Herbal remedies are rapidly gaining popularity throughout the world, especially in developing countries. It is frequently used in primary healthcare setups for the treatment of various diseases and disorders because they thought that herbal medicine is safe and free from any type of side effects [17]. Although therapies involving herbal products are established most of them have not been studied well, and their usage is either not monitored at all or is monitored very poorly. As a result, there is insufficient understanding of their mechanism of action, potential side effects, contraindications, and interactions with currently available modern medications. Hence, a toxicity study of every product is necessary to establish the safety profile of every herbal preparation before use as efficacy [4]. Kong et al described that the cytoprotective effect of the plant extracts is highly dependent on their concentration, bioavailability, and together with complex interaction among the phytochemicals [18]. Our results demonstrated that both FMB and conventional extract are safe.
The modulation of cellular immune response by plant extract is an interesting area for inflammation and immunity. Hence, investigating the effects of plant extracts that modulate pinocytes and splenocytes response gives evidence to study immunomodulation and anti-inflammatory [19]. In our study, we cultured mouse peritoneal macrophages in RPMI medium with or without supplementation of external stimuli and, after 48 hours of incubation, used neutral red, which is readily taken up by macrophages to measure the pinocytic activity. After the incubation of macrophages with neutral red, macrophages were lysed to measure the concentration of neutral red and determine the extent of pinocytic activity by the macrophages. Macrophages are the first-line defence against the antigen, and pinocytic activity is one of the distinguished methods to determine macrophage activation. Phagocytosis cells (neutrophils, monocytes and macrophages) are key components of the innate immune response. However, Macrophages play a dominant role in phagocytosis which carried out the intracellular killing of antigens and other apoptotic cells. So, the modulation of macrophage activity seems to be the foremost role in the regulation of innate immunity [20]. The spleen is the body's largest immune organ, capable of producing a large number of lymphocytes. Splenocyte proliferation is a crucial event in the activation cascade of both cellular and humoral immune responses [21]. Our in vitro studies showed comparable spleen proliferation with the previously reported study [22].
Zinc is a crucial trace element with anti-inflammatory and antiviral effects that helps in immunity. Zn deficiency has been noted to result in increased susceptibility to infectious diseases [23]. It is also noticeable that calcium plays calcium seems to play a central role in the activation of cells of the immune system. T lymphocytes, which play a central role in cell-mediated immunity, require a sustained calcium ion flow for regulation, survival, activation and proliferation [24]. Iron and immunity are very tightly related to one another. Iron promotes the growth and pathogenicity of pathogens and is necessary for the activation and expansion of immune cells [25]. ETEs are generally found in very little amounts in soil, plants, and living things, but they work as enzyme catalysts in the body's systems to strengthen defences and minimise inflammations. Additionally, ETEs enhance the pharmacokinetic profile and cellular uptake of the bioactive chemicals by enhancing their mobility and solubility
TNF-α plays a key function among the many inflammatory cytokines in causing tissue-specific inflammation, which can erode the body's immune system. According to earlier research, a number of anti-inflammatory compounds have been shown to reduce tissue damage while directly controlling the development, function, and activation of the immune system's key components [26]. According to reports, ETEs boost the levels of anti-inflammatory cytokines in the body's inflammatory response, stabilising the immunological defence system and having strong antioxidant potential. Zinc supplementation has been demonstrated to partially correct zinc deficiency-induced oxidative stress and inflammatory TNF-α in rat models [27]. Similar to calcium, iron is a necessary element for numerous bodily functions, including erythropoiesis, immunological response, and host defence, as well as vital cellular processes including DNA replication and repair, as well as enzymatic reactions that need iron as a cofactor [25]. On the other side, problems with iron, homeostasis, or inflammation ultimately result in chronic disease. Additionally, it has been claimed that taking calcium and vitamin D supplements lowers inflammation and bacterial dysbiosis [12].