Synthesis of novel 3-benzyl-2-phenyl-4(5H)-(substituted phenylhydrazino)-1, 3-oxazolidines and their antihyperglycemic activity

A number of 3-benzyl-2-phenyl-4(5H)-(substituted phenylhydrazino)-1, 3-oxazolidines 6a-f derivatives were synthesized and their antihyperglycemic activity was evaluated by streptozotocin (STZ) and sucrose-loaded model. Many of oxazolidine derivatives as well as their corresponding substituted phenylhydrazine showed significant reduction in blood glucose level in STZ and sucrose-loaded rat models.


INTRODUCTION
Diabetes mellitus is a main threat to human health in the 21 st century. [1] The incidence of the disease currently is estimated to reach 300 million by the year 2025. [2] Oxazolidine is an important group of heterocyclic compounds that plays a vital role in many biological activities in the areas of medicine and molecules containing oxazolidine ring systems and exhibits many activities such as anti-TB activity, [3] antibacterial activity, [4] mono-amine oxidase inhibition, [5] and GP IIb/IIIa antagonism [6] neuroleptic activity. [7] The information on oxadiazolidine as antihyperglycemic agents has been reported previously. [8] In view of the literature led to the conception that a novel series of 3-benzyl-2-phenyl-4(5H)-(substituted phenylhydrazino)-1, 3-oxazolidines 6a-f derivatives were synthesized and their chemical structure were confi rmed by IR, 1 H-NMR, mass spectral and elemental analyses. These compounds were screened for their antimicrobial activities.

Materials
Synthetic starting material, reagents and solvents were of analytical reagent grade or of the highest quality that were commercially available and were purchased from Aldrich Chemical Co (Hyderabad, AndhraPradesh, India), Merck Chemical Co (Mumbai, Maharashtra, India) and Acros Organics (Mumbai, Maharashtra, India) and were dried when necessary.
The melting points were taken in open capillary tube and uncorrected. IR spectra were recorded with KBr pellets (ABB Bomem FT-IR spectrometer MB 104 ABB Limited, Bangaloru, India). Proton ( 1 H) NMR spectra (Bruker 400 NMR spectrometer, Mumbai, India) were recorded with TMS as internal references. Mass spectral data were recorded with a quadrupole mass spectrometer (Shimadzu GC MS QP 5000, Chennai, India) and microanalyses were performed using a vario EL V300 elemental analyzer (Elemental Analysensysteme GmbH, Chennai, India). The purity of the compounds was checked by TLC on pre-coated SiO 2 gel (HF 254 , 200 mesh) aluminium plates (E. Merck) using ethyl acetate:benzene ratio of 1:3 and visualized in UV chamber. IR, 1 H-NMR, mass spectral data and elemental analyses were consistent with the assigned structures.
The in vivo antihyperglycemic activity of synthesized compounds was evaluated using male Sprague-Dawley rats in two diff erent models.

Streptozotocin model
A solution of streptozotocin (STZ) (60 mg/kg) in 100 mM citrate buff er, pH 4.5 was prepared and calculated amount of the fresh solution was dosed to overnight fasted rats (60 mg/kg) intraperitoneally. The blood sugar level was measured a er 48 h by glucometer. Animals showing 200-400 mg/dL were selected for antidiabetic screening. The diabetic animals were divided into groups of six animals each. Rats of experimental group were administered a suspension of the desired test sample (prepared in 1% gum acacia) orally (100 mg/kg body weight). Controlled group animals were also fed with 1% gum acacia. The blood glucose levels were measured at 1-, 2-, 3-, 4-, 5-, 6-, 7-and 24-h intervals. The % fall in blood glucose from 1 to 24 h by test sample was calculated according to the area under curve (AUC) method. The average fall in AUC in experimental group compared to control group provided % antihyperglycemic activity.

Sucrose-loaded (SLM) model
Overnight-fasted male Sprague-Dawley rats were used for sucrose-loaded experiment. Blood was collected initially and there a er test compounds were given to the test group consisting of five rats by oral gavage at a dose of 100 mg/ kg body weight. A er half an hour post-test treatment, a sucrose load of 10 gm/kg body weight was given to each rat. Blood was collected at 30, 60, 90 and 120 min post-sucrose load. The % fall in blood glucose level was calculated according to the AUC method.  Figure 1: 3-benzyl-2-phenyl-4(5H)-(substituted phenyl hydrazino)-1, 3-oxazolidines

Antihyperglycemic activity
Among the six screened compounds only 6e, 6a, 6b exhibit antihyperglycemic activity in STZ model and SLM model. Compound 6c displayed 27.3% blood glucose lowering activity at 100 mg/kg dose in SLM model and 6d showed 13.4% blood glucose lowering activity at 100 mg/kg dose in STZ model. In these compounds, 3-benzyl-2-phenyl-4(5H)-4'acetamidophenyl hydrazino-1, 3-oxazolidine 6e reduced blood glucose level to 54 and 71% in STZ and SLM models, respectively. It is evident from Table 1 that, the presence of benzyloxy (6f) caused a complete loss of antihyperglycemic activity. A closer inspection of data from this table indicates that compounds acetamido, bromo and chloro phenylhydrazino substitution of oxazolidine having excellent antihyperglycemic activity compared with standard. The transformation order of screened compounds is 6e>6a>6b>6c>6d>6f. Metformin and glybenclamide were used as standard antidiabetic drug in both the models.