Journal of Traditional and Complementary Medicine xxx (2017) 1e7 Contents lists available at ScienceDirect Journal of Traditional and Complementary Medicine journal homepage: http://www.elsevier.com/locate/jtcme Original Article Phenolic constituents and modulatory effects of Raffia palm leaf (Raphia hookeri) extract on carbohydrate hydrolyzing enzymes linked to type-2 diabetes Felix A. Dada a, b, Sunday I. Oyeleye b, c, Opeyemi B. Ogunsuyi b, c, Tosin A. Olasehinde d, * Stephen A. Adefegha b, Ganiyu Oboh b, , Aline A. Boligon e a Science Laboratory Department (Biochemistry Unit), Federal Polytechnic, Ede, P.M.B 231, Ede, Nigeria b Functional Foods and Nutraceuticals Unit, Department of Biochemistry, Federal University of Technology, Akure, P.M.B. 704, Akure, 340001, Nigeria c Department of Biomedical Technology, Federal University of Technology, Akure, P.M.B. 704, Akure, 340001, Nigeria d Nutrition and Toxicology Division, Food Technology Department, Federal Institute of Industrial Research, Oshodi, PMB 21023, Lagos, 10001, Nigeria e Phytochemical Research Laboratory, Department of Industrial Pharmacy, Federal University of Santa Maria, Building 26, Room 1115, Santa Maria, 97105-900, RS, Brazil article info abstract Article history: This study sought to investigate the effects of Raffia palm (Raphia hookeri) leaf extract on enzymes linked Received 21 June 2016 to type-2 diabetes mellitus (T2DM) and pro-oxidant induced oxidative stress in rat pancreas. The extract Received in revised form was prepared and its a-amylase and a-glucosidase inhibitory effects were determined. Radical [2,2- 9 August 2016 þ þ diphenyl-1-picrylhydrazyl (DPPH)] scavenging and Fe2 -chelating abilities, and inhibition of Fe2 - Accepted 12 January 2017 induced lipid peroxidation in rat pancreas homogenate were assessed. Furthermore, total phenol and Available online xxx flavonoid contents, reducing property, and high performance liquid chromatography diode array de- tector (HPLC-DAD) fingerprint of the extract were also determined. Our results revealed that the extract Keywords: a ¼ m a ¼ m Raffia palm leaf inhibited -amylase (IC50 110.4 g/mL) and -glucosidase (IC50 99.96 g/mL) activities in concen- ¼ m Type-2 diabetes tration dependent manners which were lower to the effect of acarbose (amylase: IC50 18.30 g/mL; 2þ Antioxidant glucosidase: IC50 ¼ 20.31 mg/mL). The extract also scavenged DPPH radical, chelated Fe and inhibited 2þ Phenolic compounds Fe -induced lipid peroxidation in rat pancreas all in concentration dependent manners with IC50 values Anti-hyperglycemia of 402.9 mg/mL, 108.9 mg/mL and 367.0 mg/mL respectively. The total phenol and flavonoid contents were 39.73 mg GAE/g and 21.88 mg QAE/g respectively, while the reducing property was 25.62 mg AAE/g. The HPLC analysis revealed the presence of chlorogenic acid (4.17 mg/g) and rutin (5.11 mg/g) as the major phenolic compounds in the extract. Therefore, the ability of the extract to inhibit carbohydrate hydro- lyzing enzymes and protect against pancreatic oxidative damage may be an important mechanisms supporting its antidiabetic properties and could make Raffia palm leaf useful in complementary/alter- native therapy for management of T2DM. However, further studies such as in vivo should be carried out. © 2017 Center for Food and Biomolecules, National Taiwan University. Production and hosting by Elsevier Taiwan LLC. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/ licenses/by-nc-nd/4.0/). 1. Introduction insulin resistance, pancreatic b-cell dysfunction and hyperglyce- mia.1 The progression of T2DM has been reported to cause some Type-2 diabetes mellitus (T2DM) is a chronic metabolic disease metabolic complications such as ketoacidosis, hypertension, e and the most common form of diabetes. It is characterized by atherosclerosis, cardiovascular diseases, pancreatitis and so on.2 4 More so, previous reports have revealed that the development and progression of T2DM is associated with glucolipotoxicity and * Corresponding author. Functional Foods and Nutraceuticals Unit, Department of oxidative stress.5 Glucolipotoxicity which involves increased levels Biochemistry, Federal University of Technology, Akure, P.M.B. 704, Akure, 340001, of plasma glucose and fatty acids have been implicated in the Nigeria. impairment of pancreatic b-cell function.5,6 The b-cells play major E-mail address: [email protected] (G. Oboh). Peer review under responsibility of The Center for Food and Biomolecules, role in the secretion of insulin which regulates the absorption of National Taiwan University. glucose. However, at the early onset of T2DM, glucolipotoxicity http://dx.doi.org/10.1016/j.jtcme.2017.01.003 2225-4110/© 2017 Center for Food and Biomolecules, National Taiwan University. Production and hosting by Elsevier Taiwan LLC. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). Please cite this article in press as: Dada FA, et al., Phenolic constituents and modulatory effects of Raffia palm leaf (Raphia hookeri) extract on carbohydrate hydrolyzing enzymes linked to type-2 diabetes, Journal of Traditional and Complementary Medicine (2017), http://dx.doi.org/ 10.1016/j.jtcme.2017.01.003 2 F.A. Dada et al. / Journal of Traditional and Complementary Medicine xxx (2017) 1e7 induces mitochondria b-cell dysfunction, decreased glucose up- distilled. A JENWAY UVevisible spectrophotometer (Model 6305; take, induce reactive oxygen species (ROS) production and induce Jenway, Barlo world Scientific, Dunmow, United Kingdom) was hyperglycemia.6,7 Hyperglycemia has been reported as a major used to measure absorbance. factor in ROS production which can induce tissue damage and pathological conditions such as diabetic neuropathy, nephropathy 2.2. Determination of total phenol content and retinopathy.8 The use of synthetic drugs such as acarbose and miglitol in the reduction of blood glucose via the inhibition of ac- The total phenol content of the extract was determined ac- tivities of carbohydrate hydrolyzing enzymes (a-amylase and a- cording to the method of Singleton et al.18 Briefly, appropriate di- glucosidase) as therapeutic targets for the management of T2DM lutions of the extract were oxidized with 2.5 mL 10% has been widely reported.9 The inhibition of these enzymes delays FolineCiocalteau's reagent (v/v) and neutralized by 2.0 mL of 7.5% the rate of carbohydrate hydrolysis in the small intestine and sodium carbonate. The reaction mixture was incubated for 40 min consequently reduces the absorption of glucose concentration into at 45 C, and the absorbance was measured at 765 nm in the the blood stream, thereby repressing postprandial hyperglycemia spectrophotometer. The total phenol content was subsequently and preventing hyperglycemia-induced ROS.10 calculated and expressed as gallic acid equivalent (GAE). The use of natural compounds for the treatment of diabetes and its complications has gained lots of interests recently. This is due to 2.3. Determination of total flavonoid content the fact that the use of synthetic drugs causes several side effects such as flatulence, diarrhea, abdominal dilatation, and so on in the The total flavonoid content was determined using a slightly body and hence, the need for an alternative therapy.9 Previous re- modified method.19 Briefly, 0.5 mL of diluted extract was mixed ports have shown that some medicinal plants and plant foods have with 0.5 mL of absolute methanol, 50 mL of 10% AlCl3,50mLof1M blood glucose lowering abilities and could be essential for man- Potassium acetate and 1.4 mL of distilled water, and allowed to e agement of T2DM.11 13 Phenolic compounds from natural source incubate at room temperature for 30 min. The absorbance of the have been reported to be potent inhibitors of a-amylase and a- reaction mixture was subsequently measured at 415 nm; the total glucosidase activities with little or no side effects.8,14 The use of flavonoid content was subsequently calculated and expressed as medicinal plants for the management of T2DM could be more ad- quercetin equivalent (QAE). vantageous than synthetic drugs due to their ability to scavenge free radicals which is a major risk factor of diabetic complications, 2.4. High performance liquid chromatography analysis (HPLC-DAD) as antidiabetic drugs such as acarbose lack significant antioxidant effects.8 Raphia hookeri samples at a concentration of 12 mg/mL were Raffia palm [Raphia hookeri Mann and Wendl (Family: Palma- injected by means of a model SIL-20A Shimadzu Auto sampler. ceae)] is a monocotyledon plant, commonly found in West Africa Separations were carried out using Phenomenex C18 column and very abundant in lowlands and swampy areas in Southwest (4.6 mm  250 mm  5 mm particle size). The mobile phase was Nigeria where it grows in water to about 1 m deep.15 Raffia palm is a water with 1% formic acid (v/v) (solvent A) and HPLC grade very tall plant of about 16 m with a single stem, 1e4 suckers, large methanol (solvent B) at a flow rate of 0.6 mL/min and injection leaves and trunks which contains palm wine.15 The oil processed volume 40 mL. The composition gradient was: 5% solvent B reaching from this plant is used for cooking and making margarine while the 15% at 10 min; 30% solvent B at 25 min, 65% solvent B at 40 min and pulp is usually consumed with boiled cassava.16 Apart from its 98% solvent B at 45 min, followed by 50 min at isocratic elution culinary uses, the Raffia palm roots have a variety of medicinal uses. until 55 min. At 60 min the gradient reached the initial conditions In traditional medicine, the root extract is used as laxative and to again, following the method described by Oboh et al.13 with slight alleviate stomach pain in infants.17 Although, there are many re- modifications. The sample and mobile phase were filtered through ports on the antidiabetic properties of Raffia palm root and seed 0.45 mm membrane filter (Millipore) and then degassed by ultra- extracts11,16; however, to the best of our knowledge, there is little or sonic bath prior to use.