Overview
Tylosema fassoglense (TFG) is used as an antidiarrheal traditional medicine in Western Kenya. This study aimed to investigate the antidiarrheal activity of its aqueous extracts in-vivo and the putative mechanism (s) of action ex-vivo using Sprague-Dawley rats and New Zealand white rabbits respectively. The in-vivo antidiarrheal effects of the extract were evaluated in castor oil-induced diarrhea, the castor oil-induced enteropooling, and phenol red gastric motility tests. On the other hand, isolated rabbit’s jejunal segments were used to evaluate the spasmolytic effect of TFG on spontaneous contraction, in acetylcholine-induced contraction, in presence of 80mMK+, calcium chloride-induced contraction as well as in presence of the following antagonists: naloxone, methylene blue, L-NAME, prazosin, and propranolol in the ex-vivo studies. The data were express as Mean ± S.E.M and analyzed by one-way ANOVA and Tukey’s post hoc test in cases of significance which was set at p < 0.05. The extract was phytochemically characterized using Liquid chromatography Mass spectroscopy (LC-MS).The extract possessed significant inhibitory effect in the in-vivo experiments. The extract exhibited significant spasmolytic effect on both spontaneous contraction and in jejunal segment pre-contracted acetylcholine as well as in presence of 80mMK+ solution. It also attenuated the spasmogenic effect of various concentration of calcium chloride. The extract’s spasmolytic effect was, however, significantly attenuated in presence of several antagonists (methylene blue and L-NAME) but the adrenergic blockers (prazosin and propranolol) had no significant effect in the ex-vivo studies. LC-MS identified thirty compounds where Proathocyanidin (11.54%), Syringic acid (7.30%), and 4-Hydroxybenzoic acid (6.19%) had the highest percentage abundance. In conclusion, the results obtained in this study partially validate the traditional uses of the tubers of this plant species as an antidiarrheal. These antidiarrheal effects are probably mediated via modulation of nitrous oxide pathway, voltage gated calcium channels, and muscarinic receptors.
Sponser
Washika Amos Mapesa*, 
Mwangi Peter Waweru, Frederick Bukachi and Kayaja David Wafula
Principal Investigator
Abstract
Diarrhea is a gastrointestinal disorder characterized by an increase in frequency of bowel movement of at least three times or more per day (Levine et al., 2017), increased liquidity of stool (Shane et al., 2017), and an increase in amount of stool of at least 200 g/day (Levine et al., 2017). Diarrheal diseases lower the quality of life, prolong hospitalization stays, raise healthcare costs and are the leading cause of death in children aged below five years (DuPont, 1995; Kotloff et al., 2017). These health impacts are attributed to the link between diarrhea and pre-renal acute kidney injury (Seifter and Chang, 2017) , malnutrition in patients (Guerrant et al., 1992) , and deterioration of health especially when comorbid with chronic diseases (World Health Organization, 2019). Indeed, diarrheal diseases account for one in nine deaths of children under the age of 5 (Centers for Disease Control and Prevention, 2015). The death rate is eleven-fold higher for children with comorbid conditions such as HIV (Centers for Disease Control and Prevention, 2015). Developing countries inordinately bear a huge global burden of diarrheal diseases (Merson, 1981; Kotloff et al., 2017).
Diarrhea is classified into five interrelated categories: secretory (Harrell and Cheng, 2018), malabsorptive (Hammer et al., 1990; Walters and Pattni, 2010), osmotic (Binder, 2010), motility, and congenital (Na+ or Cl/HCO−3) disorder diarrhea (Kere et al., 1999; Janecke et al., 2016). Secretory diarrhea is caused by toxins of infectious agents such as Vibrio cholerae which possess inhibitory effect on GTPase enzyme. As such, there is continuous activation of cAMP-adenylyl cyclase pathway, affecting gating of cystic fibrotic transmembrane regulator (CFTR) ion channel leading to hypersecretion of chloride ions into gut’s lumen and consequently water via osmosis (Harrell and Cheng, 2018). Digestive enzymes deficiency, inflammatory bowel diseases or absence of certain membrane transporters result in malabsorptive diarrhea (Surawicz, 2010). Lactase deficiency, for instance, cause the transition of the undigested lactose to the colon where microbiota convert it anaerobically into osmotically active short fatty acid anions (SCFAs) that mediate osmotic diarrhea (Binder, 2010). Similarly, unabsorbed bile salts due to the absence of apical bile salt transporters (ABST) as seen in terminal ileum resection, cause diarrhea via intestinal membrane irritation and increased mucus production (Barkun et al., 2013). Motility disorder diarrhea is mediated via neurohumoral effect where elevation of neurotransmitters or hormones such as secretin produced by tumors of enterochromaffin cells (carcinoid syndrome), shorten intestinal transit time via serotoninergic signaling (Ohevon der Ohe et al., 1993). The absence of brush boarder sodium-proton exchanger isoform (NHE3) in congenital Na+ disorder, causes sodium and accompanying copious fluid loss that manifest with hyponatremia (Surawicz, 2010).
Current treatment modalities of diarrhea aim at restoring fluid volume and electrolyte balance using various approaches i.e. the use of antibiotics in bacterial infectious diarrhea, bile acid sequestrant for patients with bile salts malabsorption (Porges, 1958), fecal microbiota transplant for patients with gut microbiota dysbiosis (Niccum et al., 2018) , and rotavirus vaccines for children under the age of 5 years who are susceptible to rotavirus infection (Dennehy, 2008). The use of these approaches is limited by numerous adverse effect and financial constraints. As such, there has been an increase in adoption of alternative medicine approaches in the management of diarrheal diseases (Patel et al., 2013). Some of the antidiarrheal herbal drugs in use include but not limited to Idigofera spicata Forssk(Fabaceae) (Awe et al., 2011), Pyrenacantha staudtii (Engl.) Engl. (Icacinaceae) (Birru et al., 2016), and Alpinia oxyphylla Miq. (Zingiberaceae) (Wang et al., 2015). The increase in the adoption of herbal medicine is attributed to their availability, affordability, efficacy, and reduced side effects (Patel et al., 2013). The tubers of Tylosema fassoglense (Kotschy ex Schweinf.) Torre and Hill (Fabaceae.) (http://www.theplantlist.org/) is a traditional antidiarrheal herbal remedy in Western Kenya (Maundu et al., 1999).
