, interacted with the surrounding residues, occupied the binding site for substrate, and induced the changes in PPO's secondary structure, thus inhibiting the catalytic activity of PPO. This study may provide novel views for the inhibition of hesperetin on PPO and theoretical guidance for developing flavonoids as new and efficient PPO inhibitors.