Fluorescence Measurements Research Articles

Nondestructive diagnosis of mineral deficiencies in wheat leaves by one- or two-shot saturation pulse measurement of chlorophyll fluorescence and P700+ absorbance with machine learning

We hydroponically cultivated wheat and subjected them to mineral deficiency by a week-long exposure to Hoagland's solution minus one essential mineral. Chlorophyll fluorescence and P700+ absorbance were measured under a single 300 ms saturating light pulse (one-shot) or the conventional two pulses (two-shot). A machine learning-based multiclass classification model was developed to assess mineral deficiency diagnosis. The results indicated that partial least squares discriminant analysis (PLS-DA) was ineffective. Conversely, linear discriminant analysis (LDA) without principal components analysis (PCA) successfully identified 13 or 11 (with a class combining the control with Mo- and B-deficiencies; and ten other distinct classes for respective mineral deficiencies) classes of mineral deficiencies, achieving over 81 % diagnostic performance across all five macro metrics of precision, recall, specificity, accuracy, and F1 measure. The one-shot method exhibited only a slight decrease in performance compared to the two-shot method, highlighting the advantage of substantially reduced measurement time. Applying the developed LDA models to outdoor soil-grown leaf data revealed their practicality and limitations. In multiple deficiency scenarios, the model predicted one predominant deficiency or incorrectly diagnosed a different deficiency. This study is the first to quantitatively demonstrate the efficacy of ultra-fast nutritional diagnostic technology in identifying root zone mineral deficiencies using chlorophyll fluorescence and P700+ absorbance measurements. It delineates the requirements for constructing multilabel or quantitative calibration models for field deployment.

The effect of magnesium impurities on the optical and spectral characteristics of calcium orthovanadate single crystals doped with chromium and manganese

The effect of magnesium co-doping on growth conditions, crystal composition, optical and spectral properties of manganese or chromium-doped Ca3(VO4)2 (CVO) crystal was studied. The single crystals were grown by the Czochralski method. Effective segregation coefficients of manganese and chromium in the CVO matrix with magnesium co-doping were calculated. A redistribution of the absorption lines intensities was observed in the absorption spectra of CVO:Mn:Mg and CVO:Cr:Mg. Low (15 K) temperature fluorescence measurements confirmed that co-doping CVO crystal with Mg ions leads to the redistribution of Mn ions concentration in different valence states, but does not cause changes in their spectral properties. In the presence of magnesium, additional annealing of the materials resulted in a slower reduction of the valence state of doping ions and a noticeable change in crystal coloration. All investigated impurities have practically no effect on the specific ferroelectric domain structure of CVO.

Liquid Chromatography Mass Spectrometric Method and a Fluorometric 96 Well Plate Assay for Determination of Thiamine in Salmonid Eggs.

Thiamine deficiency is a large contributor to reduced reproduction success among salmonids throughout the northern hemisphere. In Scandinavia, this reproduction disorder is known as M74; while in North America, it is known as early mortality syndrome (EMS). The disorder fluctuates in magnitude from year to year. During years with high prevalence of the disorder, salmonid hatcheries that stock various aquatic systems to maintain the population size experience difficulties filling their quotas without thiamine treatment of alevins. The disorder is monitored both by observing the survival rate and by measuring the thiamine content of prefertilized eggs in the hatcheries. Here, a simple extraction procedure is presented, which allows for quantitative determination of the various phosphorylated forms of thiamine using liquid chromatography mass spectrometry but also allows for extraction in 96 deep-well plates and measurement of the total thiamine content using fluorescence monitoring with a plate reader, following oxidation of thiamine to thiochrome. The latter procedure could also be integrated into a highly portal system where the thiochrome is determined using the DeNovix QFX analyzer. The newly developed extraction procedure and cleanup method for fluorescence measurement represent the most versatile and simple methods to date for monitoring of thiamine in salmonid eggs. The methods produced accurate and precise data with quantification limits below the limit where the deficiency causes 100% lethality.

A comprehensive investigation of the nano-[Cu2-(DIP)2-EA] effects on HSA through spectroscopic procedures and computer simulations

In this research, the toxicity of nano-[Cu2-(DIP)2-EA], a metal nano-complex consisting of ellagic acid and bathophenanthroline ligands, on human serum albumin (HSA) at a protein level was investigated. Molecular docking simulations and spectral analyses were conducted in a simulated physiological environment at pH 7.4 to explore the interaction of nano-[Cu2-(DIP)2-EA] with HSA. The results represented an increase in albumin absorption upon exposure to nano-[Cu2-(DIP)2-EA], demonstrating significant interaction between the two compounds. Steady-state and time-resolved fluorescence measurements pointed out that nano-[Cu2-(DIP)2-EA] induced static quenching of the albumin's intrinsic fluorescence with a high binding affinity of approximately 106 mol/L in a 1:1 interaction ratio. The thermodynamic variables clarified that binding of nano-[Cu2-(DIP)2-EA] to albumin occurs spontaneously and primarily driven by van der Waals interactions and H-bonds. The results of the computer simulations and the binding displacement experiments utilizing the site markers warfarin and ibuprofen revealed that nano-[Cu2-(DIP)2-EA] binds to site I within the subdomain IIA of albumin. Circular dichroism analysis elaborated that nano-[Cu2-(DIP)2-EA] slightly perturbed the microenvironment around of tryptophan residues and diminished the α-helix structure stability to a negligible amount.

Fluorometer “FLS 10s”

Many biological objects have the ability to fluoresce under the influence of electromagnetic radiation in the optical range. Measuring the fluorescence of a biological object helps obtain information about its internal state. Therefore, the creation of instruments for scientific research in the field of studying biological objects has been a pressing task for many areas of human activity for many years. In particular, these are areas such as agriculture, ecology, food industry and medicine. This article is devoted to a description of the recently developed “FLS 10s” device for studying plant chlorophyll fluorescence, as well as description of software for operating the device and analyzing the information obtained. This device provides data of well known OJIP test. Keywords: fluorescence, fluorometer, fluorescence measurement, Kautsky effect, OJIP test, fluorescence induction, fast chlorophyll α fluorescence induction.

Monitoring cancer treatments in melanoma cells using a fluorescence lifetime nanoprobe based on a CdSe/ZnS quantum dot functionalized with a peptide containing D-penicillamine and histidine

Anticancer therapies with cisplatin and volasertib (BI-6727) were monitored by fluorescence lifetime imaging microscopy (FLIM) in live SK-Mel-2 melanoma cells. A CdSe/ZnS quantum dot functionalized with a peptide containing D-penicillamine and histidine (CdSe/ZnS-PH) was used as intracellular pH fluorescent probe. A faster cytosol acidification was observed for cells treated with cisplatin when compared to volasertib. The first changes in the intracellular pH were found after 2 hours of treatment with cisplatin and 8 hours with volasertib. Additionally, the relationship between cytosol acidification and apoptosis was investigated using an innovative methodology based on time-resolved fluorescence measurements. Similar low percentages of apoptotic cells were observed after short incubation periods (2 – 8 hours) with both drugs. In contrast, late apoptosis and death were found for a large fraction of cells during 24-hour incubation with cisplatin but not volasertib. Thus, the early acidification observed in cisplatin treatment could accelerate apoptosis and cell death. Despite volasertib treatment shows slower mechanism of action than cisplatin, similar inhibitory effects were found for both drugs at longer incubation periods (72 hours). This study proves the potential of CdSe/ZnS-PH nanoparticle as a fluorescence lifetime probe in the study of the mechanism of action of antitumor drugs.

Regime difference between macrophyte and Cyanophyta dominance regulate microbial carbon sequestration mode in lake sediments

Different nutrient load in the lake ecosystems trigger the regime difference and change of predominant biotype. The regulation of carbon (C) sequestration mode in the lacustrine sediments in response to this process need prudent clarification. Fluorescence measurements and high-throughput sequencing for functional genes cbbL and cbbM encoding C-fixing bacteria genus were executed for sediments from two representative regimes dominated by macrophyte and Cyanophyta, respectively. The results showed that humic-like and fulvic-acid like materials dominated the dissolved organic matter (DOM) from the algae and macrophyte-dominated lake regions, respectively. Microbial assimilation played critical influence on C fixation into the sediments in both of the two regimes. However, higher diversity was detected in macrophyte-dominated regime compared to that in Cyanophyta-dominated regime, suggesting that moderate nutrient levels facilitated the species richness of bacteria encoding functional genes concerning C fixation. Bacterial species and diversities varied between two regimes including predominant and rare taxa, suggesting that community structure alteration due to regime difference triggered the regulation the C sequestration mode and stability. Predominant genera manipulated the abundance of C-fixing bacteria genes in response to the regulation of nutrient levels. Noted that rare genera also responded to the regime difference and played key role in C sequestration into lacustrine sediments. Our results suggest that more abundant macrophyte-dominated regime facilitated the C sequestration in the lake ecosystems for atmospheric C reduction.

Laser absorption correction for hydroxyl planar laser induced fluorescence measurements in a centrally staged combustor at elevated pressures

Planar laser-induced fluorescence (PLIF) is a crucial spectroscopic technique for measuring minor species [e.g., hydroxyl (OH), methylene (CH), and nitric oxide (NO) radicals] in combustion research, owing to its non-intrusive nature and high sensitivity. However, laser energy attenuation due to absorption poses significant challenges to its application under high-pressure conditions, which may cause asymmetric image intensity distribution along the light propagation direction. An absorption correction method for OH PLIF based on the concept of maximum number density is proposed in the present study. This method offers several key advantages, including simplicity, high accuracy, and versatility, allowing for correcting both time-averaged and instantaneous OH PLIF images. OH PLIF data obtained from a centrally staged combustor at elevated pressures (i.e., 0.3, 0.6, and 1.0 MPa) are utilized to validate the method. Correction for the time-averaged PLIF images achieves a much more symmetric distribution of OH, revealing the overall flame structures that would not have been completely visualized from the original images. The fronts of the pilot and main stage flames have also been recovered from the corrected instantaneous images. This correction algorithm provides an effective way of enhancing data quality for high-cost OH PLIF measurements at pressurized conditions.

Synthesis and fluorescence properties of europium complex functionalized fiberglass paper.

The development of novel rare earth fluorescent materials and the exploration of their applications have consistently been focal points of research in the fields of materials science and chemistry. In this work, a novel rare earth composite material with good photo-fluorescence properties and self-supporting has been prepared via a simple ultrasonic solvent reaction method. Initially, the Phen moieties is immobilized onto the surface of a self-supporting fiberglass paper using ICPTES, followed by the coordination of Eu(TTA)3 moieties with Phen moieties through a convenient ultrasonic solvent reaction. The resulting GF-Phen-Eu(TTA)3 has been characterized using FTIR, UV-Vis DRS, fluorescence measurements, and so on. The results indicate that the composite material exhibits strong fluorescent emission and presents a vivid red color under ultraviolet light. Further research has shown that the fluorescence of GF-Phen-Eu(TTA)3 strips demonstrated a pronounced quenching effect in response to some transition metal ions (1 mM). Hence, the rare earth composite materials presented here can be utilized not only for the production of optical materials, but also for the development of fluorescence sensing strips.

A portable multi-taxa phenotyping device to retrieve physiological performance traits

Organismal phenotyping to identify fitness traits is transforming our understanding of adaptive responses and ecological interactions of species within changing environments. Here we present a portable Multi-Taxa Phenotyping (MTP) system that can retrieve a suite of metabolic and photophysiological parameter across light, temperature, and/or chemical gradients, using real time bio-optical (oxygen and chlorophyll a fluorescence) measurements. The MTP system integrates three well-established technologies for the first time: an imaging Pulse Amplitude Modulated (PAM) chlorophyll a fluorometer, custom-designed well plates equipped with optical oxygen sensors, and a thermocycler. We demonstrate the ability of the MTP system to distinguish phenotypic performance characteristics of diverse aquatic taxa spanning corals, mangroves and algae based on metabolic parameters and Photosystem II dynamics, in a high-throughput capacity and accounting for interactions of different environmental gradients on performance. Extracted metrics from the MTP system can not only provide information on the performance of aquatic taxa exposed to differing environmental gradients, but also provide predicted phenotypic responses of key aquatic organisms to environmental change. Further work validating how rapid phenotyping tools such as the MTP system predict phenotypic responses to long term environmental changes in situ are urgently required to best inform how these tools can support management efforts.

Synthesis and characterization of nickel and molybdenum catalysts supported on alumina derived from bauxite

In this study, a natural ore (bauxite), which is characterized by its high content of various aluminum minerals, such as gypsite, , bohmite, and diaspore, AlO(OH), was used to obtain alumina ( ), which was used as a support for the catalyst. The components of the ore were studied using multiple techniques, such as The X-ray energy dispersion (EDX) technique, X-ray diffraction (XRD) and X-ray fluorescence (XRF) technology. Then, undesirable components that negatively affect the behavior of the prepared catalyst were removed, such as carbonates, iron, and amorphous silica, and then aluminum oxide (alumina) was prepared from the ore using a series of chemical treatments. Then the mixed catalyst consisting of (nickel/molybdenum) carried on alumina was prepared, its properties were studied, and its components were identified by measuring energy dispersion with X-rays (EDX) and ensuring the extent of its thermal stability by conducting a thermal analysis measurement. Gravimetric analysis (TGA), differential thermal analysis (DTA), measurement of the surface area of ​​the prepared catalyst (BET), scanning electron microscope (SEM) measurement, as well as X-ray diffraction (XRD) and X-ray fluorescence (XRF) measurements to identify the percentages of metals, and then estimating the percentage of these elements in their oxide form. KEY WORDS: Catalyst, Bauxite, Nickel, Molybdenum Bull. Chem. Soc. Ethiop. 2024, 38(6), 1715-1724. DOI: https://dx.doi.org/10.4314/bcse.v38i6.17

Inner Mitochondrial Membrane Peptidase 2-Like Deletion Aggravates Mitochondrial Apoptosis and Inhibits Autophagy After Hyperglycemia Stroke.

This study investigated the effects of inner mitochondrial membrane peptidase 2-like (Immp2l) deletion on mitochondrial apoptosis and mitochondrial autophagy under hyperglycemic conditions. The middle cerebral artery occlusion (MCAO) model was established in wild-type (WT) mice and Immp2l+/- mice; animals were then exposed to hyperglycemic (induced using 1% streptozotocin) and normoglycemic conditions. Tissues were collected at various time points post-reperfusion. The production of reactive oxygen species (ROS) was assessed by fluorescent measurements, and mitochondrial membrane potential was evaluated using a JC-1 assay kit. Autophagy was analyzed by measuring LC3II/LC3I protein expression and Beclin 1 expression. Mitochondrial ultrastructure was examined through transmission electron microscopy (TEM); neuronal autophagosomes were also assessed. Immp2l mutation in a hyperglycemic environment exacerbated brain injury by increasing ROS production, compromising mitochondrial membrane potential, inducing apoptotic cascades, and impairing mitochondrial autophagy. These findings highlight the critical role of Immp2l in modulating the response to hyperglycemic cerebral ischemia-reperfusion (I/R) injury. Furthermore, the deficiency of Immp2l appears to contribute to increased oxidative stress, mitochondrial dysfunction, and cell death, thereby exacerbating brain injury. These data may provide new insights into therapeutic strategies for reducing the impact of diabetes on stroke outcomes.

Portable X‐ray fluorescence spectrometry accurately measures metal concentrations in aqueous Mehlich III soil extraction solutions

AbstractAccurate measurement of metal concentrations in soil and water is vital for healthy crop production and decision making for environmental surveys. While there are a multitude of laboratory‐based soil analysis methods, such as inductively coupled plasma‐optical emission spectroscopy (ICP‐OES), flame emission spectrometry, and atomic absorption spectroscopy, most are time and resource intensive. Additionally, there is a lack of information for rapid analysis of elements for aqueous soil extractions. The goal of this research is to establish elemental correlations between portable X‐ray fluorescence (pXRF) measurements of Mehlich III soil extractions and traditional elemental measurements via ICP‐OES. We hypothesize that certain metals can be accurately measured in aqueous soil extraction solutions by pXRF to the same degree as they are measured by ICP‐OES. To test this hypothesis, Mehlich III and 2% nitric acid solutions with known elemental concentrations were analyzed via ICP‐OES and pXRF. Soil samples extracted using Mehlich III were compared between ICP‐OES and pXRF to verify correlations. High correlations were found for As, Ca, Cd, Cr, Cu, Fe, Mn, Mo, Ni, Pb, Se, V, and Zn in both the Mehlich III and 2% nitric acid solutions at concentrations between 5 and 85 mg L−1. P, S, and Si did not show high correlations at concentrations <100 mg L−1. These results indicate that between 5 and 85 mg L−1, pXRF analysis of aqueous solutions and soil extractions is a reliable technique; however, at low concentrations (i.e., <5 mg L−1 for metals and <100 mg L−1 for P and S), pXRF is not well suited.

UV irradiation during fluorescence measurements affects the structures and fluorescence properties of isolated lignins in solution

There are no reports on the stability of autofluorescence, although lignins and their solutions are known to exhibit autofluorescence. An aim of this study is to investigate spectral change of lignin autofluorescence under prolonged excitation with ultraviolet (UV). Both fluorescence spectra of softwood milled wood lignin (MWL) and kraft lignin (SKL) in propylene glycol significantly changed upon an UV irradiation at 280 nm under O2 and Ar atmospheres even for 10 min. By the further irradiation, the autofluorescence intensities were remarkably changed together with the emission wavelengths. However, no tendency in the autofluorescence change was observed. The change varied between lignin preparations and gas atmosphere. These phenomena should be caused by autoxidation upon UV irradiation, and the reaction resulted in not only photodegradation but also photopolymerization.

Classification accuracy and compatibility across devices of a new Rapid-E+ flow cytometer

Abstract. The study evaluated a new model of a Plair SA airflow cytometer, Rapid-E+, and assessed its suitability for airborne pollen monitoring within operational networks. Key features of the new model are compared with the previous one, Rapid-E. A machine learning algorithm is constructed and evaluated for (i) classification of reference pollen types in laboratory conditions and (ii) monitoring in real-life field campaigns. The second goal of the study was to evaluate the device usability in forthcoming monitoring networks, which would require similarity and reproducibility of the measurement signal across devices. We employed three devices and analysed (dis-)similarities of their measurements in laboratory conditions. The lab evaluation showed similar recognition performance to that of Rapid-E, but field measurements in conditions when several pollen types were present in the air simultaneously showed notably lower agreement of Rapid-E+ with manual Hirst-type observations than those of the older model. An exception was the total-pollen measurements. Comparison across the Rapid-E+ devices revealed noticeable differences in fluorescence measurements between the three devices tested. As a result, application of the recognition algorithm trained on the data from one device to another led to large errors. The study confirmed the potential of the fluorescence measurements for discrimination between different pollen classes, but each instrument needed to be trained individually to achieve acceptable skills. The large uncertainty of fluorescence measurements and their variability between different devices need to be addressed to improve the device usability.

Elucidating the Binding Interaction of a Highly Efficient Phytochemical-Sinapic Acid with Human Serum Albumin: A Spectroscopic and Calorimetric Approach

Polyphenolic compounds have received much attention due to their major function as antioxidants. Sinapic acid (SA) is an orally bioavailable phytochemical with excellent free radical scavenging property transported in the blood by human serum albumin (HSA) which is the chief carrier protein found abundantly in human plasma. SA contains a single phenolic group. This study investigates the binding and interaction mechanism between SA and HSA by multiple spectroscopic and calorimetric techniques, such as UV/visible absorption spectroscopy, intrinsic fluorescence studies, Fourier transform infrared (FTIR) spectroscopy and isothermal titration calorimetry (ITC). UV/visible absorption spectroscopy showed hyperchromicity in the absorption spectra, suggestive of structural change due to complex formation between HSA and SA. Intrinsic fluorescence measurements, performed at three different temperatures, showed quenching of the fluorescence spectra and the mechanism of quenching was static in nature which occurred due to ground state complex formation. The FTIR results demonstrated a 16% decrease in α-helical content of the secondary structure of HSA in the presence of SA. Hydrogen bonds and hydrophobic forces were the main forces of interaction, according to the thermodynamic characteristics found through ITC. Moreover, the data obtained through ITC indicated strong binding affinity between HSA and SA and supported the exothermic and spontaneous nature of the interaction.

Depth detection limit of a fluorescent object in tissue-like medium with background emission in continuous-wave measurements: a phantom study.

Although the depth detection limit of fluorescence objects in tissue has been studied, reports with a model including noise statistics for designing the optimum measurement configuration are missing. We demonstrate a variance analysis of the depth detection limit toward clinical applications such as noninvasively assessing the risk of aspiration. It is essential to analyze how the depth detection limit of the fluorescence object in a strong scattering medium depends on the measurement configuration to optimize the configuration. We aim to evaluate the depth detection limit from theoretical analysis and phantom experiments and discuss the source-detector distance that maximizes this limit. Experiments for detecting a fluorescent object in a biological tissue-mimicking phantom of ground beef with background emission were conducted using continuous wave fluorescence measurements with a point source-detector scheme. The results were analyzed using a model based on the photon diffusion equations. Then, variance analysis of the signal fluctuation was introduced. The model explained the measured fluorescence intensities and their fluctuations well. The variance analysis showed that the depth detection limit in the presence of ambient light increased with the decrease in the source-detector distance, and the optimum distance was in the range of 10 to 15mm. The depth detection limit was found to be with this optimum distance for the phantom. The presented analysis provides a guide for the optimum design of the measurement configuration for detecting fluorescence objects in clinical applications.

Identification and characterisation of functional Kir6.1-containing ATP-sensitive potassium channels in the cardiac ventricular sarcolemmal membrane.

The canonical Kir6.2/SUR2A ventricular KATP channel is highly ATP-sensitive and remains closed under normal physiological conditions. These channels activate only when prolonged metabolic compromise causes significant ATP depletion and then shortens the action potential to reduce contractile activity. Pharmacological activation of KATP channels is cardioprotective, but physiologically, it is difficult to understand how these channels protect the heart if they only open under extreme metabolic stress. The presence of a second KATP channel population could help explain this. Here, we characterise the biophysical and pharmacological behaviours of a constitutively active Kir6.1-containing KATP channel in ventricular cardiomyocytes. Patch-clamp recordings from rat ventricular myocytes in combination with well-defined pharmacological modulators was used to characterise these newly identified K+ channels. Action potential recording, calcium (Fluo-4) fluorescence measurements and video edge detection of contractile function were used to assess functional consequences of channel modulation. Our data show a ventricular K+ conductance whose biophysical characteristics and response to pharmacological modulation were consistent with Kir6.1-containing channels. These Kir6.1-containing channels lack the ATP-sensitivity of the canonical channels and are constitutively active. We conclude there are two functionally distinct populations of ventricular KATP channels: constitutively active Kir6.1-containing channels that play an important role in fine-tuning the action potential and Kir6.2/SUR2A channels that activate with prolonged ischaemia to impart late-stage protection against catastrophic ATP depletion. Further research is required to determine whether Kir6.1 is an overlooked target in Comprehensive in vitro Proarrhythmia Assay (CiPA) cardiac safety screens.

Reduced diffusional limitations in carnation stems facilitate higher photosynthetic rates and reduced photorespiratory losses compared with leaves.

Green stem photosynthesis has been shown to be relatively inefficient but can occasionally contribute significantly to the carbon budget of desert plants. Although the possession of green photosynthetic stems is a common trait, little is known about their photosynthetic characteristics in non-desert species. Dianthus caryophyllus is a semi-woody species with prominent green stems, which show similar photosynthetic anatomy with leaves. In the present study, we used a combination of gas exchange and chlorophyll fluorescence measurements, some of which were taken under varying O2 and CO2 partial pressures, to investigate whether the apparent anatomical similarities between the species' leaves and stems translate into similar photosynthetic physiology and capacity for CO2 assimilation. Both organs displayed high photosynthetic electron transport rates (ETR) and similar values of steady-state non-photochemical quenching (NPQ), albeit leaves could attain them faster. The analysis of OJIP transients showed that the quantum efficiencies and energy fluxes along the photosynthetic electron transport chain are largely similar between leaves and stems. Stems displayed higher total conductance to CO2 diffusion, similar biochemical properties, significantly higher photosynthetic rates and lower water use efficiency than leaves. Leaf ETR was more sensitive to sub-ambient O2 and super-ambient CO2 partial pressures, while leaves also displayed a higher relative rate of Rubisco oxygenation. We conclude that the highly responsive NPQ and the enhanced photorespiration and WUE in leaves represent photoprotective and water-conserving adaptations to the high incident light intensities they experience naturally, at the expense of higher CO2 assimilation rates, which the vertically orientated stems can readily attain.

In vitro and in silico DNA interaction studies of a series of 3-carboxamide-4-quinolone derivatives prepared from an one-pot stepwise synthesis (OPSS) method

The 3-carboxamide-4-quinolone framework is highly regarded in the fields of Medicinal and Bioorganic Chemistry mainly due to its widespread bioactive properties. This versatile scaffold allows for the development of compounds with various pharmacological effects. In this sense, the present work explores a novel one-pot stepwise (OPPS) method for synthesizing fifteen 3-carboxamide-4-quinolone derivatives, some of which exhibit antitumor profiles in previous works. These compounds were synthesized using the OPPS methodology with yields ranging from 35 % to 88 %, streamlining the synthetic process and reducing solvent usage. Furthermore, interaction studies with calf-thymus DNA were done by UV–Vis absorption and steady-state fluorescence measurements combined with molecular docking calculations. Compounds 10a-b, 10d-g, 10k, and 10n were selected for the CT-DNA interaction assays, from which it was possible to demonstrate their DNA-binding property through peripheral secondary interactions. Molecular docking results suggested van der Waals and hydrogen bonds as the main binding forces responsible for the interactive profile between the nucleobases and the 3-carboxamide-4-quinolone derivatives. All these results unveiled promising insights into their role as DNA replication inhibitors, highlighting their potential in developing antiproliferative drugs.