Acute and Long-Term Toxicity of Mango Leaves Extract in Mice and Rats

The acute toxicity of mango leaves extract (MLE) at the maximal dose (18.4 g/kg) was studied in ICR mice and no abnormalities were detected during the experiment. The long-term studies at various doses of MLE (100 mg/kg, 300 mg/kg, and 900 mg/kg) in SD rats for 3 consecutive months revealed that, compared with the control group, rats in MLE treated groups showed slight body weight increase and higher fat weight; the serum TG and CHOL levels and the epididymis weight of male rats were a little higher; the serum K+ level of female rats was on the low side but the weights of liver, kidney, and adrenal gland were on the high side. In addition to this, no other obvious abnormalities were detected.


Introduction
Mango tree (Mangifera indica L.), a tropical plant belonging to Anacardiaceae, has been distributed worldwide as the most cultivated fruits in the tropics. Mango leaves were used for diabetes and asthma treatment in traditional Chinese medicine (TCM). Mango leaves contain phenolic constituents such as caffeic acid [1], polyphenols such as mangiferin and gallic acid [2], flavonoids [3], volatile compounds [4], and so forth. Pharmacology studies showed that the extract of mango leaves possesses many effects like antioxidant, antimicrobial, antihelminthic, antidiabetic, antiallergic, and so forth [5]. In previous study, we reported benzophenone Cglycosides with triglyceride accumulation inhibitory effects in adipocyte [6,7]. We also reported that ethanol extract of mango leaves dose-dependently decreased serum glucose and triglyceride in KK-A y mice, and mechanism on glucose and lipid homeostasis is mediated, at least in part, through PI3K/AKT and AMPK signaling pathway [8]. Although mango tree leaves were used for a long period in TCM clinic, there are few reports on the safety evaluations. In this study, we carried out the acute toxicity and long-term toxicity of mango leaves extract (MLE), aiming at providing reference basis for other safety evaluation studies and selecting clinical dosage.  Ten mice in each group; no difference was considered to be significant between the two groups.   intragastric administration at the corresponding dose, in a volume of 0.1 mL per 10 g, for 3 consecutive months (6 times a week), while the control group received an equal volume of deionized water. During the study, all rats were allowed access to food and water ad libitum.

Materials and Methods
(2) Observational Indices. After oral administration, observe the general symptom, such as appearance, behavior, glandular secretion, breathing, and so on. The body weight and food consumption of each animal were recorded weekly and the differences among groups were compared. After 90 days of treatment, all the 48 SD rats were sacrificed, and blood samples were collected from the abdominal aorta for hematology and coagulation tests. The white blood corpuscles (WBC) count, red blood corpuscles (RBC) count, hemoglobin concentration (HGB), hematocrit (HCT), mean corpuscular volume (MCV), and prothrombin time (PT) were carried out.
Serum was separated by spinning the blood and was used for biochemical studies. ALT, AST, ALP, BUN, CREA, TP, ALB, GLU, CHOL, Na + , K + , and Cl − contents in serum and many other blood biochemical parameters were determined using the automatic biochemistry analyzer.
Organs (liver, heart, spleen, lung, kidney, brain, thymus, etc.) were collected from each sacrificed rat and weighed. The relative organ weights (organ/body weight ratio and organ/ brain weight ratio) were calculated and compared with the value of the control.

Statistical Analysis.
The intragroup difference of measurement data was detected with the -test. The data obtained were subjected to SPSS NPar Tests Mann-Whitney Test. Values were expressed as the mean ± standard error and were considered statistically significant at < 0.05.

General Observation.
During the course of the study, all the mice were healthy without any abnormal responses, and no distinct lesions were revealed anatomically.

Body Weight.
After the second oral administration of MLE, the body weight of the mice in both of the two groups decreased slightly than before the first dose, and the control Evidence-Based Complementary and Alternative Medicine 5 group decreased more, but there was no significant difference ( > 0.05). It is speculated that the body weight loss may result from the fasting between the two doses and the high drug concentration (the maximum dispensing concentration) that induced satiety may affect the short-time body weight change. Throughout the recovery period, the animal weight, both of the two groups, showed a general increase, but in several mice it decreased a day after the treatment (one in control group and six in MLE treated group); and after fourteenday treatment, five mice in MLE treated group showed a slight weight loss (less than 1 g). Speculated by the whole growth trend, we guess that the body weight change after oneday treatment, which was recovered three days later, may be associated with the drug's effect and the change after fourteen days was likely to be coursed by physiological fluctuations, but it produced no significant difference ( > 0.05) compared with control group ( Table 1). As a result, the effect of MLE on mice body weight was not obvious.

Gross Anatomy and Histopathological Examination.
After a fourteen-day recovery, all the mice were executed for gross anatomy check. Because there were no gross lesions on major organs, no histopathological examination was conducted.

General Observation.
After three consecutive months of oral administration, all the animals showed no marked abnormalities during the study.

Body Weight and Food
Consumption. The body weight of rats (presented in Table 2) in each group showed a steady increase trend, while MLE treated groups had a higher bodymass index than control group. However, there were no significant differences ( > 0.05), except in the female rats in low and medium dose groups at day 10 and in high dose group at days 21 and 49 ( < 0.05).