Galina Nikolova

The protective effects of herbal feed additives Withania somnifera, Silybum marianum, Centella asiatica, and silymarin (administered at feed levels of 4000 ppm, 5000 ppm, 4600 ppm, and 25,000 ppm, respectively) against the toxic effects of ochratoxin A (OTA) (administered at 2 ppm) were investigated in 48 New Zealand White rabbits (37 days old) over an 80-day experimental period. A decrease in body weight was seen in rabbits exposed to OTA alone, but that decrease was less pronounced in rabbits receiving herbal supplements. The most severe lesions in OTA-treated rabbits were found in the liver, kidneys and spleen, while milder lesions were seen in the heart, intestine and lungs. The intensity of macroscopic, histopathological and biochemical changes was highest in rabbits exposed to OTA alone, followed by rabbits additionally supplemented with herbal additives, as evidenced by histopathological findings and serum levels of blood urea nitrogen (BUN), creatinine, aspartate aminotransferase (AST), and alanine aminotransferase (ALT). The most pronounced protective effects of the herbal additives were observed in the kidneys and liver. Oxidative stress was significantly increased in rabbits exposed to OTA alone, as evidenced by elevated reactive oxygen species (ROS), nitric oxide (●NO), ascorbyl radicals (●Asc), protein oxidation (PO), procollagen type I alpha 1 (COL1A1), malondialdehyde (MDA), kidney injury molecule-1 (KIM-1), hydroxyproline (Hyp), advanced glycation end products (AEGs), heme oxygenase-1 (HO-1), catalase (CAT), and 8-hydroxy-2′-deoxyguanosine (8-OHdG), as well decreased superoxide dismutase (SOD) and glutathione peroxidase-1 (GPx-1) activities, while protective effects were seen for all herbal additives, and better expressed for Centella asiatica and Silybum marianum.

Background/Objectives: The aim of the present study was to conduct a systematic in vitro assessment of the biofunctionalities of newly synthesized lignin (LNPs) and lignin–chitosan nanoparticles (LCNPs) via a comparative in vitro estimation of their cytotoxicity, cytocompatability potential, radical-scavenging activity, and antimicrobial performance, thereby establishing a benchmark for their sustainable design and biomedical applications. Methods: LNPs and LCNPs were synthesized via “green” self-assembly and co-assembly methods. Results: In vitro cytotoxicity studies on L929 fibroblasts and HaCaT keratinocytes demonstrated higher long-term viability for LCNPs (half-maximal inhibitory concentration IC₅₀ = 3.05 mg/mL at 72 h) compared with LNPs (IC₅₀ = 1.37 mg/mL), while both formulations maintained >76% viability at a concentration of 0.5 mg/mL. Electron Paramagnetic Resonance (EPR) and spectrophotometric antioxidant assays displayed strong radical scavenging activity, with LNPs excelling in ^●OH, NO, and ABTS scavenging and LCNPs exhibiting enhanced lipid peroxidation and superoxide inhibition potential. Antimicrobial testing revealed minimal inhibitory concentration (MIC) reductions of the nanoparticles up to 8–13-fold compared to lignin solutions, with LCNPs showing higher activity against Gram-positive and Gram-negative microbial strains. Conclusions: These results highlight LCNPs as biocompatible, antioxidant, and antimicrobial nanoplatforms with potential for regenerative medicine, oxidative stress mitigation, and infection control.

Donkey milk has been increasingly studied in recent years and has been proposed to be a functional food. However, its components undergo changes during lactation, including its lipid profile and redox-related properties. This study analyzed the fatty acid composition, antioxidant parameters, and redox-modulating properties of donkey milk from the Bulgarian local donkey breed at three lactation stages (0–30, 31–60, and 61–90 days postpartum). Milk samples from 40 clinically healthy donkeys were grouped by days postpartum. A cross-sectional design with three lactation stage groups was used; one-way ANOVA tested group differences with Tukey’s post hoc test, and associations with days postpartum were evaluated using regression models. Fatty acid methyl esters were analyzed by GC-FID, and the atherogenic (AI) and thrombogenic (TI) indices were calculated. Antioxidant enzymes (SOD, CAT, and GPx-1), GSH, MDA, TAC, and EPR-based redox markers (DPPH, Asc•, ROS, NO•, TEMPOL, and 5-MSL) were analyzed. During lactation, monounsaturated fatty acids decreased (approximately 32% in the first month to ~30% by the third month), while AI increased from ~1.9 to ~2.2, and TI increased to ~2.5. SOD and GPx-1 activities increased with advancing lactation, while total antioxidant capacity decreased (213.4 to 199.7 µmol). DPPH radical scavenging activity remained stable during lactation. EPR-detected ROS and NO• values increased with advancing lactation stage, while thiol-bound 5-MSL decreased, suggesting a shift in the balance between oxidative challenge and antioxidant defense during lactation. Regression modeling confirmed a significant effect of lactation period on multiple compositional and redox-related parameters. Therefore, the stage of lactation should be taken into account when interpreting the biological value, redox stability, and potential functional properties of milk, as well as when developing milk management and yield strategies.

Background/Objectives: Acetaminophen (paracetamol or APAP) overdose is a major cause of acute liver injury mediated by oxidative stress, inflammation, and hepatocellular necrosis. The present study investigates the in vivo hepatoprotective potential of morin (M), lignin nanoparticles (LN), and morin-encapsulated lignin nanoparticles (LMN) against APAP-induced hepatotoxicity in mice. The specific goal was to determine whether LMN could strengthen hepatic antioxidant and anti-inflammatory defenses prior to toxic insult, which aligns with a prophylactic model rather than a post-injury clinical rescue approach. This study was guided by the primary hypothesis that LMN pretreatment would markedly reduce APAP-induced hepatic injury. Methods: Experimental groups included control, APAP, M, LN, LMN, M+APAP, LN+APAP, and LMN+APAP treatments. Serum hepatic biomarkers, oxidative stress parameters, and inflammatory cytokines were analyzed to assess protective responses. Results: APAP exposure markedly elevated aspartate aminotransferase (AST) and alkaline phosphatase (ALP) levels, indicating severe hepatic dysfunction, accompanied by increased lipid peroxidation and pro-inflammatory cytokine production. LMN+APAP treatment significantly restored hepatic enzyme levels to approximately normal values and suppressed malondialdehyde (MDA) formation, while enhancing superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) activities. LMN also downregulated interleukin 6 (IL-6), tumor necrosis factor α (TNF-α), and interleukin 1β (IL-1β), while upregulating interleukin 10 (IL-10), suggesting effective attenuation of inflammatory signaling. Correlation analyses demonstrated positive interactions between MDA, cytokines, and hepatic enzymes, whereas antioxidant enzyme levels were inversely correlated with liver injury markers. Histopathological analysis revealed that treatment with LMN enhanced hepatoprotection, demonstrating predominantly mild, reversible lesions and suggesting a synergistic antioxidant and immunomodulatory effect. Conclusions: It could be concluded that LMN provided superior hepatoprotection compared to M or LN. These findings establish LMN as a promising bio-based nanotherapeutic agent for mitigating drug-induced hepatotoxicity through coordinated antioxidant and anti-inflammatory mechanisms.

Purpose: This study aimed to evaluate the radical scavenging capacity and potential modulation of oxidative stress exerted by Rosa alba L. absolute in a model of gentamicin-induced nephrotoxicity. Material/Methods: The antioxidant activity of R. alba L. absolute was examined using electron paramagnetic resonance (EPR) spectroscopy. In vivo EPR measurements were performed to assess scavenging activity against hydroxyl (•OH) and superoxide (•O2-) radicals, as well as the DPPH radical scavenging capacity (DPPH•). Tissue homogenates were prepared from renal samples and serum obtained from Balb/c mice. Animals were randomly allocated into four experimental groups (n = 6): (a) the control group, receiving an intraperitoneal injection (i.p.) of 1 mL isotonic NaCl solution (0.9%) daily along with a standard diet; (b) the R. alba group, administered R. alba absolute orally at 80 mg·kg-¹ body weight·day-¹; (c) the gentamicin (GM) group, injected i.p. with GM at 200 mg·kg-¹ body weight·day-¹ to induce acute nephrotoxicity; and (d) the combination group, receiving GM (200 mg·kg-¹) followed 2 hours later by R. alba absolute (80 mg·kg-¹·day-¹) administered orally. EPR spectroscopy was further employed to analyze oxidative alterations and conformational modifications in renal proteins, with a particular focus on sulfhydryl (-SH) group integrity in albumin. Protein carbonylation (PC) levels were quantified using an enzyme-linked immunosorbent assay (ELISA). Results: The findings demonstrated that R. alba L. absolute exhibited pronounced antioxidant activity, effectively scavenging hydroxyl and superoxide radicals and displaying notable DPPH radical neutralization capacity. Conclusions: Moreover, oral administration of R. alba significantly attenuated protein oxidation and carbonylation, thereby mitigating oxidative damage to renal proteins (p < 0.05).

Background: Goat milk and its fermented products exhibit unique nutritional and therapeutic characteristics corresponding to the management of metabolic disorders, such as type 2 diabetes mellitus (T2DM) and its associated complications, including diabetic nephropathy (DN). Due to its rich content of bioactive compounds and superior digestibility compared to cow’s milk, goat milk enhances nutrient assimilation and exhibits notable anti-inflammatory and antioxidant effects. The primary aim of this investigation was to systematically evaluate the efficacy of goat milk-based nutritional interventions as an integral component of a multifaceted therapeutic approach aimed at attenuating oxidative stress (OS), restoring metabolic homeostasis, and mitigating the progression of long-term complications in patients with diabetes mellitus and concurrent renal dysfunction. Methods: Participants diagnosed with T2DM were stratified into three subgroups based on the severity of renal dysfunction. The results were analyzed in comparison with those of healthy control subjects. Results: Following a dietary regimen that included goat milk enriched with Aronia (Aronia melanocarpa) fruit juice patients—particularly those with DN—exhibited marked reductions in free radical concentrations, decreased cytokine production, and diminished levels of lipid and protein oxidation byproducts. Moreover, a significant improvement was observed in nitric oxide (NO) levels, along with partial restoration of the nitric oxide synthase (NOS) system and an upregulation of endogenous antioxidant enzyme activity (p < 0.05) relative to pre-intervention measurements. Conclusions: These outcomes suggest that the dietary intervention not only attenuated OS but also contributed to improved renal function in affected individuals. The results support the therapeutic potential of functional dairy-based diets—specifically those incorporating bioactive ingredients such as Aronia melanocarpa fruit juice and goat milk—in mitigating oxidative damage and enhancing metabolic and renal health in patients with T2DM and DN.

Objectives: The current study focused on the kidney protection and antioxidant properties along with the potential anti-ferroptotic activity of Geranium macrorrhizum L. (G. macrorrhizum) oil to ameliorate the acute renal oxidative tissue damage and toxicity of the aminoglycoside antibiotic gentamicin (GM) in an experimental murine model. Methods: The research was carried out with mature Balb/c mice distributed into four groups (n = 6). Application of GM (200 mg kg⁻¹ intraperitoneal injection for 10 days) was performed to induce kidney injury. Only saline was administered to the controls. The remaining groups were administered G. macrorrhizum oil (50 mg kg⁻¹ per dose) either used alone or in combination with GM. To assess the renal antioxidant status, the activities of specific antioxidant enzymes, indicators of lipid and DNA peroxidation and renal functional damage were examined using standard commercial kits, ELISA and EPR spectroscopy. Results: G. macrorrhizum oil analysis revealed 20 organic components belonging to mono- and sesquiterpenoids and long-chain hydrocarbons. The antioxidant and anti-inflammatory effects of G. macrorrhizum oil were demonstrated by reduced malondialdehyde, ROS, 8-hydroxy-2′-deoxyguanosine and cytokine levels (especially interleukin-1β) compared with GM. Furthermore, increased activation of superoxide dismutase, catalase and glutathione (GSH) were observed in the kidney homogenates of the animals which received GM in combination with G. macrorrhizum oil compared with the GM group. Additional changes in the GSH/glutathione peroxidase-4 axis were detected, suggesting the possible anti-ferroptotic potential of the oil. Nephroprotection was also demonstrated by elevated PGC-1α expression (peroxisome proliferator-activated receptor γ coactivator 1-alpha) and reduced KIM-1 levels (kidney injury molecule-1) following application of the oil. Conclusions: The preserved kidney antioxidant and functional properties in the groups treated with oil suggest that Geranium macrorrhizum L. could be utilized clinically to mitigate the toxic effects of GM application.

This study investigates the association between diabetic retinopathy (DR) and its complication, diabetic macular edema (DME), and compared it with biomarkers of oxidative stress. This study aimed to compare the main indicators of the development of diabetic retinopathy measured as parameters of oxidative stress and compared to lipid oxidation, DNA damage, and cytokine levels and to monitor their quantitative manifestation in DME. This study evaluated 134 patients (62 males and 72 females; aged 62.10 ± 11.22 years) and divided them into two groups: type 2 diabetes mellitus with DR and type 2 diabetes mellitus with DME. All results were compared with healthy volunteers (n = 94) and showed that patients with DME had significantly higher levels of ROS, cytokine production, lipid oxidation, and DNA damage. In addition, patients with DME had decreased levels of nitric oxide (NO) and an impaired NO synthase (NOS) system (p < 0.05). These findings suggest that patients with DR and DME are unable to compensate for high levels of oxidative stress. Reduced NO levels in patients with DME may be due to impaired NO availability. This study highlights compromised oxidative status as a contributing factor to DME in patients with decompensated type 2 diabetes mellitus. An assessment of oxidative stress levels and inflammatory biomarkers may aid in the early detection and prediction of diabetic complications.

Oxidative stress is a critical factor in the onset of gestational diabetes and its associated complication, pre-eclampsia. This study aimed to evaluate (1) reactive oxygen species, reactive nitrogen species, and superoxide radical levels as indicators of oxidative stress, (2) lipid and protein oxidation, (3) antioxidant enzyme activity, and (4) cytokine production in pregnant women with gestational diabetes, as well as those with both gestational diabetes and pre-eclampsia, comparing these with biomarkers of gestational diabetes mellitus. The study categorized pregnant patients with gestational diabetes mellitus into two groups based on the presence of new-onset hypertension, measured twice every four hours, and a 24 h urine protein test showing 300 mg/day or ≥1+ proteinuria detected via a visual dipstick at ≥20 weeks of gestation. These groups were compared with normotensive pregnant patients. The findings revealed that patients with both gestational diabetes and pre-eclampsia exhibited significantly elevated levels of reactive oxygen species, cytokine production, and lipid and protein oxidation end products compared to normotensive pregnant women. Additionally, these patients showed reduced nitric oxide (•NO) levels, impaired NO synthase systems (eNOS and iNOS), and decreased antioxidant enzyme activities (p < 0.05). These results indicate that patients with gestational diabetes and pre-eclampsia are unable to counteract oxidative stress effectively. The study underscores the compromised oxidative status as a contributing factor to these complications. The findings provide insights into the pathogenesis of gestational diabetes and the subsequent pre-eclampsia, the role of oxidative stress, and the resulting complications. Measuring oxidative stress levels and inflammatory biomarkers could help in the early detection and prediction of gestational-diabetes-related complications in pregnant women.

The production of soybean meal (SBM) can be linked to various issues related to the environment (e.g., deforestation, water waste, and transportation costs), and reducing its inclusion in pig diets by using alternative protein sources, such as insect meal, is an important challenge for nutritionists. This study aimed to compare the productive performance, dietary digestible energy (DE), nutrient digestibility, and some blood indices of growing Danube White pigs fed graded levels of Black Soldier fly (Hermetia illucens L.) larvae meal (BSFLM) at 0, 30, 60, 90, and 120 g/kg of diets, in replacement of SBM for 38 days, from 119 to 157 d old. Each diet was fed to eight pigs in individual boxes following randomization. Pigs grew according to breeders’ recommendations and did not have any clinical health problems. Replacing SBM did not change (p > 0.05) the pigs’ growth performance and DE, as only dietary fat digestibility increased in a linear fashion (p < 0.001), possibly due to the high BSFLM, i.e., the high-fat inclusion rate. There was a simultaneous rise in some oxidative damage indicators and an increase in antioxidant status, thus suggesting that further research involving longer feeding periods is needed to identify a potential time sequence of events. Overall, BSFLM is a promising ingredient in pig nutrition.