3D Fluorescence Research Articles

Network response of brain microvasculature to neuronal stimulation

Neurovascular coupling (NVC), or the adjustment of blood flow in response to local increases in neuronal activity is a hallmark of healthy brain function, and the physiological foundation for functional magnetic resonance imaging (fMRI). However, it remains only partly understood due to the high complexity of the structure and function of the cerebrovascular network. Here we set out to understand NVC at the network level, i.e. map cerebrovascular network reactivity to activation of neighbouring neurons within a 500×500×500 μm3 cortical volume (∼30 high-resolution 3-nL fMRI voxels). Using 3D two-photon fluorescence microscopy data, we quantified blood volume and flow changes in the brain vessels in response to spatially targeted optogenetic activation of cortical pyramidal neurons. We registered the vessels in a series of image stacks acquired before and after stimulations and applied a deep learning pipeline to segment the microvascular network from each time frame acquired. We then performed image analysis to extract the microvascular graphs, and graph analysis to identify the branch order of each vessel in the network, enabling the stratification of vessels by their branch order, designating branches 1–3 as precapillary arterioles and branches 4+ as capillaries. Forty-five percent of all vessels showed significant calibre changes; with 85 % of responses being dilations. The largest absolute CBV change was in the capillaries; the smallest, in the venules. Capillary CBV change was also the largest fraction of the total CBV change, but normalized to the baseline volume, arterioles and precapillary arterioles showed the biggest relative CBV change. From linescans along arteriole-venule microvascular paths, we measured red blood cell velocities and hematocrit, allowing for estimation of pressure and local resistance along these paths. While diameter changes following neuronal activation gradually declined along the paths; the pressure drops from arterioles to venules increased despite decreasing resistance: blood flow thus increased more than local resistance decreases would predict. By leveraging functional volumetric imaging and high throughput deep learning-based analysis, our study revealed distinct hemodynamic responses across the vessel types comprising the microvascular network. Our findings underscore the need for large, dense sampling of brain vessels for characterization of neurovascular coupling at the network level in health and disease.

An update on liver surgery- a new terminology and modern techniques.

Liver surgery is the cornerstone of the curative treatment of malignant liver tumors. However, the liver anatomy is very complex, and liver surgery is still associated with relevant morbidity despite many technical advances. The Brisbane nomenclature is used worldwide to classify liver resection. However, this nomenclature has several limitations as multiple terms are used for the same type of resection. Non-anatomical resections, multiple resections, and combined bilio-vascular resections were not mentioned. Therefore, new terminologies have been proposed for the precise and simple classification of liver resection. Furthermore, in recent years, many technical innovations have been introduced in liver surgery, such as 3D imaging systems and indocyanine green fluorescence, for better preoperative and intraoperative identification of tumor localization and critical vascular structures. Minimally invasive techniques are used more frequently in liver surgery. Potential benefits include less intraoperative blood loss, less pain, and a shorter hospital stay. The implementation of robotic systems also has an impact on liver surgery, and the number of cases reported in the literature is constantly increasing. The potential benefits of robotic liver resection over laparoscopic liver resection will be the subject of future studies.

Interaction of food colorant indigo carmine with human and bovine serum albumins: A multispectroscopic, calorimetric, and theoretical investigation

In this work we have studied the interaction of the food dye Indigo-Carmine (IndC) with the most studied model transport proteins i.e. human and bovine serum albumin (HSA & BSA). A multispectroscopic approach was used to analyze the details of the binding process. The intrinsic fluorescence of both the albumins was significantly quenched by IndC and the quenching was both static and dynamic in nature with the former being dominant. The HSA-lndC and BSA-IndC distance after complexation was determined by Förster resonance energy transfer (FRET) method which suggested efficient energy transfer from the albumins to IndC. Thermodynamics of serum protein-IndC complexation was estimated by isothermal titration calorimetry (ITC) which revealed that the binding was enthalpy driven. Circular dichroism (CD) and FTIR spectroscopy revealed that the binding of IndC induced secondary structural changes in both the serum proteins. Synchronous and 3D fluorescence spectroscopy revealed that the binding interaction caused microenvironmental changes of protein fluorophores. Molecular docking analysis suggested that hydrogen bonding and hydrophobic interactions are the major forces involved in the complexation process.

Investigation on the inhibition mechanism and binding behavior of cryptolepine to α-glucosidase and its hypoglycemic activity by multi-spectroscopic method

Cryptolepine, one important active ingredient of Cryptolepis sanguinolenta (Lour), exhibits various beneficial pharmacological activities. But its anti-α-glucosidase and hypoglycemic activities are still not cleared. Thence, in this study, we investigated its inhibitory effects on α-glucosidase by a multi-spectroscopic method and in vivo hypoglycemic activity by the oral glucose tolerance test (OGTT). The results showed that cryptolepine is a reversible mixed-type α-glucosidase inhibitor with IC50 value of 0.44 ± 0.05 mM, ∼ 2-fold stronger than positive control acarbose (IC50: 0.72 ± 0.02 mM). Fluorescence quenching confirmed cryptolepine could quench the endogenous fluorescence of α-glucosidase in a static process. Fluorescence quenching, synchronous fluorescence, CD spectra, and 3D fluorescence results showed that the binding of cryptolepine with α-glucosidase caused chromophore microenvironment and conformation changes of α-glucosidase, consequently inhibiting its catalytic activity. Molecular docking revealed the detailed binding interactions between cryptolepine and α-glucosidase. Furthermore, oral administration of cryptolepine could improve the glucose tolerance in mice to reduce the postprandial hyperglycemia. These findings provide a solid foundation for understanding the inhibitory effects on α-glucosidase and hypoglycemic activity of cryptolepine.

Использование 3D-спектров флуоресценции для идентификации подлинности масла виноградной косточки

Falsification of grape seed oil causes serious damage to consumers and shows unfair competition in the market. 3D fluorescence spectroscopy was used to characterize oil flurophores and differentiate samples by identification criteria: method of preparation, raw material used, and manufacturer. During the study, 3D excitation-emission spectra of several samples of grape seed oil from various manufacturers available on the Russian market were obtained. One of the samples with confirmed production histories was used for comparison. The results of analysis of the obtained spectra by zones of maxima on the hypersurface of the absorption-emission matrix were correlated with individual fluorophores and their groups of functional ingredients of the oil, providing its physiological activity. The possibility of using the obtained data for identification of grapeseed oil samples and prompt detection of falsifications is shown. All samples were tested in hexane. It has been shown that genuine oil samples have distinctly differentiated regions of very high intensity on the hypersurface of the excitation-radiation matrix intensity with peaks characteristic of all isomers of tocopherols and tocotrienols with a predominance of the gamma isomer with an EEM maximum of 292/324 nm. There are also intense regions of absorption of carotenoids 325/453 nm and isomeric forms of chlorophyll 405–436/650–670 nm, the  form being the prevailing one. The absorption regions of procyanidins are intense 280 /316–324 nm; gallic acid 260/390 and coumaric acid 270/420. The uniqueness of the 3D fluorescence spectra for each oil and the formation of the resulting data pool in the form of a three-dimensional matrix allows them to be used for machine learning and the isolation of significant parameters by standard chemometrics.

Third-order calibration applied to process surfactant-modulated excitation-emission matrix four-way fluorescence data for the direct determination of four polycyclic aromatic hydrocarbons in oilfield produced water

Fluorescence spectroscopy is a powerful tool to determine polycyclic aromatic hydrocarbons (PAHs) owing to the strong endogenous fluorescence of these compounds. However, the presence of unknown interferences and overlapped spectra hinders the accurate determination of PAHs in oilfield produced water. Moreover, surfactants frequently coexist in oilfield produced water and will seriously affect the fluorescence signals of PAHs. Herein, a new methodology applying third-order calibration to process four-way (4D) fluorescence data was proposed to solve these problems and achieve accurate determination of pyrene, fluorene, phenanthrene, and fluoranthene as an example in oilfield produced water. The methodology is based on excitation-emission matrix fluorescence modulated by different concentrations of sodium dodecyl benzene sulfonate (SDBS) in the analyzed samples. The 4D fluorescence data were processed by third-order calibration methods including four-way parallel factor analysis (4-PARAFAC) and alternating weighted residue constraint quadrilinear decomposition (AWRCQLD), and the results were compared with those of second-order calibration methods. It was proved that third-order calibration was capable of accurately identifying and quantifying PAHs together with SDBS in oilfield produced water, which has better quantitative results and figures of merit compared to second-order calibration. This study provided a new approach to generating 4D fluorescence data and opened up an avenue for the accurate determination of PAHs in complex oilfield produced water with surfactants.

Detection and quantification of olive oil adulteration using fluorescence spectroscopy and chemometric tools

The increasing incidence of fraud in the olive oil market, particularly through adulteration with cheaper oils, poses a major challenge for the industry. This study examines the effectiveness of fluorescence spectroscopy combined with chemometric tools as a method to detect and quantify olive oil adulteration. Chemical analyses were used to measure parameters such as specific absorption coefficients K232 and K270, peroxide value, and acidity, in accordance with International Olive Council (IOC) standards. These measurements were conducted on samples of olive oil blended with varying percentages of adulterating oils. 3D fluorescence spectra were analyzed to examine the changes induced by adulteration. The results show that free acidity and peroxide value decrease with increasing adulteration degree, while the parameters K232 and K270 increase with the degree of adulteration. Principal component analysis (PCA) was effectively used to differentiate samples based on the percentage of adulterant. A partial least squares (PLS) regression model was developed, achieving a correlation coefficient of 0.999 for predicting the percentage of adulterant in olive oil. This PLS model also proved effective in predicting oil quality parameters such as free acidity and K232 and K270 indices. The results of this study demonstrate the significant potential of fluorescence spectroscopy as a rapid and non-destructive method for olive oil authentication. PCA emerges as a powerful tool for characterizing pure oils without requiring sample preparation or destruction. Furthermore, PLS models provide accurate means to predict both the presence of adulterants and olive oil quality parameters. This approach offers a promising solution to combat fraud in the olive oil industry, ensuring product integrity and quality.

Calibrating Fluorescence Microscopy With 3D-Speckler (3D Fluorescence Speckle Analyzer).

Fluorescence microscopy has been widely accessible and indispensable in cell biology research. This technique enables researchers to label targets, ranging from individual entities to multiple groups, with fluorescent markers. It offers precise determinations of localization, size, and shape, along with accurate quantifications of fluorescence signal intensities. Furthermore, an ideal fluorescence microscope can achieve approximately 250 nm in lateral and 600 nm in axial resolution. Despite its integral role in these measurements, the calibration of fluorescence microscopes is often overlooked. This protocol introduces the use of 3D-Speckler (3D fluorescence speckle analyzer), a semi-automated software tool we have recently developed, for calibrating fluorescence microscopy. Calibration of fluorescence microscopy includes determining resolution limits, validating accuracy in size measurements, evaluating illumination flatness, and determining chromatic aberrations. 3D-Speckler is user-friendly and enables precise quantification of fluorescence puncta, including nanoscale 2D/3D particle size, precise locations, and intensity information. By utilizing multispectral fluorescence beads of known sizes alongside 3D-Speckler, the software can effectively calibrate imaging systems. We emphasize the importance of routine calibration for imaging systems to maintain their integrity and reproducibility, ensuring accurate quantification. This protocol provides a detailed step-by-step guide on using 3D-Speckler to calibrate imaging systems. Key features • Semi-automated particle detection. • Accurate three-dimensional measurement of fluorescent particle sizes. • High-precision three-dimensional localization of fluorescent particles. • Precision analysis of point spread function and chromatic aberration in fluorescence microscopy.

A sensitive lateral flow test strip sensor for visual detection of acid red 18 in food using bicentric-emission carbon dots.

Hazardous synthetic colorants have found widespread use in food production, and excessive consumption of these pigments can pose potential risks to human health. In this study, we propose an ultrasensitive fluorescence method for the analysis of Acid Red 18 (AR18) in food products. The method is based on the nitrogen-doped carbon dots (N-CDs) derived from tris and resorcinol through a hydrothermal way. The as-synthesized N-CDs exhibit two emission centers at 425 nm and 541 nm, corresponding to the excitation wavelengths of 377 nm and 465 nm, respectively. Upon the addition of AR18, the fluorescence intensity at 541 nm significantly decreases with a simultaneous, though less pronounced, reduction in the intensity at 425 nm, which is attributed to the localization of fluorescence resonance energy transfer (L-FRET). Specifically, a novel ratiometric fluorescent probe was constructed based on the extracted data from the 3D fluorescence excitation-emission matrix. This probe demonstrates a wide linear range from 0.0539 to 30 μM and a low limit of detection (LOD) of 53.9 nM. For practical applications, a portable fluorescent sensor based on a lateral flow test strip (LFTS) was designed for real-time monitoring of AR18. Color channel values were determined using a smartphone application, resulting in a satisfactory LOD of 75.3 nM. Furthermore, the suitability of the proposed ratiometric fluorescent probe was validated through the detection of AR18 in real food samples, consistently achieving recovery rates in the range of 99.7-101.4%. This research not only expands the scope of CDs in sensing fields, but also provides an effective strategy for the development of an excellent platform for real-time AR18 detection, contributing to public food safety.

Noninvasive Visualization of Amyloid-Beta Deposits in Alzheimer's Amyloidosis Mice via Fluorescence Molecular Tomography Using Contrast Agent.

Alzheimer's disease is pathologically featured by the accumulation of amyloid-beta (Aβ) plaque and neurofibrillary tangles. Compared to small animal positron emission tomography, optical imaging features nonionizing radiation, low cost, and logistic convenience. Optical detection of Aβ deposits is typically implemented by 2D macroscopic imaging and various microscopic techniques assisted with Aβ-targeted contrast agents. Here, we introduce fluorescence molecular tomography (FMT), a macroscopic 3D fluorescence imaging technique, convenient for in vivo longitudinal monitoring of the animal brain without the involvement of cranial window opening operation. This chapter aims to provide the protocols for FMT in vivo imaging of Aβ deposits in the brain of rodent model of Alzheimer's disease. The materials, stepwise method, notes, limitations of FMT, and emerging opportunities for FMT techniques are presented.

Satellite-Based On-Orbit Printing of 3D Tumor Models.

Space three dimension (3D） bioprinting provides a precise and bionic tumor model for evaluating the compound effect of the space environment on tumors, thereby providing insight into the progress of the disease and potential treatments. However, space 3D bioprinting faces several challenges, including prelaunch uncertainty, possible liquid leakage, long-term culture in space, automatic equipment control, data acquisition, and transmission. Here, a novel satellite-based 3D bioprinting device with high structural strength, small volume, and low weight (<6kg) is developed. A microgel-based biphasic thermosensitive bioink and suspension medium that supports the on-orbit printing and in situ culture of complex tumor models is developed. An intelligent control algorithm that enables the automatic control of 3D printing, autofocusing, fluorescence imaging, and data transfer back to the ground is developed. To the authors' knowledge, this is the first time that on-orbit printing of tumor models is achieved in space with stable morphology and moderate viability via a satellite. It is found that 3D tumor models are more sensitive to antitumor drugs in space than on Earth. This study opens up a new avenue for 3D bioprinting in space and offers new possibilities for future research in space life science and medicine.

Investigation of Praseodymium Ions Dopant on 9/65/35 PLZT Ceramics’ Behaviors, Prepared by the Gel-Combustion Route

In this work, were synthesized (Pb0.91La0.09)(Zr0.65Ti0.35)0.9775O3 ceramic materials with different concentrations of praseodymium (0, 0.1, 0.3, 0.5, 1 wt.%) via gel-combustion route and sintered by the hot uniaxial pressing method. Measurements were conducted on the obtained ceramics using X-ray powder diffraction (XRD), scanning electron microscope (SEM), EDS analysis, and examination of dielectric and ferroelectric optical properties. Results give us a detailed account of the influences of the praseodymium ions on the structural, microstructural, and dielectric properties. 3D fluorescence maps and excitation and emission spectra measurements show how a small admixture changes the ferroelectric relaxor behavior to an optically active ferroelectric luminophore.

A novel 3D-fluorescence sensing strategy based on HN-chitosan polymer probe for rapid identification and quantification of potential adulteration in saffron.

Saffron is a candidate for various kinds of fraud to make huge profits. The present study proposed an efficient three-dimensional (3D) fluorescence sensing strategy based on hydrophilic hydrazine-naphthalimide functionalized chitosan (HN-chitosan) polymer probe for rapid identification and quantification of potential adulteration in saffron. The amino functional group in the HN-chitosan probe reacted specifically with the Oxygen-containing group of active ingredients in saffron, amplifying the signal difference between saffron and the adulterants, which was comprehensively characterized by 3D fluorescence. Four advanced chemometrics methods were applied for the classification of saffron and adulterated saffron, and good performance were obtained in both training and prediction sets. Furthermore, the PLS regression model was applied to the prediction of adulteration level in saffron and showed satisfactory accuracy. This strategy provides a new solution for rapid identification and quantification of potential adulteration in saffron, which contributes to the healthy development of its industry.

Partial inhibition of complex I restores mitochondrial morphology and mitochondria‐ER communication in hippocampus of APP/PS1 mice

AbstractBackgroundAlzheimer’s Disease (AD) has no cure. We showed that partial inhibition of mitochondrial complex I (MCI) with small molecule CP2 induced adaptive stress response activating multiple neuroprotective mechanisms. Chronic treatment reduced inflammation, improved synaptic and mitochondrial functions, and blocked neurodegeneration in symptomatic APP/PS1 mice, a translational model of AD.MethodsAnalysis of mitochondrial morphology and mitochondria‐endoplasmic reticulum (ER) contacts (MERCS) was done in the hippocampal CA1 tissue from APP/PS1 mice treated with CP2 and compared to APP/PS1 and non‐transgenic mice treated with vehicle using serial block‐face scanning electron microscopy (SBFSEM) and three‐dimensional (3D) EM reconstructions combined with western blot analysis and next generation RNA sequencing.ResultsWe demonstrate that CP2 treatment restores mitochondrial morphology and mitochondria‐ ER communication, reducing ER stress in the APP/PS1 mouse brain. Using 3D EM reconstructions, we found that mitochondria in AD dendrites existed primarily as mitochondria‐on‐a‐string (MOAS), a morphological phenotype that is difficult to detect using conventional 2D EM or fluorescence spectroscopy. Compared to other morphological phenotypes, MOAS are extensively enveloped in the ER membranes forming multiple MERCS known to contribute to abnormal lipid and calcium homeostasis, and ER stress in AD. CP2 treatment specifically reduced MOAS formation, consistent with improved energy homeostasis in the brain, with concomitant reduction in MERCS, ER stress, and improved lipid homeostasis.ConclusionsThese data provide novel information on the role MOAS play in AD. Our data demonstrate that in response to energetic stress, mitochondria in dendrites acquire MOAS phenotype with increased interaction with the ER, which may synergistically contribute to the AD development. Since CP2 treatment alleviated the ER stress and improved mitochondrial dynamics and function, these data further support the development of partial MCI inhibitors as disease modifying strategy for AD.

Adsorption of clarithromycin on polystyrene nanoplastics surface and its combined adverse effect on serum albumin

Emerging contaminants, such as antibiotics and nanoplastics, have garnered significant attention due to their potential adverse effects on diverse ecosystems. Antibiotic adsorption on the surface of nanoplastics potentially facilitates their long-range transport, leading to the synergistic effects of the complex. This research aims to examine the adsorption behavior of clarithromycin binding with polystyrene nanoplastics surface as well as their interaction between drug adsorbed polystyrene nanoplastics with serum albumin. Different spectroscopic methods were used to find out the interaction between clarithromycin and nanoplastics, under stimulated physiological conditions UV–vis spectroscopy showed a maximum of 22.8% percentage of the drug adsorbed with the polystyrene nanoplastics surface after 6 h of incubation. The fluorescence spectroscopic results demonstrated that the fluorescence intensity of serum albumin was quenched by the clarithromycin-polystyrene nanoplastics (CLA-PSNP) complex through static quenching. We calculated the number of binding stoichiometry, binding constants, and thermodynamic parameters. This study revealed that the CLA-PSNP binds to serum albumin spontaneously and its hydrophobic interactions played a significant role. The conformational changes in the structure of serum albumin were revealed from the findings of synchronous fluorescence spectra, CD spectra, and 3D fluorescence spectra, leading to the disturbance in functional activity. This study focuses valuable insights into the intermolecular interactions between clarithromycin-adsorbed polystyrene nanoplastics and serum albumin and its potential molecular-level biological toxicity.

Small molecule linoleic acid inhibiting whey syneresis via interact with milk proteins in the fermentation of set yogurt fortified with c9,t11-conjugated linoleic acid.

The study aimed to investigate the impact of fermentation conditions on c9,t11-conjugated linoleic acid (CLA) synthesis by Lactobacillus casei, as well as its effects on whey syneresis, water holding capacity (WHC), and texture characteristics of set yogurt. The amount of whey syneresis decreased about 30% with the adding of 0.1% linoleic acid (LA). The interaction between LA and casein (CS), β-lactoglobulin (β-Lg) and bovine serum albumin (BSA) was observed by UV-Vis absorption spectroscopy, 3D fluorescence spectroscopy and CD spectroscopy. It found that LA changed the microenvironment and polarity around amino acids, as well as the conformation of the three milk proteins. Scanning electron microscope (SEM) analysis revealed that the addition of LA resulted in a more uniform and compact microstructure of the set yogurt. It indicates that LA can promote the crosslink of milk proteins, which may be the reason for the reduction of whey syneresis in set yogurt.

Monitoring of cellulose-rich biowaste co-digestion with 3D fluorescence spectroscopy and mass spectrometry-based metabolomics

Anaerobic digestion (AD) is a promising waste management strategy that reduces landfilling while generating biogas. Anaerobic co-digestion involves mixing two or more substrates to enhance the nutrient balance required for microorganism growth and thus improve the degradation. Monitoring AD is crucial for comprehending the biological process, optimizing process stability, and achieving efficient biogas production. In this work, we have used three dimensional excitation emission fluorescence spectroscopy and mass spectrometry metabolomics, two complementary techniques, to monitor the anaerobic co-digestion (AcoD) of cellulose, ash wood or oak wood with food waste. The two approaches were compared together and to the biogas production records. Results of this experiment demonstrated the complementarity of both analytical techniques with the measurement of the biogas production since 3D fluorescence spectroscopy and MS metabolomics revealed the earlier molecular changes occurring in the bioreactors, mainly associated with the hydrolysis step, whereas the biogas production data reflected the biological activity in the last step of the digestion. Moreover, in all cases, the three data sets effectively delineated the differences among the substrates. While the two wood substrates were poorly degradable as they were richer in aromatic compounds, cellulose was highly degradable and was characterized by the production of several glycolipids. Then, the three tested AcoDs resulted in a similar 3D EEM fluorescence and metabolomics profiles, close to the one observed for the AD of food waste alone, indicating that the incorporation of the food waste drove the molecular degradation events in the AcoDs. Substrate-specific differences were appreciated from the biogas production data. The overall results of this research are expected to provide insight into the design of guidelines for monitoring AcoD.

Synthesis and biological evaluation of indole derivatives containing thiazolidine-2,4-dione as α-glucosidase inhibitors with antidiabetic activity

In order to develop potential α-glucosidase inhibitors with antidiabetic activity, twenty-six indole derivatives containing thiazolidine-2,4-dione were synthesized. All compounds presented potential α-glucosidase inhibitory activities with IC50 values ranging from 2.35 ± 0.11 to 24.36 ± 0.79 μM, respectively compared to acarbose (IC50 = 575.02 ± 10.11 μM). Especially, compound IT4 displayed the strongest α-glucosidase inhibitory activity (IC50 = 2.35 ± 0.11 μM). The inhibition mechanism of compound IT4 on α-glucosidase was clarified by the investigation of kinetics studies, fluorescence quenching, CD spectra, 3D fluorescence spectra, and molecular docking. In vivo antidiabetic experiments demonstrated that oral administration of compound IT4 would suppress fasting blood glucose level and ameliorate their glucose tolerance and dyslipidemia in diabetic mice.

Interaction mechanism of okra (Abelmoschus esculentus L.) seed protein and flavonoids: Fluorescent and 3D-QSAR studies

The binding capacity of 10 flavonoids with okra seed protein (OSP) was studied by fluorescence spectroscopy. The structure of flavonoids had an obvious impact on binding performance. The binding ability of flavanone was lower than that of flavone, isoflavone and dihydrochalcone. The binding capacity of flavonoid glycoside was superior to that of the corresponding flavonoid aglycone. The binding ability was positively correlated with the number of phenolic hydroxyl groups on the B ring. The steric field and electrostatic field model constructed by 3D-QSAR method could well explain the above interaction behavior. Thermodynamic analysis suggested that the quenching mechanism of OSP caused by flavonoids was static quenching, and the binding-site number was 1. In addition, hydrogen bonding and van der Waals force dominated this interaction. The 3D and synchronous fluorescence spectra showed that there was no significant change in the polarity of the environment around tryptophan and tyrosine residues during binding.

Steady-state and 3D fluorescence study reveals the binding of a dicoumarol analogue in subdomain IIA of human serum albumin with structural variation

Drugs exhibit pharmacological properties through their binding interactions with the target receptors. Coumarins and dicoumarols are known to exhibit diverse pharmacological potential. 3,3′-Carbonylbis (7-diethylamino coumarin) (CDC) is a bis-coumarin derivative with antioxidant activity. The interaction of CDC to human serum albumin (HSA) has been reported here. The fluorescence quenching of HSA by CDC at 303, 308, and 313 K revealed a dynamic quenching with a quenching constant of 104 M−1 order. The binding constant from spectroscopic methods was found to be 103 M−1 order. Thermodynamic parameters (∆S0, ∆H0, and ∆G0) were calculated from temperature studies using the Van't Hoff equation. A negative ∆G0 value showed the spontaneity of the interaction. A positive ∆S0 value and negative ∆H0 value indicated the major role of an electrostatic force. Further the binding was found to be an entropy-driven process. Site-specific marker displacement studies with 1-Anilino-8-naphthalene sulfonate (ANS), warfarin, and ibuprofen revealed CDC binding at site-1 in subdomain IIA. HSA conformational changes were verified using synchronous and 3D fluorescence studies. Secondary structural changes in HSA were visible from circular dichroism results. Molecular docking results corroborated the experimental finding showing CDC binding at site-1 in HSA.