Hepato-renal toxicity of Myristica fragrans Houtt. (Myristicaceae) seed extracts in rats

1 Introduction

Foods derived from plants have been widely acclaimed as sources of nutrients needed to promote good health and wellbeing (Hirschi, 2009). Although many plants are rich in nutritional factors and are medicinal, which are beneficial in combating nutritional deficiency and managing common health challenges, their abuse could be harmful. Considering that natural source of nutritional factors for nutrient-fortification is perceived to be safer and maybe more acceptable by the consumers, using common spices for nutritional fortification of food and beverages need to be investigated. However, there is a need to investigate the safety of using a high dose of these spices over a long time to avoid creating problems while trying to solve another. The need to confirm the safety of this plant is because a handful of plants are toxic, majorly when consumed at higher doses (Mounanga et al., 2015), including the common spice, Myristica fragrans Houtt. (Myristicaceae) (Benzeid et al., 2018).

Myristica fragrans is traditionally used in managing paralysis because it increases blood circulation, scavenge radical species, and kill cancer cells (Ginting et al., 2021). Other properties include aphrodisiac, hypolipidaemic, anti-thrombotic, anti-fungal, nervous stimulant, aromatic, anti-dysenteric and anti-inflammatory, fungicidal (Fernando and Senevirathne, 2020; Zhao et al., 2020), anti-diarrheal, and memory-enhancing (Nikolic et al., 2021) potentials have been documented. On the other hand, it has been shown to induce convulsions, palpitations, and hallucinations, indicating that it elicits psychoactive effects (Gupta, 2020). Essential oils from the seed formulated as potential drugs in combination with magnesium aluminometasilicate as an excipient are currently being examined for approval for broad-spectrum therapeutic purposes (Matulyte et al., 2020). Recently, the seeds were demonstrated to contain substantial amounts of essential amino and fatty acids and moderate levels of essential minerals (Anaduaka et al., 2020). Considering the nutritive and pharmacological properties of M. fragrans, the seeds have potential application in the biofortification of food to mitigate malnutrition. However, this can only be possible if the plant is demonstrated to be safe for consumption in large quantities over a long time. The involvement of the liver and kidney in detoxification processes exposes them to damage by cytotoxic chemicals. Hence, monitoring of liver and kidney status is crucial in establishing food safety. This study, therefore, seeks to evaluate the phytochemical constituents and effects of n-hexane and methanol extracts of M. fragrans on the liver and kidney status of rats.

2 Materials and methods

3 Results

4 Discussion

The increased understanding of herbs and spices as sources of natural antioxidants in addition to nutrients has encouraged more researches on them with a view of identifying those with little or no toxicity (Chatterjee et al., 2007). The seeds of M. fragrans are widely used as spices and drugs by locals, some in large amounts without knowing the potential safety concerns associated with the consumption of large doses of the plant. This study investigated the impact of short-term administration of large doses of extracts of M. fragrans on two crucial organs, the liver and kidney. The presence of flavonoids, terpenoids, alkaloids, phenols, steroids, and tannins in both extracts of M. fragrans (though in varying quantities) agrees with earlier studies. Nikolic et al. (2021) reported the presence of alkaloids, flavonoids, cardiac glycosides, and saponins in M. fragrans seed essential oil. Similar to our findings, the presence of terpenoids and phenolics was also reported by El-Alfy et al. (2009) and Naikodi et al. (2011) in their separate studies. The existence of these secondary metabolites in the plant extracts suggests that the extracts may elicit some health-promoting potentials (Huang et al. 2010). It was shown that the extracts did not cause mortality or conspicuous sign of toxicity. The observed tolerance of mice to the extracts in this study supports an earlier report by George and Ioana (2007) that myristic acid, a constituent of M. fragrans (Anaduaka et al., 2020), elicits a low order acute oral toxicity in rodents. This finding implies that the extracts have a low acute toxicity profile, making the seed relatively safe for consumption at low doses over a short time.

Urea is produced as a waste product of protein digestion by the liver and is typically removed from the body through ultra-filtration in the glomeruli of the kidneys (Kasper et al., 2005). Similarly, creatinine is a metabolite of the muscle’s creatine phosphate whose concentration is relatively constant due to the kidney’s role in extracting it (Miller et al., 2005). Hence, urea and creatinine levels are essential indicators of renal health because higher than expected serum urea, and creatinine levels suggest impaired kidney function. The significantly higher urea and creatinine levels after 14 days daily administration of the extracts, as seen in groups 3 and 5 compared to the control, suggest that the extracts at 1000 mg/kg b.w. for 14 days elicits nephrotoxicity.

One of the liver function enzyme markers, ALP, participates in membrane transport, protein and glycogen metabolisms in addition to secretory properties (Akpanabiatu et al., 2005). Hence, elevated circulating ALP levels may arise from leakage of the enzymes from the hepatic membrane in hepatotoxic conditions (Malomo et al., 2006). The increase in the activity of aminotransferases is generally produced by cellular necrosis (Etim et al., 2008). The elevated levels of ALT and AST in some of the treated groups on days 7 and 14 treatment could increase the metabolism of toxic metabolites and enhance the production of free radicals, thereby causing hepatocellular injury. In all the liver marker enzymes activities assessed, there was a general elevation in their activities in serum, and the activities increased on day 14 compared with day 7, suggesting that M. fragrans seed extracts exhibit dose and duration of consumption-dependent hepatotoxicity, with n-hexane extract causing being more toxic.

After 14 days of oral ingestion, the extracts did not alter lactate dehydrogenase (LDH) activity in serum. This indication of no toxicity could be attributed to the presence of myristicine that has been shown to prevent lipopolysaccharides-induced liver injury (Morita et al., 2003). As a marker for liver and cardiac injuries, the presence of a large amount of LDH in circulation indicates severe damage to cardiac and hepatic tissues (King and Perry, 2001). LDH activity increased after 14 days of consuming n-hexane extract, indicating that high doses of this extract caused severe liver damage. Luster et al. (2000) reported that hepatotoxins' first point of assault is the centrilobular regions that harbor CYP 450 mixed-function oxidases. This group of enzymes catalyzes the detoxification of toxic metabolites with the release of free radicals as by-products. Among these groups of enzymes, CYP 2E1 is mainly found in the liver with a smaller amount in extra-hepatic tissues, including lungs, kidney, brain, endothelium of large blood vessels, heart, bone marrow, and nasopharyngeal tissue (Kang et al., 2008). CYP 2E1 is responsible for the breakdown of many low molecular weight toxins and carcinogens (Malhotra, 1998). Methanol and n-hexane extracts of M. fragrans seeds at 500 and 1000 mg/kg b.w. for 14 days caused elevation (though not statistically significant) in liver CYP 450-2E1 compared with control. Yokota et al. (1988) earlier demonstrated that 1-terpinen-4-ol and eugenol increases phase II detoxifying enzymes' activities for xenobiotics in the liver, such as UDP-glucuronyl transferase, DT-diaphorase, and glutathione-S-transferase without significantly increasing CYP 450. A higher-than-expected level of bilirubin in serum is a good marker of hepatic malfunction (Eteng et al., 2009). Hence, the ability of the extracts to elevate total bilirubin at 1000 mg/kg after 14 days of oral consumption suggests hepatotoxicity. The elevated bilirubin level in this study might have resulted from a reduction in the liver’s ability to metabolize and conjugate them due to compromise, such as obstruction of bile ducts. It could also be due to a reduction in levels of reducing equivalents such as NADPH and GSH. Reduced glutathione contributes to protecting the membrane of erythrocytes from hemolysis, decreasing bilirubin levels (Noori et al., 2009). The result of the histological examination shows indications of hepatotoxicity only at 1000 mg/kg. This indication of hepatotoxicity agrees with the observed degeneration of cells of the kidney, heart, and stomach characterized by cellular necrosis involving disruption and distortion of cellular membranes structural and functional integrity by M. fragrans seeds and its component, myristin (Eweka and Eweka, 2010; AL-Mosaibih, 2015; Anaduaka et al., 2020). Generally, despite the numerous health benefits of the spice, M. fragrans, consumers are advised to consume only a small quantity over a short duration to avert toxicological effects associated with chronic consumption of a high dose of the spice.

5 Conclusions

The presence of substantial quantities of phytochemicals including terpenoids, flavonoids, alkaloids, phenols, steroids, and tannins were detected in methanol and n-hexane of Myristica fragrans seeds, although at varying concentrations. This study also demonstrated that at high doses, the extracts induced hepatotoxicity and renotoxicity, with a higher impact on the liver. Compared to the methanol extract, the n-hexane extract was shown to be more toxic. Based on the findings of this study, consumption of a large quantity of M. fragrans seed over a long duration is discouraged. Further study is needed to ascertain the molecular mechanism behind the observed hepato-renal toxicity induced by the seed extracts.