Diabetes Mellitus (DM) is the commonest serious, complex and multifarious group of metabolic disorders of multiple etiology that disturbs carbohydrate, fat and protein metabolism, characterized by chronic hyperglycemia as a result of relative or absolute lack of insulin or mounting resistance to its action [1] . Chronic hyperglycemia gives rise to the risk of microvascular (retinopathy, nephropathy and neuropathy) and macro vascular (ischaemic heart disease, stroke and peripheral vascular disease) damage with attendant reduction in life expectancy and diminished quality of life [2] . The world prevalence of diabetes among the adult population aged 20 years to 79 years in 2010 was 6.4% affecting 285 million adults and expected to rise to 7.7% and 439 million adults by the year 2030 [3] . A report by the International Diabetes Federation said that more than 371 million people (8.3%) of the world’s population had diabetes in 2012 [4] . This dramatic increase is noted to be more in developing countries particularly in Sub-Saharan Africa and Asia [5] as a result of on-going quest in adopting “western life style” and diet. The currently available orthodox medicines to control hyperglycemia in DM management include: “insulins, insulin secretagogues (sulfonylureas, meglitinides), insulin sensitizers (biguanides, thiazolidinedione), agents that enhance incretin secretion and action (incretin analogues, incretin mimetics, dipeptidyl peptidase IV (DPP-IV) inhibitors), agents that decrease gastrointestinal glucose absorption (alpha glucosidase inhibitors, alpha amylase inhibitors, sodium-glucose co-transporter (SGLT-1) selective inhibitors), agents that promote renal glucose excretion (sodium-glucose co-transporter (SGLT-2) inhibitors) and others (amylin analogue, bile acid sequesterants, bromocriptine) [6] -[8] . Despite the knowledge of the pathological processes involved in causation/progression of the disease and the wide range of therapeutic agents designed to fight hyperglycemia, the statistical projections are still alarming and the stability of communities is being threatened. Alternative strategies to the current pharmacological options of DM management are therefore urgently needed [9] to manage this crippling global health problem. The plant kingdom has become a target for the search of biologically active lead compounds for complementary/alternative management of diabetes mellitus. Medicinal plants are noted to play an important role in the management of diabetes. The effect of these plants may delay the development of diabetic complications and correct the metabolic abnormalities [10] . Similarly, it has been reported that some bioactive drugs isolated from plants showed antidiabetic activity with more efficacy than hypoglycemic agents used clinically [11] . Ethnobotanical and ethnopharmacological surveys report that more than 1200 plants are being used in many ethnic societies around the world in traditional medicine for their alleged hypoglycemic activity [12] -[15] and Newbouldia laevis (Bignoniaceae) is one of such plants.

The morphology, habitat and cultivation of Newbouldia laevis have been described [16] -[18] . In some parts of Southeastern Nigeria, it is usually planted at the grave to mark the position of the head of the deceased. This plant popularly known as boundary tree, chieftaincy tree, fertility plant or tree of life is locally called “Ogirisi” in Igbo, “Akoko” in Yoruba and “Aduruku” in Hausa languages [19] . The leaves, stem bark, roots and root bark of N. laevis have been reported to have versatile applications and are used in more than 25 medical purposes throughout the tropical Africa [17] , including but not limited in Nigeria, Togo, Senegal, Ghana, Congo, Cote de Voire, and Cameroun. Some of the documented medical uses include in the folk treatment of fevers (including yellow fever), malaria, stomach ache, cough, sexually transmitted infections, skin infections, tooth ache, breast cancer, constipation, pain (pelvic pain in females, chest pain, ear ache), gonococcal orchitis, elephantiasis, sorefeet, ulcer, epilepsy, convulsion, migraine, sickle cell anaemia, as a febrifuge, as a vermifuge, in female reproductive healthcare (fibroids, infertility, hemorrhage), as aphrodisiacs, eye problems, snake bites, wound healing, diabetes, arthritis, rheumatism and other inflammatory conditions [20] -[28] .

Pharmacological studies on extracts of different parts of N. laevis have revealed the antioxidant [29] , free radical scavenging [30] , antimicrobial [31] [32] , antimalarial [33] , sedative and anticonvulsant [34] -[36] , analgesic, antinociceptive and antiinflamatory [37] [38] , hepatoprotective [29] , anticancer [39] , uterine contraction [40] , wound healing and antiulcer [41] , antisickling [42] , hypoglycemic [43] , antihypertensive [44] [45] , entomocide [46] activities among others. Scientific reports on the phytochemical constituents of different plants of the plant revealed the presence of alkaloids, phenylpropaniod glycosides, flavonoids, tanins, saponins, phenols, essential oils, terpenoids, triterpenoids, quinoids, ceramides among others [29] [47] -[49] .

In Southeastern and Midwestern Nigeria, a hydro-alcoholic decoction of N. laevis leaves has been used in folk medicine for the management of diabetes mellitus. Previously, there are preliminary reports on the antihyperglycaemic and antidiabetic studies which were on the crude extract of the leaves [4] [43] [50] [51] . In this study, we evaluated the crude leaf extract of N. laevis and its solvent fractions for antihyperglycemic activity in alloxan diabetic rats (acute study) in order to explore the fraction(s) that has/have the active principle(s) responsible for the antihyperglycemic activity.

Hyperglycemia is the defining feature of diabetes mellitus. Chronic hyperglycemia is associated with both microvascular and macrovascular complications in patients with diabetes [2] , both of which are responsible for increased morbidity and mortality in diabetic subjects. Chronic hyperglycemia is directly and proportionately related to glycosylation of haemoglobin and lipid peroxidation, both of which are markers that herald the onset of diabetic complications [60] . Evidence has shown that tight and optimal blood glucose control eliminates diabetic complications [61] . The global incidence of the disease is geometrically rising to pandemic proportions. The bane of diabetes management has been to keep both short-term and long-term blood glucose levels optimized within acceptable limits in order to prevent or reduce these complications. This can only be achieved by tightly controlling fasting blood glucose levels as well preventing post prandial glucose excursions [62] . The later has been recognised lately as a better marker of glycaemic control than fasting blood glucose since glycation of haemoglobin is primarily favoured by post prandial hyperglycaemia [63] . Plasma glycated haemoglobin (HbA1c) level is an index of glycaemic control [60] .

Many of the available orthodox hypoglycaemic medications have not effectively addressed this global health problem. The reasons include affordability, accessibility, side-effects, acceptability, use restrictions in certain populations, and limitation of the agents in addressing underlying biochemical and pathological aberrations associated with the disease among many other reasons. Phytomedicines as an alternative have proven to be relatively safer, cheaper, more available and acceptable among the cultural society where they are found. Phytotherapy have demonstrated at least in animal models to prevent and/or halt the disease at different points of its natural history [64] [65] .

Prior to evaluating the effectiveness of antidiabetic plants, World Health Organization (WHO) advocates that their safety be evaluated in order to standardize their use. Marles and Farnsworth (1994) [66] earlier noted that about one third of medicinal plants used in traditional ethnomedical treatment of diabetes mellitus are toxic. Therefore, the oral acute toxicity of the dichloromethane-methanol (1:1) leaf extract of Newbouldia laevis was evaluated and found to be greater than 5000 mg/kg body weight. This finding agrees with previous reports on the LD₅₀ of the leaf extract [43] . Toxicological assessment and safety of various parts of N. laevis has been reported [29] [49] [67] . Their findings suggest a high safety profile of N. laevis extracts and could be suitable for use in management of chronic diseases such as diabetes mellitus.

Alloxan is known to induce free radical generation of which the pancreatic beta cells are particularly vulnerable [68] leading to massive destruction of insulin secreting cells and subsequent hyperglycemia. In the present study, it was observed and demonstrated that oral administration of extract and fractions of N. laevis leaf exhibited effective and dose dependent reduction in blood glucose levels of alloxan diabetic rats and could have possibly reversed the effect of alloxan in causing and maintaining hyperglycemia in animal studies. Previous reports on antihyperglycemic effects of plant extracts on alloxan diabetic animal models favour regeneration of islet beta cells as the primary mechanism of recovery of alloxan injected animals [68] . The reported antioxidant and free radical scavenging properties of N. laevis extracts [30] [45] [69] [70] may have contributed to the recovery of the pancreatic beta islet cells and restoration of its secretory function. Methanol fraction slightly lowered the fasting blood glucose level of normoglycemic rats suggesting weak insulin secretagogue activity in nonglycemic state. The methanol fraction however, demonstrated a significant improvement in oral glucose tolerance in glucose fed hyperglycaemic non-diabetic rats which was comparable to that of the standard drug glibenclamide. This may suggest glucose dependent insulinotropic and insulinomimetic properties of the methanol fraction [10] . Inhibition of intestinal sodium-glucose co-transporter (SGLT 1) and/or inhibition of renal glucose reabsorption (SGLT 2) may contribute to the observed effects [71] .

Although this study was not designed to investigate the antihyperglycemic mechanism of action of the extract and fractions in hyperglycemic non-diabetic and alloxan diabetic rats, a number of possible mechanisms (based on phytochemistry result) that may augment glucose disposal could help to explain the result of this study. Several phytoconstituents such as alkaloids, steroids, carbohydrates, glycosides, flavonoids, terpenoids, saponins, tannins, resins, peptides and amino acids, lipids, phenolics, glycopeptides and iridoids with putative antihyperglycemic activity have been reported [72] [73] . The preliminary phytochemical analysis of sub-fractions f₄ (showing highest activity of 74.57% reduction) showed high levels of alkaloids, flavonoids, saponins, resins and tannins. This suggests that the observed antihyperglycemic effects of the extract and fractions could have been mediated by one or more of these phytoconstituents.

Flavonoids have been shown to possess remarkable hypoglycaemic effects which have been linked to their capacity to avoid glucose absorption or improve glucose tolerance [74] . It has been demonstrated that flavonoids act as insulin secretagogues or insulin mimetics, attenuate diabetic complications, stimulate glucose uptake in peripheral tissues and regulate the activity and/or expression of the rate limiting enzymes involved in carbohydrate metabolism [75] -[77] . Flavonoids have been reported to show potent inhibitory activity against a wide range of enzymes such as lipo-oxygenases, cyclo-oxygenases, and prevent the generation or the action of free radicals which cause tissue damage during inflammatory processes [65] [78] [79] . They are therefore potent antioxidants and could modulate the activities of various enzymes involved in blood glucose homeostasis. This may have been responsible for the observed reduction of the blood glucose level by the extract and fractions of N. laevis leaf in alloxan-induced diabetic rats. Recently, there were similar reports on the antioxidant activities of the ethanol leaf extract of N. laevis and its ability to attenuate haemoglobin glycosylation and lipid peroxidation [51] . These reported antidiabetic potentials of N. laevis leaf extracts could only be explained wholly or in part by the result of this study since hyperglycemia favours these enzymatically induced changes in diabetic situations. Polyphenolic compounds have been implicated in the production of incretin hormone (glucagon-like peptide GLP-1) [65] that appears to act through many mechanisms towards effective glucose disposal, including stimulation of insulin secretion, suppression of glucagon release, slowing of gastric emptying, improving insulin sensitivity, and reduction of food intake. In rodents and cell line experiments, GLP-1 has been shown to promote β-cell regeneration, proliferation, mass, and function [65] . There is documented evidence that polyphenols slow down glucose absorption by competitive inhibition of the intestinal brush border membrane sodium-glucose co-transporter (SGLT 1) [80] as well as inhibition of disacharidases [81] . Kolawole and Abanji (2013) [82] reported the inhibition of alpha glucosidase and alpha amylase enzymes by the ethanolic leaf extract of N. laevis on sucrose fed rats. Phenolic compounds are known to directly cause regeneration of alloxan damaged islet beta cells by their action as antioxidants and free radical scavengers causing restoration of pancreatic beta cell function [83] .

Phlorizin-like effects of plant extracts have also been reported [84] . Phlorizin a non-selective, competitive inhibitor of sodium-glucose co transporter (SGLT-1 & SGLT-2), is a naturally occurring phenol glycoside, first isolated from the bark of an apple tree in 1835 [85] used to enhance glucose excretion through the kidneys. Modification of chemical structure of this prototype gave more selective and more potent SGLT-2 inhibitors, most of which are C-glycosides [85] . Studies have shown that oral administration of sergiflozin, a SGLT-2 inhibitor in normal and streptozotocin induced diabetic rats, increased urinary excretion in a dose dependent manner and attenuated the increase in blood glucose level following an OGTT without stimulating insulin release [86] . Similarly, dapagliflozin, a potent SGLT-2 inhibitor caused a dose dependent reduction in blood glucose level by as much as 60% in STZ-induced diabetic rats over a 5-hr period following oral administration [87] . These findings are consistent with the findings of this study. Obute and Adubor [88] reported that the flavonoid compound in N. laevis leaves is majorly quercetin which occurs naturally as glycosides [89] . Quercetin glycoside was noted to be a potential substrate for lactose phlorizin hydrolase (LPH) [89] and a similar pharmacological activity with phlorizin was suggested. Interaction of quercetin glycosides with intestinal SGLT-1 has also been reported [89] [90] . Flavonoids (quercetin glycosides), saponins (triterpenoid/ steroidal glycosides) present in the extract and fractions may have acted in the above manner to cause the observed effects.

The saponin present in the fraction may have played a role to explain the improvement in the oral glucose tolerance of normoglycemic rats pretreated with methanol fraction of N. laevis leaf extract in this study. In other studies, saponins have been shown to possess antihyperglycemic activities or other beneficial activities relevant in the management of diabetes. For examples, charantin, a steroidal glycoside saponin from Momordica charantia is known to possess insulin-like effects, stimulates insulin release and exhibits rat lens aldose reductase inhibitory activity [91] . Similarly, saponin isolated from Gymnema sylvestre (Gymnemic acid) exhibited marked hypoglycemic effects in animal models showing stimulation of insulin secretion and release, regeneration of pancreatic beta cells and activation of glucose metabolising enzymes [92] .

Alkaloids from plant extracts are reported to show antihyperglycemic activity through a number of mechanisms that regulate glucose homeostasis. Berberine effectively inhibited the activity of disaccharidases (alpha glucosidase) and decreased glucose transport through the intestinal epithelium [93] . Berberine was also reported to show antihyperglycemic activity by promoting the secretion of glucagon-like peptide-1 [94] . Catharanthine, vindoline and vindolinine are alkaloids isolated from Catharanthus roseus with potent hypoglycemic activity [95] . Arecoline, an alkaloid isolated from Areca catechu was reported to have hypoglycemic activity in animal model [96] . Aegeline, an alkaloidal amide from the leaves of Aegle marmelos was found to possess antihyperglycemic activity with suggested β₃-adrenergic receptor activity [97] . Trigonelline isolated from a known hypoglycemic plant Trigonella foeum graceum could lower blood glucose level in alloxan treated diabetic rats through regeneration of new islets [98] . The β-carboline alkaloids obtained from Peganum harmala stimulates insulin secretion in a dose dependent manner [99] [100] and are known insulin secretagogues from plant sources.

The results of this study suggest that the extract and fractions of N. laevis leaves possess antihyperglycemic activity in alloxan diabetic rats. Alkaloids, flavonoids, tannins, resins and saponins may be responsible for activity. This study gave credence to the ethnomedicinal use of N. laevis leaves in the management of diabetes. Further work is suggested to isolate the fraction(s)/compound(s) responsible for the activity.

The authors report no conflict of interest. The authors alone are responsible for the conduct and writing of this manuscript.

Chinyelu C.Osigwe,Peter A.Akah,Chukwuemeka S.Nworu,Theophine C.Okoye,Michel K.Tchimene, (2015) Antihyperglycemic Studies on the Leaf Extract and Active Fractions of Newbouldia laevis (Bignoniaceae). Pharmacology & Pharmacy,06,518-532. doi: 10.4236/pp.2015.611054