Hematological and biochemical evaluation of Ameliorative effects of mangiferin on deltamethrin induced toxicity in rats

Introduction

Deltamethrin is a type II synthetic pyrethroid insecticide extensively used for crop protection and vector control. Although considered relatively safe at recommended doses, prolonged or excessive exposure has been reported to cause oxidative stress, hepatic damage, renal impairment and haematological alterations in experimental animals (Mongi et al., 2011) [10] . Toxic effects are primarily mediated through free radical generation and membrane damage. Haematological parameters serve as important indicators of toxic insult and physiological stress. Alterations in haemoglobin concentration, erythrocyte count, leukocyte profile and packed cell volume reflect systemic toxicity (Ncir et al., 2020) [11] . Serum biochemical markers such as liver enzymes, total protein, urea and creatinine are sensitive indicators of hepatic and renal function (Manna et al., 2006) [8] . Mangiferin is a naturally occurring xanthone glycoside found in Mangifera indica. It possesses antioxidant, anti-inflammatory and hepatoprotective properties. Previous experimental studies have demonstrated its ability to scavenge free radicals and protect against chemical-induced organ toxicity. The present study was undertaken to evaluate the haematological and biochemical alterations induced by deltamethrin in rats and to assess the protective efficacy of mangiferin against such toxicity.

Female Wistar rats aged 6-8 weeks, non-pregnant and nulliparous, were used for the experiment. The animals were maintained under controlled environmental conditions with a temperature of 22±3 °C, relative humidity of 50-60%, and a 12-hour light/12-hour dark cycle. Standard laboratory feed and water were provided ad libitum throughout the study period. A total of 40 rats were randomly allocated into five groups, with eight animals in each group. Group I (normal control) received Peanut oil orally. Group II (positive control) was administered deltamethrin where deltamethrin of pure form was dissolved in Peanut oil where Peanut oil was used as a vehicle, at a dose of 5.2 mg/kg body weight (1/10th of LD50 dose). Group III received mangiferin alone at a dose of 30 mg/kg body weight. Group IV was treated with deltamethrin (5.2 mg/kg body weight) in combination with mangiferin (30 mg/kg body weight). Group V received deltamethrin (5.2 mg/kg body weight) along with a higher dose of mangiferin (60 mg/kg body weight).

Body weight Measurement

The body weight of all the experimental animals were recorded weekly once and the dosing was adjusted accordingly.

Hematology

The blood samples collected in EDTA vials on 14th and 28th day of the experiment were subjected for haemoglobin (Hb), total erythrocyte count (TEC), total leucocyte count (TLC), total platelet count, packed cell volume (PCV) Mean Corpuscular Volume (MCV), Mean Corpuscular Haemoglobin (MCH) and Mean Corpuscular Haemoglobin Concentration (MCHC) estimation using Mindray BC-2800VET auto Haematology auto analyser in the Department of Teaching Veterinary Clinical Complex, Veterinary College, Hassan.

Serum Biochemistry

For serum biochemical parameters analysis, blood was collected on 14th and 28th day into serum vacutainers, centrifuged at 3000 rpm for 15 min and serum separated and stored at-20 °C till further analysis. Biochemical estimation for AST, ALT, ALP, Urea, BUN and Creatinine levels were analysed using standard laboratory procedure by Robonik, India Semi-automatic Biochemistry Analyzer (Prietest Compact) using reagent kits from Lifechem, Ahmedabad.

Results

General observations

Oral administration of deltamethrin to rats for 28 days did not result in mortality. However, animals in Group II showed noticeable clinical signs, including roughened hair coats and reduced feed intake compared with Group I. During the last two weeks of treatment, Group II rats exhibited anorexia, lethargy, weakness, abnormal gait, aggressive behavior, and dizziness. In contrast, rats in Groups III, IV, and V displayed normal behavior and activity throughout the experimental period. Body weights were recorded weekly and are presented in Table 1. The rate of body weight gain was significantly influenced by deltamethrin administration, both alone and in combination with mangiferin. From the second week onward, the mean body weight of rats in Group II was significantly lower compared with Groups I, III, IV, and V.

Conversely, Group III animals showed a progressive increase in body weight throughout the study period.

Absolute organ weights

A significant increase in absolute liver weight was observed in Group II compared with Group I. In Groups IV and V, liver weights were significantly reduced relative to Group II, while liver weights in Group III were comparable to those of Group I. Similarly, the absolute kidney weight was significantly higher in Group II than in Group I. In Groups IV and V, kidney weights were significantly lower compared with Group II, whereas a marginal increase was noted in Group III relative to Group I.

Relative organ weights

The relative weights of the liver and kidneys were significantly increased in Group II compared with Group I. In contrast, Groups IV and V showed a significant reduction in relative liver and kidney weights when compared with Group II. No significant differences in relative liver or kidney weights were observed between Group III and Group I.

Hematological parameter

The mean values of hematological parameters with standard error of mean on 14 th and 28th day are presented in Table 2. On days 14 and 28, Group II showed a significant decrease in total erythrocyte count (TEC) compared to Groups I, IV, and V. In contrast, TEC levels were significantly higher in Groups IV and V when compared with Group II. TEC levels in Group III were similar to those in Group I. During the same periods, total leukocyte count (TLC) was significantly increased in Group II compared to Group I. However, Groups IV and V showed a significant decrease in TLC levels compared to Group II. No significant difference in TLC levels was observed between Group III and Group I. Hemoglobin (Hb) levels on days 14 and 28 were significantly lower in Group II compared to Group I. In contrast, Hb levels were significantly increased in Groups IV and V relative to Group II. Group III showed Hb levels comparable to Group I.

Serum Biochemistry

The mean values of Serum Biochemistry parameter values with standard error of mean on 14th and 28th day are presented in Table 3. Serum ALT, AST and ALP levels were significantly elevated in Group II compared to Group I. In contrast, Groups IV and V showed a significant reduction in these enzyme levels relative to Group II. Enzyme levels in Group III remained comparable to those in Group I. Renal function markers, including urea, blood urea nitrogen (BUN), and creatinine, were significantly increased in Group II compared to Group I. However, Groups IV and V exhibited a significant decrease in these parameters when compared with Group II. No significant differences were observed between Group III and Group I.

Discussion

Body weight gain is an important indicator of growth. In the present study, rats treated with deltamethrin alone (Group II) showed a significant reduction in both absolute and percentage body weight gain after 28 days compared to the control group (Group I). Although Group III showed an increase in body weight gain, the difference was not statistically significant compared to Group I. These findings are consistent with earlier reports showing reduced body weight gain following deltamethrin exposure (Gerbed et al., 2014; Ncir et al., 2018) [6,11] . The reduction in body weight may be due to the metabolic and hormonal effects of deltamethrin (Manna et al., 2005) [7] . Similar effects on body weight have also been reported in fipronil-induced toxicity in rats (Mohan, 2023) [9] . Relative organ weight is a key indicator of organ toxicity. In this study, a significant increase in relative liver weight was observed in Group II compared to Group I, which may be attributed to inflammatory changes and liver enlargement caused by deltamethrin exposure, as previously reported (Sharma et al., 2014) [18] . Similarly, Group II showed a significant increase in relative kidney weight compared to Group I. This increase may result from inflammatory cell accumulation and altered renal function, consistent with earlier findings ( [16,12,7] . Hematological analysis is commonly used to assess disease conditions and toxic effects caused by chemicals and pesticides (Al-Attar et al., 2010) [3] . Erythrocytes play a vital role in oxygen transport and hemoglobin levels are closely associated with total erythrocyte count (Pasini et al., 2010) [15] . In the present study, Group II showed a significant decrease in TEC and Hb levels, along with an increase in TLC, compared to Group I. These findings are consistent with earlier reports on deltamethrin toxicity (Mongi et al., 2011; Aljali et al., 2023) [10,4] and similar hematological alterations reported in fipronil-induced toxicity (Mohan, 2023) [9] . The reduction in hemoglobin may indicate regenerative anemia due to interference with erythropoiesis, while decreased TEC and increased TLC could be attributed to oxidative stress-induced damage to red blood cells (Mongi et al., 2011; Aljali et al., 2023) [10,4] . Similar protective effects of mangiferin on these parameters have also been reported by Mohan (2023) [9] supporting the present findings. A significant increase in AST, ALT, ALP, creatinine, urea and BUN levels was observed in Group II compared to Group I. These findings are consistent with previous reports showing deltamethrin-induced liver and kidney toxicity in rodents (Abdel et al., 2016) [1] . Elevated ALT and AST levels indicate liver cell damage, which may be associated with bile duct hyperplasia, hepatic congestion, Kupffer cell hyperplasia and altered membrane permeability. Increased ALP levels, which are present on hepatocyte membranes and bile duct epithelium, serve as a sensitive indicator of toxic liver damage (Tewari et al., 2014) [21] . The rise in urea levels observed in this study supports the findings of (Tewari et al., 2014) [21] and correlates with kidney tissue damage. Elevated BUN and creatinine levels further indicate impaired renal function, likely due to reduced glomerular filtration and tubular dysfunction (Mongi et al., 2011; Aljali et al., 2023) [10,4] . Meanwhile, Mangiferin is a natural xanthone compound known for its antioxidant, anti-inflammatory, and protective properties. It reduces toxicity by scavenging free radicals and strengthening antioxidant defense mechanisms (Shashirekha et al., 2015; Ajay et al., 2023) [19,2] . Based on these properties, this study evaluated the protective effect of mangiferin against sub-acute deltamethrin-induced toxicity in Wistar rats. Groups IV and V showed a significant increase in absolute and percentage body weight gain compared to Group II, indicating that mangiferin reduced deltamethrin-induced oxidative stress and neurotoxicity. Group III showed values comparable to Group I. Similar protective effects of mangiferin on body weight were reported by Song et al. (2020) [20] . Relative liver and kidney weights were significantly reduced in Groups IV and V compared to Group II, suggesting hepato-and nephroprotective effects of mangiferin. This may be due to inhibition of the NF-κB inflammatory pathway and reduced reactive oxygen species formation. These findings are consistent with earlier reports on the protective role of mangiferin (Song et al., 2020) [20] . Serum biochemical markers, including ALT, AST, ALP, urea, creatinine, and BUN, were significantly lowered in Groups IV and V compared to Group II, supporting the protective effects of mangiferin on liver and kidney function (Mongi et al., 2011) [10] . Hematological parameters such as TEC, Hb, and TLC were also significantly improved in Groups IV and V relative to Group II. These results agree with previous studies showing the hemoprotective and immunoprotective effects of mangiferin, likely through enhanced erythropoiesis and reduced oxidative damage to blood cells (Das et al., 2011; Sellamuthu et al., 2014; Mohan, 2023) [5,17,9] .

Conclusions

Sub-acute oral administration of deltamethrin in rats produced marked toxic effects, including decreased body weight gain, changes in liver and kidney weights, hematological alterations (reduced TEC and hemoglobin with elevated TLC), and increased hepatic and renal biomarkers, indicating liver and kidney toxicity. Exposure to deltamethrin significantly elevated serum ALT, AST, ALP, urea, creatinine, and BUN levels, along with increased relative liver and kidney weights, demonstrating functional impairment and structural damage to these organs. Concurrent treatment with mangiferin notably mitigated deltamethrin-induced toxicity by improving body weight gain, normalizing organ weight indices, restoring hematological parameters, and lowering elevated liver and kidney biochemical markers. The protective action of mangiferin is likely attributed to its antioxidant and anti-inflammatory properties, which reduce oxidative stress and suppress inflammatory responses, thereby maintaining hepatic, renal, and hematological health during deltamethrin exposure.

Acknowledgement

AcknowledgementI would like to convey my heartfelt appreciation to the Veterinary College, Hassan, for providing me with the opportunity to undertake my M.V.Sc. research work.

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