Nickel-induced oxidative stress causes cell death in testicles: implications for male infertility.

Aligarh region is well known for its lock industry. This lock industry utilises nickel for electroplating. There have been informal reports of infertility in men and women living near the lock industry. We analysed field water samples to investigate this link, and the results showed considerable nickel contamination. To further validate our results, we exposed male rats to relevant nickel levels in drinking water. This experimental exposure resulted in abnormal sperm morphology, decline in sperm count, significant change in activities of antioxidant enzymes, pronounced oxidative stress in the rat spermatocytes and decrease in serum testosterone level, as well as damage in the hypothalamus and pituitary (in all cases, the changes were most significant at the highest concentration used i.e 2.5 mg/l). The breeding experiments showed decline in live birth rate, while pups did not survive post birth in cages where males were given 2 and 2.5 mg/l concentrations of nickel in drinking water prior to mating. Our data strongly indicate a link between industrial nickel exposure and male infertility.

Antiglycation and antioxidant potential of coumaric acid isomers: a comparative in-vitro study.

Advanced glycation end products (AGEs) are the product of non-enzymatic glycation of serum proteins. AGEs increase reactive oxygen species (ROS) formation, which leads to diabetic complications. Phytochemicals exhibit lesser side effects as compared to conventional therapy. In this study, three isomers of coumaric acid (ortho, meta, para) were used to deduce the better one in terms of reducing diabetic complications. For this purpose, human serum albumin (HSA) was incubated with glucose in the absence and presence of isomers for 28 days. To avoid any growth, NaN3 was added and temperature was kept constant throughout the incubation period. Studies like fluorescence, circular dichroism spectroscopy, fructosamine analysis, free lysine estimation, free thiol group estimation were done. To investigate the ROS production, fluorescence microscopy of isolated lymphocytes using DAPI and dichloro-dihydro-fluorescein diacetate were performed. Molecular docking and molecular dynamic simulations (root-mean-square deviation, root-mean-square fluctuations, radius of gyration and solvent-accessible surface area) of HSA and peroxisome proliferator activated receptor (PPAR) alpha and gamma were also done. It was observed that in glycated protein samples, the level of absorbance, fluorescence, fructosamine and carbonyl group increased along with the loss of secondary structure, free lysine and thiol group. These parameters were found gradually recovered in treated samples. ROS production and apoptosis were found to be reduced in lymphocytes treated with p-Coumaric acid (pCA)-treated protein samples as compared to lymphocyte treated with glycated protein. Computational modelling suggested a stable complex formation of HSA and PPARs with pCA. Results with pCA at 200 µM were consistently better than other two isomers. Our next step is to evaluate this study in rats.Communicated by Ramaswamy H. Sarma.

Antiglycation, antifibrillation and antioxidative effects of para coumaric acid and vitamin D; an in-vitro and in-silico comparative-cum-synergistic approach.

Diabetes Mellitus is a metabolic disorder that results in impaired utilization of carbohydrates, lipids, and proteins. Severe hyperglycemia is its principal clinical symptom. Human serum albumin (HSA) is used as a model protein since it is viewed as a sign of glycaemic management because it is more likely to get glycated in diabetic people than other proteins. Para-coumaric acid (pCA), a phenolic acid, and Vitamin D (vit-D) are used as protective agents. In the present work, we deduce a synergistic-cum-comparative effect of pCA and vit-D, expecting some improvement in the efficacy of pCA when combined with vit-D. Methods employed are DPPH radical scavenging activity, In-vitro glycation of HSA, UV-vis spectroscopy, fluorescence analysis, and circular dichroism measurement. After treatment, increasein the absorbance and fluorescence intensity were reduced along with normalization of CD value. . The glycation-mediated fibrillation assessed by Congo-Red and Thioflavin T (ThT) were found to be diminishedwhen HSA was treated with equimolar concentration of p-CA and vit-D- treatment. Fructosamine adduct formation and lysine modificationwas also decreased, while inhibition to hemolysis and lipid peroxidation was found to increase upon treatment. The reactive oxygen species generation detection was also performed in lymphocytes treated with various protein samples. Docking results further confirmed theblocking some glycation-prone amino acids by both compounds. The study shows that the combination in the ratio of 1:1 has provided higher overall protection comparable to aminoguanidine (AG), the molecule which is utilized as a positive control.

Increased therapeutic effect of nanotized silibinin against glycation and diabetes: An in vitro and in silico-based approach.

BACKGROUND: The global prevalence of diabetes has increased sharply, with the number of cases expected to rise from 424.9 million in 2017 to 628.6 million by 2045. Flavonoids are plant derived molecules with well-established antioxidant potential in addition to other therapeutic properties. Silibinin is a naturally occurring flavonoid with antioxidant and antidiabetic properties. However, its rapid metabolism and low bioavailability limit its therapeutic effects. AIMS & OBJECTIVES: In this study, we have synthesized the nanoformulation of silibinin and compared its antiglycating and antidiabetic potential with the soluble form. METHODOLOGY: The inhibitory effect was tested on carbohydrate-hydrolyzing enzymes as well as glycation of human serum albumin (HSA). The structural and biochemical changes in HSA were assessed by spectroscopic analyses and different assays. KEY FINDINGS: The nanoforms were found to be better inhibitors of α-amylase and α-glucosidase compared to the bulk forms. Glycation of HSA in the presence of nano-silibinin resulted in the formation of lower level of early and advanced glycation products. This was also confirmed by spectroscopic studies and by estimating protein oxidation and free lysine residues. Molecular docking studies further supported the experimental outcomes. These results indicate that the nano form has significantly stronger antidiabetic and antiglycating effects than the bulk form. Nano-silibinin could therefore be recommended as a dietary supplement for diabetics to help control glycation and other associated complications.

Insight into the In Vitro Antiglycation and In Vivo Antidiabetic Effects of Thiamine: Implications of Vitamin B1 in Controlling Diabetes.

Hyperglycemia is considered to be a driving factor for advanced glycated end products (AGEs) formation. Inhibition of this process plays a vital role in reducing the problems of diabetes. This study aimed to explore the in vitro antiglycation and in vivo antidiabetic effect of thiamine. Human serum albumin (HSA) was used as a model protein to delineate the antiglycation potential of thiamine. Fructosamine levels were low in the presence of thiamine, implying the inhibition of early stages of glycation by thiamine. Furthermore, HSA-glucose assays depict the inhibition of post-Amadori products by thiamine. CD spectroscopy suggested fewer alterations in the secondary structure in the presence of thiamine. It was found that the administration of thiamine to diabetic rats leads to an increase in hexokinase activity and increased insulin secretion coupled with glycolysis utilization of glucose. Moreover, the activity of glucose-6-phosphatase and fructose- 1-6-phosphatase (increased in the liver and kidney of diabetic rats) is restored to near-normal levels upon thiamine administration. Histopathological studies also advocated that thiamine supplementation decreases the pathological abnormalities associated with diabetes in the liver and kidney. This study provides a rationale that vitamins can be implicated in controlling diabetes.

Mechanistic insight into glycation inhibition of human serum albumin by vitamin B9: Multispectroscopic and molecular docking approach.

Advanced glycation end products (AGEs) formation produces free radicals that play a role in diabetes mellitus; hence inhibition of glycation plays a part in minimizing diabetes-related complications. This study was intended to examine the AGEs formation of HSA upon prolonged incubation of 28 days at 37 °C and further investigate the antiglycation potential of folic acid (FA). FA shows a significant binding affinity to the HSA with a binding constant (K) of 104 M-1. The evaluation of enthalpy change (∆H0) and entropy change (∆So) implied that the HSA-FA complex is stabilized primarily by hydrophobic interaction and hydrogen bonding. Molecular docking analysis depicted that FA binds with HSA in subdomain IIA (Sudlow's site I) with a binding energy of -7.0 kcal mol-1. AGEs were characterized by free lysine and thiol groups, carbonyl content, and AGEs specific fluorescence. The presence of FA significantly decreased glycation from free lysine and carbonyl content estimation and AGEs specific fluorescence. Multispectroscopic observations and molecular docking and examination of various biomarkers demonstrate the antiglycation activity of FA and its capacity to prevent disease progression in diabetes.

Elucidating the interaction of aminophylline with calf thymus DNA using multispectroscopic and molecular docking approach.

Aminophylline (Am) is a methylxanthine compound clinically applied for chronic lung diseases like asthma, bronchitis or emphysema. Chemically, it comprises theophylline and ethylenediamine in a ratio 2:1. For the widening of the therapeutic window of any class of drug or for the designing of the newer therapeutic compound, an insight into the binding mechanics of available drugs with DNA is quite imperative. In view of that, here in this study we have investigated binding mechanics of aminophylline molecule with calf thymus DNA (Ct-DNA) using various spectroscopic techniques as well as molecular docking approach. Spectral analysis employing UV-visible and fluorescence approach confirmed the formation of aminophylline-Ct-DNA complex. The binding constant was calculated as 3.5 × 104 M-1 with 0.90 as the value of binding site suggestive of minor groove binding mode of aminophylline. The groove binding mode was further confirmed through spectrofluorimetric experiments like competitive displacement assay employing ethidium bromide, hoechst and rhodamine 6 G dyes as well as iodide quenching studies. The circular dichroic spectral evaluation and molecular docking study finally validated the minor groove binding mode of aminophylline with binding energy calculated as -4.5 Kcal/mol.

Causing DNA damage and stopping DNA repair - Vitamin D supplementation with Poly(ADP-ribose) polymerase 1 (PARP1) inhibitors may cause selective cell death of cancer cells: A novel therapeutic paradigm utilizing elevated copper levels within the tumour.

Inhibitors of PARP1 are currently being used to treat malignancies and as maintenance drugs post chemotherapy. Vitamin D has been shown to have an anticancer effect (mediated by multiple mechanisms) and is usually deficient in cancer patients. We had previously proposed and experimentally validated the selective cell death of cancer cells caused due to interaction of vitamin D with elevated copper within malignant cells, leading to ROS mediated DNA damage. In this communication we propose a Vitamin D supplementation strategy with PARP1 inhibitor treatment which would have multifaceted benefits for therapy. Besides exerting its anticancer effects by other mechanisms and addressing the deficiency, Vitamin D in principle would cause selective ROS mediated DNA breakage in malignant cells, while sparing healthy non malignant cells. Since Vitamin D is also a known inhibitor of PARP1, this therapeutic strategy would push the malignant cells to apoptosis due to DNA breakage via the vitamin D-copper mechanism, in addition to inhibiting DNA repair. Since the available levels copper within normal cells are several fold less as compared to malignant cells, the normal cells would be spared of the Vitamin D induced, ROS mediated DNA damage and would be less likely to die due to PARP1 inhibition.

How climate change affects parasites: the case of trematode parasite Clinostomum complanatum and its fish host Trichogaster fasiatus.

This study was undertaken to understand the impact of climate change on the ecology of infection of Clinostomum complanatum, a model trematode parasite. We analysed climate change data and data from infected fish over a period of seven years (2007-2013) from the Aligarh region (India) in this retrospective study. We show that infection of the trematode parasite Clinostomum complanatum (Rudolphi, 1814) in the forage fish Trichogaster facsiatus (Bloch & Schneider, 1801) is dependent on surface air temperature amongst the (ecologically) relevant climate change variables for both the parasite and its host. This study is the first to implicate surface air temperature as an environmental variable that may contribute towards parasitism, particularly for parasites with a piscine host. The biological relevance of changing climate on the ecology of this parasite is discussed.

Redox cycling of copper by coumarin-di(2-picolyl)amine hybrid molecule leads to ROS-mediated modulation of redox scavengers, DNA damage and cell death in diethylnitrosamine induced hepatocellular carcinoma.

Targeted therapy is a new strategy for cancer treatment that targets chemical entities specific to cancer cells than normal ones. One of the features associated with malignancy is the elevated copper which plays an integral role in angiogenesis. Work is in progress in our lab to identify new copper chelators to target elevated copper under targeted therapy for the killing of cancer cells. Recently, a coumarin-based copper chelator, di(2-picolyl)amine-3(bromoacetyl)coumarin hybrid molecule (ligand-L) has been synthesized by us, and also studied its copper-dependent macromolecular damage response in copper overloaded lymphocytes. The present study investigates the anticancer activity of ligand-L and its mode of action in rat model of diethylnitrosamine (DEN) induced hepatocellular carcinoma. It has been found that liver tissue has a marked increase in copper levels in DEN induced hepatocellular carcinoma. Ex vivo results showed that ligand-L inhibited cell viability, induced reactive oxygen species (ROS) generation, DNA damage, loss of mitochondrial membrane potential and caspase-3 activation in isolated hepatocellular carcinoma cells (HCC). All these effects induced by ligand-L were abrogated by neocuproine and N-acetylcysteine (ROS scavenger). Further, ligand-L treatment of animals bearing hepatocellular carcinoma results in an increment in the cellular redox scavengers, lipid peroxidation and DNA breakage in malignant hepatocytes. In vivo studies using ligand-L also showed that ligand-L possesses anticancer properties as evidenced by improvement in liver marker enzymes and liver surface morphology, and reduced alpha-fetoprotein in the treated group compared to untreated cancer-induced group. Overall, this study suggests that copper-ligand-L interaction leads to ROS generation which caused DNA damage and apoptosis in malignant cells. This study provides enough support to establish ligand-L as a clinically relevant lead molecule for the treatment of different malignancies.

Nickel toxicology with reference to male molecular reproductive physiology.

The toxicity of metals is a known phenomenon. Nickel toxicity is very common since nickel is used extensively both industrially and in items of personal use such as utensils and jewellery. Here we discuss human exposure to nickel and its toxicity in the light of the available scientific evidence to understand its underlying pathophysiology. The ability of Ni+2 to get oxidized to Ni+3 renders it's potential of generating reactive oxygen species (ROS) in the system leading to oxidative stress. Carcinogenesis, apoptosis induction, contact dermatitis, epigenetic changes, and alteration in gene regulation are a result of overexposure of nickel. Our focus is on how nickel affects the male reproductive physiology. Nickel primarily drives ROS mediated perturbations in the male reproductive system. It influences zinc metabolism, which is critical for sperm stability and affects the structure of DNA binding proteins, including protamines, thereby affecting sperm function.

Anti-diabetic study of vitamin B6 on hyperglycaemia induced protein carbonylation, DNA damage and ROS production in alloxan induced diabetic rats.

Oxidative stress performs an imperative role in the onset and progression of diabetes. Metabolic enzymes and cellular organelles are detrimental to increased levels of free radicals and the subsequent reduction in anti-oxidant defence. Pyridoxamine (vitamin B6) is an indispensible nutrient for humans and is considered to be an important food additive too. The aim of this research was to examine the effect of vitamin B6 in a diabetic environment. This study reports the effects of pyridoxamine supplementation in alloxan induced diabetic rats. Diabetes was induced by the single intra peritoneal dose of alloxan (120 mg per kg body weight). Diabetic rats were treated with pyridoxamine (10 and 15 mg per kg body weight) and compared with a control set of diabetic rats without supplementation. Pyridoxamine treatment showed dose dependent recovery in all parameters. A notable decline in oxidative stress parameters and ROS production with reductions in fasting blood glucose levels along with normal patterns of the glucose tolerance test has been reported here. Histological studies reveal damage recovery in the liver as well as kidney tissues. A notable amount of recovery was observed in cellular DNA distortion and damage. It is thus advocated that pyridoxamine might help in reducing problems associated with diabetes. A probable mechanism pertaining to the action of pyridoxamine is proposed as well.

A comparative insight into the oxidative damage and cell death potential of photoilluminated aminophylline - riboflavin system in normal and cancer lung cells of swiss albino mice.

Photosensitisation of riboflavin (Rf) activates aminophylline (Am) resulting into the formation of a highly pro-oxidant Am-Rf system. We have previously shown its macromolecular damaging response in human peripheral lymphocytes, however, its potential inside a cancer cell is yet to be explored. Since, altered redox status of a cancer cell is a reliable therapeutic window in designing anticancer strategies, therefore, it's imperative to investigate whether the reactive oxygen species (ROS) generated by this system readily triggers apoptosis or it is countered by elevated antioxidant machinery of a cancer cell. Here, we have demonstrated DNA damaging and cytotoxic potential of this system in benzopyrene induced lung carcinoma cells. Using various biochemical assays significant macromolecular damage was observed along with mitochondrial membrane disruption as evaluated by rhodamine 6G membrane permeant. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay showed decreased cell viability, confirming cytotoxic action whereas fluorescence and electron microscopic evaluation confirmed apoptosis. ROS scavengers ameliorated the oxidative damage and inhibited cell death, thus confirming, pivotal role of ROS in causing cell death. It was evidently found out that the lung cancer cells were more sensitive towards the photodynamic action of this system, which can be attributed to the upregulated riboflavin metabolism in cancer cell. Hence, we propose a photodynamic mechanism to kill lung cancer cell that exhibits enhanced sensitivity towards cancer cells.

Glyoxal Induced Transition of Transferrin to Aggregates: Spectroscopic, Microscopic and Molecular Docking Insight.

BACKGROUND & OBJECTIVE: The present study was aimed at characterizing the conformational alterations induced in human transferrin, the iron regulatory protein by glyoxal. Since protein aggregation is at the core of many disorders, thus interest in this domain has increased significantly during the past years. METHODS: In our present study, the effect of glyoxal was monitored on human transferrin using multispectroscopic and multi-microscopic studies. RESULTS: Intrinsic fluorescence spectroscopy suggested changes in native conformation of human transferrin evident by decreased fluorescence and blue shift in the presence of glyoxal. Further, extrinsic fluorescence was retorted and the results showed the formation of aggregates; apparent by increased Congo red (CR) absorbance, Thioflavin T (ThT) and ANS fluorescence and TEM of human transferrin in the presence of glyoxal. Molecular docking was also employed to see which residues are at core of human transferrin and glyoxal interaction. Reactive oxygen species (ROS) generation assays revealed enhanced ROS levels by human transferrin after treatment with glyoxal. CONCLUSION: Thus, our study proposes that glyoxal induces the formation of aggregates in human transferrin. These aggregates further generate ROS which are key players in the complications associated with diabetes mellitus, giving our study clinical perspective.

Chronic unpredictable environmental stress impair biochemical and physiological homeostasis: Role in diabetes mellitus.

AIMS: Chronic unpredictable environmental stress (CUES) may induce predisposition to diabetes mellitus. This study investigates the role of CUES on impaired homeostasis. MATERIAL AND METHODS: Stressed group mice (n = 20) were exposed to CUES for 16 weeks. Weekly body weight, feed consumption, feed efficiency ratio, fasting blood glucose were monitored. Plasma HbA1c, plasma cortisol, plasma epinephrine and plasma insulin, serum lipids, antioxidants and carbohydrate metabolizing enzymes activity were assessed along with DNA damage and histopathological examination of liver, kidney, pancreas, spleen and skeletal muscles. RESULTS AND CONCLUSION: s: Fasting blood glucose levels & HbA1c in the stressed were significantly higher compared to control (p < 0.001). Serum lipids were found insignificantly higher in stressed mice compared to control. Body weights of the stressed mice and feed efficiency ratio were found significant (p < 0.001). Plasma corticosterone, plasma epinephrine, HOMA-IR was found to be significantly higher in the stressed group (p < 0.001). Plasma insulin level was found to be significantly lower in the stressed group (p < 0.001). Significant changes were observed in antioxidants level, carbohydrate metabolizing enzymes activity, peripheral tissues and DNA integrity. CUES initiates pathogenesis of diabetes.

Probing the interaction of a coumarin-di(2-picolyl)amine hybrid drug-like molecular entity with human serum albumin: Multiple spectroscopic and molecular modeling techniques.

HSA is an important plasma protein responsible for transport of drug molecules. Coumarin derivatives play critical role as anticancer, antidiabetic and antiparkinson agents. In our lab we have synthesized coumarin-based pharmacophore, di(2-picolyl)amine-3(bromoacetyl) coumarin (ligand-L) endowed with anticancer activity. Anticancer agents binding mode of HSA provides valuable pharmacological information and is a structural guidance in synthesizing new drugs with greater efficacy. Thus, binding mechanism of ligand-L with HSA was explored using spectroscopic and molecular docking techniques. UV-Vis spectroscopy demonstrates hyperchromism in the absorbance spectra of HSA on addition of ligand-L suggesting interaction of ligand-L with HSA. Fluorescence spectroscopy indicates quenching in the fluorescence of HSA in the presence of ligand-L confirming the complex formation and this binding follows static mechanism. Steady state fluorescence spectroscopy revealed high binding affinity between ligand-L and HSA with a 1:1 stoichiometry. Thermodynamic parameters obtained by ITC suggest that the interaction between ligand-L and HSA is mainly driven by van der Waals forces and hydrogen bonds, and the negative value of ΔG is an indication of spontaneous binding process. Competitive binding and molecular docking experiments showed that the binding site of ligand-L mainly resides in sub-domain IIA of HSA. CD experiments revealed no significant conformational changes in the secondary structure of HSA on binding of ligand-L. We also found that esterase-like activity of HSA was not affected by ligand-L. In conclusion, this study demonstrates binding mechanism of ligand-L with HSA, and the binding did not induce conformational changes in HSA. This study is likely to provide better understanding of transport and delivery of ligand-L via HSA. Overall, it will provide insights into pharmacokinetic properties of ligand-L and designing new ligand-L based derivatives with greater efficacy.

Interaction of C20-substituted derivative of pregnenolone acetate with copper (II) leads to ROS generation, DNA cleavage and apoptosis in cervical cancer cells: Therapeutic potential of copper chelation for cancer treatment.

Cervical cancer is a leading cause of cancer-related deaths among women in developing countries. Therefore, development of new chemotherapeutic agents is required. Unlike normal cells, cancer cells contain elevated copper levels which play an integral role in angiogenesis. Thus, targeting copper via copper-specific chelators in cancer cells can serve as effective anticancer strategy. In this work, a copper chelator pregnenolone acetate nucleus-based tetrazole derivative (ligand-L) was synthesized and characterized by elemental analysis, ESI-MS, 1H NMR and 13C NMR. DNA binding ability of ligand-L was studied using UV-Vis and fluorescence spectroscopy. Fluorescence spectroscopy studies reveal that quenching constant of ligand-l-DNA and ligand-L-Cu(II) were found to be 7.4 × 103 M-1 and 8.8 × 103 M-1, respectively. In vitro toxicity of ligand-L was studied on human cervical cancer C33A cancer cells. Results showed that ligand-L exhibit significant cytotoxic activity against cervical cancer C33A cells with IC50 value 5.0 ±â€¯1.8 µM. Further, it was found that ligand-L cytotoxicity is due to redox cycling of copper to generate ROS which leads to DNA damage and apoptosis. In conclusion, this is the report where we synthesized pregnenolone acetate-based tetrazole derivative against C33A cells that targets cellular copper to induce pro-oxidant death in cancer cells. These findings will provide significant insights into the development of new chemical molecules with better copper chelating and pro-oxidant properties against cancer cells.

Interaction of aminophylline with photoilluminated riboflavin leads to ROS mediated macromolecular damage and cell death in benzopyrene induced mice lung carcinoma.

Riboflavin (Rf) or vitamin B2 is a known photosensitizer whose photophysical and photochemical properties are well established. Aminophylline (Am) is a phosphodiesterase inhibitor and is currently used as a bronchodilator. Although there are several reports of haemolytic and proteolytic interaction of photoilluminated riboflavin with aminophylline, the cytotoxicity of this system against malignant tissue is not well defined and fully unravelled. Here, we are evaluating anticancer activity of this system against B(a)P induced lung carcinoma in swiss albino mice. We observed marked increment in the level of cellular redox scavengers as well as oxidative stress markers. A significant DNA damage was observed using comet assay. Histopathological studies further confirmed induction of apoptosis in lung tissues of Am-Rf treated animals. Scanning electron microscopy revealed altered surface morphology of the malignant tissue, which characteristically improved in the treatment group. Since malignancy is characterised by compromised redox status, therefore, further increment in ROS due to the action of this system derives cellular system towards extensive macromolecular damage and consequent ROS mediated apoptosis. We anticipate the usage of this system in developing efficient photodynamic therapy against lung cancer that can be clinically realised.

ROS mediated antibacterial activity of photoilluminated riboflavin: A photodynamic mechanism against nosocomial infections.

Nosocomial infections are a major threat to modern therapeutics. The major causative agent of these infections is multidrug-resistant gram-negative bacteria, which impart high morbidity and mortality rate. This has led to an urge for the development of new antibiotics. Antimicrobial photodynamic therapy is a promising strategy to which till date no resistant strain has been reported. Since the efficacy of photodynamic therapy largely depends on the selection and administration of an appropriate photosensitizer, therefore, the realization of clinically active photosensitizers is an immediate need. Here, by using E. coli as a study model we have demonstrated the antimicrobial photodynamic potential of riboflavin. Intracellular ROS formation by DCFH-DA assay, lipid peroxidation, protein carbonylation, LDH activity was measured in treated bacterial samples. Enzymatic (SOD, CAT, GSH) antioxidants and non-enzymatic (GSH) was further evaluated. Bacterial death was confirmed by colony forming assay, optical microscopy and scanning electron microscopy. The treated bacterial cells exhibited abundant ROS generation and marked increment in the level of oxidative stress markers as well as significant reduction in LDH activity. Marked reduction in colony forming units was also observed. Optical microscopic and SEM images further confirmed the bacterial death. Thus, we can say that photoilluminated riboflavin renders the redox status of bacterial cells into a compromised state leading to significant membrane damage ultimately causing bacterial death. This study aims to add one more therapeutic dimension to photoilluminated riboflavin as it can be effectively employed in targeting bacterial biofilms occurring on hospital wares causing several serious medical conditions.