Thesis
Phytochemical and biological investigation of Lathyrus linofilius as a possible treatment in diabesity
- Creator
- Rights statement
- Awarding institution
- University of Strathclyde
- Date of award
- 2019
- Thesis identifier
- T15483
- Person Identifier (Local)
- 201552203
- Qualification Level
- Qualification Name
- Department, School or Faculty
- Abstract
- Diabetes is a world-wide issue which can affect people at any age. It is ranked as oneof the top 10 diseases responsible for death worldwide. According to the InternationalDiabetes Federation, 424.9 million people worldwide were estimated to have diabetesin 2017 (90% with type 2 diabetes) and this number is projected to reach 629 millionpatients by 2045. The current treatments for managing diabetes are linked with anumber of side effects including severe hypoglycaemia, permanent neurologicaldeficit, stomach-ache, headache, lactic acidosis, liver damage, dizziness and death insome cases. Therefore, there is a need for improved anti-diabetic drugs with fewer sideeffects to enhance patient compliance and to control blood glucose levels more tightly.Natural products and plants in particular, offer an alternative to syntheticdrugs. Lathyrus linifolius is a plant whose tubers have historically been used as anappetite suppressant during medieval times in times when food was scarce. Due to theclose link between appetite suppressants and the treatment of diabetes/obesity, theplant’s leaves and tubers were examined for their potential anti-diabetic and antiobesity activity.This project aimed to: 1) produce Lathyrus linifolius extracts using Soxhlet apparatusextraction from which bioactive compounds could be isolated by columnchromatography and characterised by nuclear magnetic resonance (NMR); 2) assessthe anti-diabetic and anti-obesity activity of the extracts and compounds; 3) investigatethe anti-inflammatory and anti-oxidant activity of the extracts and compounds in invitro assays; 4) identify the effects of the tuber treatment on the gene expression ofappropriate diabesity components in normal rat pancreatic tissues obtained from aprevious study in which rats were fed with tubers; and 5) re-assess Lathyrus linifoliuseffects on obese Zucker rats and monitor the effects on body weight, food intake, waterintake and blood glucose levels.1) Four compounds were isolated for the first time from the ethyl acetate extract of thetubers; these were betulinic acid, lupeol, stigmasterol and β-sitosterol. The isolatedcompounds are known for their beneficial effects on hyperglycaemia and obesity tosome extent. Betulinic acid was the major component in the tubers and leaves.2) In biological assays, the ethyl acetate extract and betulinic acid from the tubers werepotent α-glucosidase inhibitors (P<0.05) with IC50 of 9.5 µg/ml and 5.5 µM,respectively. Moreover, the leaf ethyl acetate and methanol extracts were also stronginhibitors of α-glucosidase (P<0.05) with IC50 of 0.58 µg/ml and 4.3 µg/ml,respectively. In an anti-obesity assay, based on pancreatic lipase, the ethyl acetatetuber extract at 30 µg/ml and betulinic acid at 30 µM showed 50% enzymeinhibition (P<0.05); and the hexane extract from the leaves showed 30% inhibition. InHepG2 cells, leaf hexane extract showed a significant increase in glucose uptake whichwas comparable to that of insulin. These findings showed promising results for theplant to be used as an anti-diabetic and anti-obesity agent. Inflammation and oxidativestress are implicated in diabesity and determined the subsequent investigation.3) Moreover, all the plant extracts and isolated compounds at 30 µg/ml or 30 µMshowed a greater than 70% protection (P<0.05) in L929 cells (mouse fibroblastscommonly used for anti-inflammatory studies) from the cytotoxic effects of tumournecrosis factor alpha (10 µM). This was followed by investigating the ability ofLathyrus linifolius to inhibit nuclear factor kappa-light-chain-enhancer of activated Bcells (NF-ƙB) in the NCTC cell line (human skin cells transfected with a NF-ƙBluciferase reporter vector, used in anti-inflammatory studies). Betulinic acid, ethylacetate extract of the tubers, hexane and ethyl acetate extracts of the leaves inhibitedNF-ƙB which gave a NF-ƙB % inhibition of 70, 40, 30 and 30, respectively. Abetulinic acid inhibition curve was performed and found to inhibit NF-ƙB expressionwith an IC50 of 22.8 µM. Oxidative stress was then investigated following the positivefindings in the anti-inflammatory assays. Lathyrus linifolius extracts and isolatedcompounds were not reactive oxygen species-generators and significantly (P<0.05)protected cells from reactive oxygen species generated by tert-butyl hydroperoxide inSH-SY5Y (neuronal), HepG2 (hepatic) and Panc1 (pancreatic) cells, except lupeol inHepG2 cells. In HepG2 cells, glutathione levels were significantly (P<0.05) increasedby ethyl acetate extract of the tubers which confirmed the potential anti-oxidantactivity of Lathyrus linifolius.4) It was considered pertinent to carry out a retrospective investigation into any genechanges using RNA-Sequencing and real-time quantitative polymerase chain reaction(RT-qPCR) on RNA isolated from rat pancreatic tissues in a feeding study withLathyrus linifolius tubers carried out by Woods (2017). It was found that theadiponectin gene (Adipoq) was highly up-regulated (P<0.05) whereas genes thatcontribute to pancreatic β-cells damage for example Il1β and Tnfrsf19 were downregulated (P<0.05).5) Then, the biological effects of Lathyrus linifolius on an animal model for diabesitywere followed up. Dried, powdered Lathyrus linifolius tubers (100-200 mg/kg BW)were dissolved in deionised water, mixed with Farley’s rusks and given to obeseZucker rats daily for 16 days. However, the results showed that there were no effectson the body weight, food intake, water intake or blood glucose of Zucker rats and thismay have occurred due to the low Lathyrus linifolius dosage used and the shortduration of the study. It was decided not to carry out any further investigation (such asRNA- Sequencing) after this as there was no significant difference in results betweenthe obese Zucker rats and normal rats as observed in the previous feed study.These findings suggest that compounds from Lathyrus linifolius could possibly be usedas anti-diabetic and anti-obesity agents in the future, based on the positive resultsobtained from the in vitro and ex vivo assays in this study. This study should befollowed by assessing the effects of Lathyrus linifolius on body weight, food intake,water intake and blood glucose in humans.
- Advisor / supervisor
- Ferro, Valerie A.
- Resource Type
- Note
- Previously held under moratorium from 10 February 2020 until 10 February 2022
- DOI
- Date Created
- 2019
- Former identifier
- 9912791092802996
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