Protein-Mediated Changes in Membrane Fluidity and Ordering: Insights into the Molecular Mechanism and Implications for Cellular Function.

Probing protein-membrane interactions is vital for understanding biological functionality for various applications such as drug development, targeted drug delivery, and creation of functional biomaterials for medical and industrial purposes. In this study, we have investigated interaction of Human Serum Albumin (HSA) with two different lipids, dipalmitoylphosphatidylglycerol (dDPPG) and dipalmitoylphosphatidylcholine (dDPPC), using Vibrational Sum Frequency Generation spectroscopy at different membrane fluidity values. In the liquid-expanded (LE) state of the lipid, HSA (at pH 3.5) deeply intercalated lipid chains through a combination of electrostatic and hydrophobic interactions, which resulted in more ordering of the lipid chains. However, in the liquid-condensed (LC) state, protein intercalation is decreased due to tighter lipid packing. Moreover, our findings revealed distinct differences in HSA's interaction with dDPPG and dDPPC lipids. The interaction with dDPPC remained relatively weak compared to dDPPG. These results shed light on the significance of protein mediated changes in lipid characteristics, which hold considerable implications for understanding membrane protein behavior, lipid-mediated cellular processes, and lipid-based biomaterial design.

A novel FA1-targeting fluorescent probe for specific discrimination and identification of human serum albumin from bovine serum albumin.

A novel probe C1 combining benzothiazole with a spiropyran section was developed for the specific detection of human serum albumin (HSA). The molecular docking suggested that the sulphonic acid group modification allowed C1 to form specific hydrogen bonds with lysine (Lys137) at fatty acid site 1 (FA1) of HSA, thus enabling fluorescence differentiation between HSA and BSA.

Developing a Copper(II) Isopropyl 2-Pyridyl Ketone Thiosemicarbazone Compound Based on the IB Subdomain of Human Serum Albumin-Indomethacin Complex: Inhibiting Tumor Growth by Remodeling the Tumor Microenvironment.

To develop a next-generation metal agent and dual-agent multitargeted combination therapy, we developed a copper (Cu) compound based on the properties of the human serum albumin (HSA)-indomethacin (IND) complex to remodel the tumor microenvironment (TME). We optimized a series of Cu(II) isopropyl 2-pyridyl ketone thiosemicarbazone compounds to obtain a Cu(II) compound (C4) with significant cytotoxicity and then constructed an HSA-IND-C4 complex (HSA-IND-C4) delivery system. IND and C4 bind to the hydrophobic cavities of the IB and IIA domains of HSA, respectively. In vivo, the HSA-IND-C4 not only showed enhanced antitumor efficacy relative to C4 and C4 + IND but also improved their targeting ability and decreased their side effects. The antitumor mechanism of C4 + IND involved acting on the different components of the TME. IND inhibited tumor-related inflammation, while C4 not only induced apoptosis and autophagy of cancer cells but also inhibited tumor angiogenesis.

Association between admission albumin levels and 30-day readmission after hip fracture surgery in geriatric patients: a propensity score-matched study.

PURPOSE: This study aimed to evaluate the correlation admission albumin levels and 30-day readmission after hip fracture surgery in geriatric patients. METHODS: In this retrospective cohort study, 1270 geriatric patients admitted for hip fractures to a level I trauma center were included. Patients were stratified by clinical thresholds and albumin level quartiles. The association between admission albumin levels and 30-day readmission risk was assessed using multivariate logistic regression and propensity score-matched analyses. The predictive accuracy of albumin levels for readmission was evaluated by ROC curves. The dose-response relationship between albumin levels and readmission risk was examined. RESULTS: The incidence of 30-day readmission was significantly higher among hypoalbuminemia patients than those with normal albumin levels (OR = 2.090, 95%CI:1.296-3.370, p = 0.003). Furthermore, propensity score-matched analyses demonstrated that patients in the Q2(35.0-37.9 g/L) (OR 0.621, 95%CI 0.370-1.041, p = 0.070), Q3(38.0-40.9 g/L) (OR 0.378, 95%CI 0.199-0.717, p < 0.001) and Q4 (≥ 41 g/L) (OR 0.465, 95%CI 0.211-0.859, p = 0.047) quartiles had a significantly lower risk of 30-day readmission compared to those in the Q1(< 35 g/L) quartile. These associations remained significant after propensity score matching (PSM) and subgroup analyses. Dose-response relationships between albumin levels and 30-day readmission were observed. CONCLUSIONS: Lower admission albumin levels were independently associated with higher 30-day readmission rates in elderly hip fracture patients. Our findings indicate that serum albumin may assist perioperative risk assessment, and prompt correction of hypoalbuminemia and malnutrition could reduce short-term readmissions after hip fracture surgery in this high-risk population.

Generation of affinity maps for thiazolidinediones with human serum albumin using affinity microcolumns. I. Studies of effects by glycation on multisite drug binding.

Human serum albumin (HSA) is known to undergo modifications by glucose during diabetes. This process produces glycated HSA that can have altered binding to some drugs. In this study, high-performance affinity microcolumns and competition studies were used to see how glycation affects the binding by two thiazolidinedione-class drugs (i.e., pioglitazone and rosiglitazone) at specific regions of HSA. These regions included Sudlow sites I and II, the tamoxifen and digitoxin sites, and a drug-binding site located in subdomain IB. At Sudlow site II, the association equilibrium constants (or binding constants) for pioglitazone and rosiglitazone with normal HSA were 1.7 × 105 M-1 and 2.0 × 105 M-1 at pH 7.4 and 37 °C, with values that changed by up to 5.7-fold for glycated HSA. Sudlow site I of normal HSA had binding constants for pioglitazone and rosiglitazone of 3.4 × 105 M-1 and 4.6 × 105 M-1, with these values changing by up to 1.5-fold for glycated HSA. Rosiglitazone was found to also bind a second region that had a positive allosteric effect on Sudlow site I for all the tested preparations of HSA (binding affinity, 1.1-3.2 × 105 M-1; coupling constant for Sudlow site I, 1.20-1.34). Both drugs had a strong positive allosteric effect on the tamoxifen site of HSA (coupling constants, 13.7-19.9 for pioglitazone and 3.7-11.5 for rosiglitazone). Rosiglitazone also had weak interactions at a site in subdomain IB, with a binding constant of 1.4 × 103 M-1 for normal HSA and a value that was altered by up to 6.8-fold with glycated HSA. Neither of the tested drugs had any significant binding at the digitoxin site. The results were used to produce affinity maps that described binding by these thiazolidinediones with HSA and the effects of glycation on these interactions during diabetes.

Insights into the effect of glucose on the binding between human serum albumin and the nonsteroidal anti-inflammatory drug nimesulide.

Glucose interacts with human serum albumin (HSA, the main protein responsible for the biodistribution of drugs in the bloodstream) and consequently affects the binding capacity of exogenous compounds. Thus, in this work, the interactive profile between HSA and the anti-inflammatory drug nimesulide (NMD, used mainly by patients with diabetic neuropathy to relieve acute or chronic pains) was characterized in nonglycemic, normoglycemic (80 mg/dL), and hyperglycemic (320 mg/dL) conditions by biophysics techniques. There is a spontaneous and ground-state association HSA:NMD under physiological conditions. Therefore, the Stern-Volmer constant (Ksv) can also be used to estimate the binding affinity. The Ksv values for nonglycemic, normoglycemic, and hyperglycemic conditions are around 104 M-1, indicating a moderate affinity of NMD to albumin that was slightly improved by glucose levels. Additionally, the binding is enthalpically and entropically driven mainly into subdomains IIA or IIIA. The binding perturbs weakly the α-helix content of albumin, however, glucose potentially stabilizes the tertiary structure, decreasing the structural perturbation upon NMD binding and improves the complex HSA:NMD stability. Overall, the biophysical characterization indicated that glucose levels might slightly positively impact the pharmacokinetic profile of NMD, allowing NMD to achieve its therapeutical potential without affecting drastically its effective dosages.

Insight into the Effect of Submicellar Concentrations of Sodium Deoxycholate on the Structure, Stability, and Activity of Bovine and Human Serum Albumin: An Interesting Comparison between Single and Double Tryptophan Proteins.

The progressive escalation in the applications of bile salts in diverse fields has triggered research on their interaction with various biological macromolecules, especially with proteins. A proper understanding of the interaction process of bile salts, particularly in the lower concentrations range, with the serum albumin seems important since the normal serum concentration of bile salts is approximately in the micromolar range. The current study deals with a comprehensive and comparative analysis of the interaction of submicellar concentrations of sodium deoxycholate (NaDC) with two homologous transport proteins: bovine serum albumin (BSA) and human serum albumin (HSA). HSA and BSA with one and two tryptophans, respectively, provide the opportunity for an interesting comparison of tryptophan fluorescence behavior on interaction with NaDC. The study suggests a sequential interaction of NaDC in three discrete stages with the two proteins. A detailed study using warfarin and ibuprofen as site markers provides information about the sites of interaction, which is further confirmed by inclusive molecular dynamics simulation analysis. Moreover, the comparison of the thermodynamics and stability of the NaDC-serum albumin complexes confirms the stronger interaction of NaDC with BSA as compared to that with HSA. The differential interaction between the bile salt and the two serum albumins is further established from the difference in the extent of decrease in the esterase-like activity assay of the proteins in the presence of NaDC. Therefore, the present study provides important insight into the effect of submicellar concentrations of NaDC on the structure, stability, and activity of the two homologous serum albumins and thus can contribute not only to the general understanding of the complex nature of serum albumin-bile salt interactions but also to the design of more effective pharmaceutical formulations in the field of drug delivery and biomedical research.

Studies on the binding of wedelolactone to human serum albumin with multi-spectroscopic analysis, molecular docking and molecular dynamic simulation.

Wedelolactone (WEL) is a small molecule compound isolated from Eclipta prostrate L., which has been reported to possess various biological activities such as anti-hepatotoxicity, anti-hypertension, anti-tumour, anti-phospholipase A2 and detoxification activity against snake venom. In the present study, we investigated the interaction of WEL with human serum albumin (HSA) using simultaneous fluorescence, UV-visible spectroscopy, 3D fluorescence spectroscopy, Fourier transform infrared spectroscopy (FTIR), molecular docking technique and molecular dynamics simulation. We found that the interaction between HSA and WEL can exhibit a static fluorescence burst mechanism, and the binding process is essentially spontaneous, with the main forces manifested as hydrogen bonding, van der Waals force and electrostatic interactions. Competitive binding and molecular docking studies showed that WEL preferentially bound to HSA in substructural region IIA (site I); molecular dynamics simulations showed that HSA interacted with WEL to form a stable complex, which also induced conformational changes in HSA. The study of the interaction between WEL and HSA can provide a reference for a more in-depth study of the pharmacodynamic mechanism of WEL and its further development and utilisation.

A Facile Access to Green Fluorescent Albumin Derivatives.

A Morita-Baylis-Hillman Adduct (MBHA) derivative bearing a triphenylamine moiety was found to react with human serum albumin (HSA) shifting its emission from the blue to the green-yellow thus leading to green fluorescent albumin (GFA) derivatives and enlarging the platform of probes for aggregation-induced fluorescent-based detection techniques. A possible interaction of MBHA derivative 7 with a lipophilic pocket within the HSA structure was suggested by docking studies. DLS experiments showed that the reaction with HSA induce a conformational change of the protein contributing to the aggregation process of GFA derivatives. The results of investigations on the biological properties suggested that GFA retained the ability of binding drug molecules such as warfarin and diazepam. Finally, cytotoxicity evaluation studies suggested that, although the MBHA derivative 7 at 0.1 µg/mL affected the percentage of cell viability in comparison to the negative control, it cannot be considered cytotoxic, whereas at all the other concentrations≥0.5 µg/mL resulted cytotoxic at different extent.

Research on the Interaction Mechanism and Structural Changes in Human Serum Albumin with Hispidin Using Spectroscopy and Molecular Docking.

The interaction between human serum albumin (HSA) and hispidin, a polyketide abundantly present in both edible and therapeutic mushrooms, was explored through multispectral methods, hydrophobic probe assays, location competition trials, and molecular docking simulations. The results of fluorescence quenching analysis showed that hispidin quenched the fluorescence of HSA by binding to it via a static mechanism. The binding of hispidin and HSA was validated further by synchronous fluorescence, three-dimensional fluorescence, and UV/vis spectroscopy analysis. The apparent binding constant (Ka) at different temperatures, the binding site number (n), the quenching constants (Ksv), the dimolecular quenching rate constants (Kq), and the thermodynamic parameters (∆G, ∆H, and ∆S) were calculated. Among these parameters, ∆H and ∆S were determined to be 98.75 kJ/mol and 426.29 J/(mol·K), respectively, both exhibiting positive values. This observation suggested a predominant contribution of hydrophobic forces in the interaction between hispidin and HSA. By employing detergents (SDS and urea) and hydrophobic probes (ANS), it became feasible to quantify alterations in Ka and surface hydrophobicity, respectively. These measurements confirmed the pivotal role of hydrophobic forces in steering the interaction between hispidin and HSA. Site competition experiments showed that there was an interaction between hispidin and HSA molecules at site I, which situates the IIA domains of HSA, which was further confirmed by the molecular docking simulation.

Biophysical and structural characterization of tetramethrin serum protein complex and its toxicological implications.

Tetramethrin (TMT) is a commonly used insecticide and has a carcinogenic and neurodegenerative effect on humans. The binding mechanism and toxicological implications of TMT to human serum albumin (HSA) were examined in this study employing a combination of biophysical and computational methods indicating moderate binding affinity and potential hepato and renal toxicity. Fluorescence quenching experiments showed that TMT binds to HSA with a moderate affinity, and the binding process was spontaneous and predominantly enthalpy-driven. Circular dichroism spectroscopy revealed that TMT binding did not induce any significant conformational changes in HSA, resulting in no changes in its alpha-helix content. The binding site and modalities of TMT interactions with HSA as computed by molecular docking and molecular dynamics simulations revealed that it binds to Sudlow site II of HSA via hydrophobic interactions through its dimethylcyclopropane carboxylate methyl propanyl group. The structural dynamics of TMT induce proper fit into the binding site creating increased and stabilizing interactions. Additionally, molecular mechanics-Poisson Boltzmann surface area calculations also indicated that non-polar and van der Waals were found to be the major contributors to the high binding free energy of the complex. Quantum mechanics (QM) revealed the conformational energies of the binding confirmation and the degree of deviation from the global minimum energy conformation of TMT. The results of this study provide a comprehensive understanding of the binding mechanism of TMT with HSA, which is important for evaluating the toxicity of this insecticide in humans.

Oxidative stress induced conformational changes of human serum albumin.

Oxidative stress, generated by reactive oxygen species (ROS), is responsible for the loss of structure and functionality of proteins and is associated with several aging-related diseases. Here, we report an in vitro study to gauge the effect of ROS on the structural rearrangement of human serum albumin (HSA), a plasma protein, through metal-catalyzed oxidation (MCO) at physiological temperature through various biophysical techniques like UV-vis absorption, circular dichroism (CD), differential scanning calorimetry (DSC), MALDI-TOF, FTIR, and Raman spectroscopy. The UV-vis spectra of oxidized HSA show an early blueshift, signifying the unfolding of the protein because of ROS followed by the broadening of the absorption peak at a longer time. The DSC data corroborate the observation, revealing an exothermic transition for the oxidized sample at a longer time, suggesting in situ aggregation. The CD and FTIR spectra indicate the associated secondary structural changes occurring with time, depicting the variation of the helical content of HSA. The amide-III analysis of Raman data also complements the structural changes, and MALDI-TOF data show the mass distribution with time. Overall, this work might help determine the effect of oxidation on the biological activity of serum albumin as it can impact the physiological properties of HSA.

Chiral Au(III) chelates exhibit unique NCI-60 cytotoxicity profiles and interactions with human serum albumin.

Au(III) bis(pyrrolide-imine) chelates are emerging as a class of versatile, efficacious metallodrug candidates. Here, we synthesised two enantiopure chiral ligands H2L1 and H2L2 (tetradentate cyclohexane-1,2-diamine-bridged bis(pyrrole-imine) derivatives). Metallation of the ligands with Au(III) afforded the chiral cationic complexes AuL1 and AuL2. The in vitro cytotoxicities of AuL1 and AuL2 determined in the NCI-60 single-dose drug screen were 56.5% and 89.1%, respectively. AuL1 was subsequently selected for a five-dose NCI-60 screen, attaining GI50, IC50, and LC50 values of 4.7, 9.3 and 39.8 µM, respectively. Hierarchical cluster analysis of the NCI-60 data indicated that the profile for AuL1 was similar to that of vinblastine sulfate, a microtubule-targeting vinca alkaloid. Reactions of AuL1 with glutathione (GSH) in vitro confirmed its susceptibility to reduction, Au(III) → Au(I), by intracellular thiols. Because human serum albumin (HSA) is responsible for transporting clinically deployed and investigational drugs, we studied the uptake of AuL1 and AuL2 by HSA to delineate how chirality impacts their protein-binding affinity. Steady-state fluorescence quenching data acquired on the native protein and data from site-specific probes showed that the compounds bind at sites close enough to Trp-214 (subdomain IIA) of HSA to quench the fluorophore. The bimolecular quenching rate constants, Kq, were ca. 102 times higher than the maximum diffusion-controlled collision constant of a biomolecule in water (1010 M-1 s-1), confirming that static fluorescence quenching was the dominant mechanism. The Stern-Volmer constants, KSV, were ∼104 M-1 at 37 °C, while the affinity constants, Ka (37 °C), measured ∼2.1 × 104 M-1 (AuL1) and ∼1.2 × 104 M-1 (AuL2) for enthalpy-driven ligand uptake targeting Sudlow's site I. Although far- and near-UV CD spectroscopy indicated that both complexes minimally perturb the secondary and tertiary structure of HSA, substantial shifts in the CD spectra were recorded for both protein-bound ligands. This study highlights the role of chirality in determining the cytotoxicity profiles and protein binding behaviour of enantiomeric Au(III) chelates.

The binding selectivity of quercetin and its structure-related polyphenols to human serum albumin using a fluorescent dye cocktail for multiplex drug-site mapping.

Human serum albumin (HSA) is a serum protein that carries flavonoids in blood circulation. In this report, the binding selectivity and strength of interactions to HSA-binding sites (sites I or II) by flavonoids were evaluated using competition experiments and the specific fluorescent dyes, dansylamide and BD140. Most tested flavonoids bound site I preferentially, with the binding strength dependent on the mother structure in the order flavonol > flavone > flavanone > flavan 3-ols. Glycosylation or glucuronidation reduced the binding of quercetin to site I of HSA, whereas sulfation increased binding. Quercetin 7-sulfate showed the strongest binding and molecular docking simulations supported this observation. Prenylation at any position or glucuronidation and sulfation at the C-4' or C-7 position of quercetin facilitated stronger binding to site II. The binding affinity of flavonoids toward site I correlated with the partition coefficient value (logP), whereas no corresponding correlation was observed for site II.

Modulation of protein-ligand interactions in the presence of ZIF-8: Spectroscopy and molecular dynamics simulation.

In this paper, we investigated the protein-ligand interactions in the presence of ZIF-8 using multi-spectroscopic approaches and molecular dynamics simulation. Fluorescence experiments and molecular docking results showed that ZIF-8 did not change the type of quenching and interaction force between ciprofloxacin (CIP) and human serum albumin (HSA), but made the binding constant of HSA-CIP to be smaller, suggesting that ZIF-8 maybe accelerate the dissociation of CIP from HSA-CIP complex. Moreover, the effect of ZIF-8 on the physiological function of HSA was explored. Multi-spectroscopic methods revealed that ZIF-8 did not significantly alter the microenvironment of amino acid groups, but cause a slight decrease in the content of α-helical conformation, and a sparse and flexible structure of the protein backbone. These peculiarities might lead to the diminution of HSA's ability to control drugs. In short, ZIF-8 might enhance drug effect due to affecting the binding of drugs to proteins. However, the present study is only a preliminary investigation of the suitability of ZIF-8 as a drug carrier in vitro, and subsequent in vivo experimental studies will be required to further confirm the idea.

Investigation of steric hindrance effect on the interactions between four alkaloids and HSA by isothermal titration calorimetry and molecular docking.

The binding of four alkaloids with human serum albumin (HSA) was investigated by isothermal titration calorimetry (ITC), spectroscopy and molecular docking techniques. The findings demonstrated that theophylline or caffeine can bind to HAS, respectively. The number of binding sites and binding constants are obtained. The binding mode is a static quenching process. The effects of steric hindrance, temperature, salt concentration and buffer solution on the binding indicated that theophylline and HSA have higher binding affinity than caffeine. The fluorescence and ITC results showed that the interaction between HSA and theophylline or caffeine is an entropy-driven spontaneous exothermic process. The hydrophobic force was the primary driving factor. The experimental results were consistent with the molecular docking data. Based on the molecular structures of the four alkaloids, steric hindrance might be a major factor in the binding between HSA and these four alkaloids. This study elucidates the mechanism of interactions between four alkaloids and HSA.

Human serum albumin as a potential drug delivery system for N-methylated hot spot insulin analogs inhibiting hormone aggregation.

The purpose of this study was to investigate whether Human Serum Albumin (HSA) can bind N-methylated analogs of hot spots of native insulin. Three N-methylated derivatives of the A13-A19 fragment of native insulin were used: L(N-Me)YQLENY (1), LYQ(N-Me)LENY (2), and L(N-Me)YQ(N-Me)LENY (3). The studied N-methylated insulin fragments possess inhibiting potential against hormone aggregation. A variety of research techniques, including spectroscopic methods and microscopy assays, were used to study the interaction of HSA with the N-methylated insulin fragments. Based on spectroscopic measurements with Congo Red and Thioflavin T, all the analyzed N-methylated peptides were able to interact with the HSA surface. The CD spectrum registered for HSA in the presence of L(N-Me)YQLENY showed the smallest content of α-helix conformation, indicating the most compact HSA structure. Based on the results of MST, the dissociation constants (Kd) for complexes of HSA and peptides 1-3 were 19.2 nM (complex 1), 15.6 nM (complex 2), and 8.07 nM (complex 3). Microscopy assays, dynamic light scattering measurements as well as computer simulation of protein-ligand interaction also confirmed the possibility of docking the N-methylated inhibitors within HSA.

Covalent immobilization of human serum albumin on cellulose acetate membrane for scavenging amyloid beta - A stepping extracorporeal strategy for ameliorating Alzheimer's disease.

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by interrupted neurocognitive functions and impaired mental development presumably caused by the accumulation of amyloid beta (Aß) in the form of plaques. Targeting Aß has been considered a promising approach for treating AD. In the current study, human serum albumin (HSA), a natural Aß binder, is covalently immobilized onto the surface of a cellulose acetate (CA) membrane to devise an extracorporeal Aß sequester. The immobilization of HSA at 3.06 ± 0.22 µg/mm2 of the CA membrane was found to be active functionally, as evidenced by the esterase-like activity converting p-nitrophenyl acetate into p-nitrophenol. The green fluorescent protein-Aß (GFP-Aß) fusion protein, recombinantly produced as a model ligand, exhibited characteristics of native Aß. These features include the propensity to form aggregates or fibrils and an affinity for HSA with a dissociation constant (KD) of 0.91 µM. The HSA on the CA membrane showed concentration-dependent sequestration of GFP-Aß in the 1-10-µM range. Moreover, it had a greater binding capacity than HSA immobilized on a commercial amine-binding plate. Results suggest that the covalent immobilization of HSA on the CA surface can be used as a potential platform for sequestering Aß to alleviate AD.

Development of Human Serum Albumin Fluorescent Probes in Detection, Imaging, and Disease Therapy.

Human serum albumin (HSA) acts as a repository and transporter of substances in the blood. An abnormal concentration may indicate the occurrence of liver- and kidney-related diseases, which has attracted people to investigate the precise quantification of HSA in body fluids. Fluorescent probes can combine with HSA covalently or noncovalently to quantify HSA in urine and plasma. Moreover, probes combined with HSA can improve its photophysical properties; probe-HSA has been applied in real-time monitoring and photothermal and photodynamic therapy in vivo. This Review will introduce fluorescent probes for quantitative HSA according to the three reaction mechanisms of spatial structure, enzymatic reaction, and self-assembly and systematically introduce the application of probes combined with HSA in disease imaging and phototherapy. It will help develop multifunctional applications for HSA probes and provide assistance in the early diagnosis and treatment of diseases.

An HDBB-based fluorescent probe for the sensitive detection of human serum albumin.

The detection of human serum albumin (HSA) in bodily fluids is of great significance in the biomedical area because HSA in bodily fluids is commonly used as a biomarker for the early diagnosis of diseases. To detect HSA, we employed HDBB, 4,4'-(hydrazine-1,2-diylidene bis(methanylylidene)) bis(3-hydroxybenzoic acid), as a fluorescent probe with a large Stokes shift. HDBB had obvious excited state intramolecular proton transfer (ESIPT) and aggregation-induced emission (AIE) features. We elucidated the ESIPT characteristics of HDBB through the DFT approach. We also performed a molecular docking simulation between HDBB and HSA, showing that HDBB primarily bonded to HSA via hydrophobic force and hydrogen bonds. The FL intensities of HDBB with HSA concentrations had a linear range of 0.01-0.2 mg mL-1 (R2 = 0.9995), and the LOD was 1.104 µg mL-1. We also used the probe to detect HSA in urine, with spiked recoveries of 98.10-105.33%. Given its high selectivity and feasible synthesis, HDBB has potential applications in detecting HSA in real biological systems.