Chronotherapeutics refers to a treatment method in which in vivo drug availability is timed to match rhythms of disease, in order to optimise therapeutic outcomes and minimise side effects. The term "chrono" basically refers to the observation that every metabolic event undergoes rhythmic changes in time. It refers to a treatment method in which in vivo drug availability is timed to match rhythms of disease in order to optimize therapeutic outcomes and minimize side effects. It is based on the observation that there is an interdependent relationship between the peak-to-trough rhythmic activity in disease symptoms and risk factors, pharmacologic sensitivity, and pharmacokinetics of many drugs. In this present study invivo evaluation of bilayered tablets of Losartan and Hydrochlorothiazide were studied on rabbits to prove that the prepared formulation shows a pulsatile drug release. The plasma levels were measured using LC-MS/MS method. Pharmacokinetic parameters of these tablets were determined by using WINNOLIN Scientific software. The pulsatile release tablet formulation prepared in the lab managed to show some lag phase initially before releasing the drug with a maximum time (Tmax) at the 12th hour. From this present work the results of invivo studies proved that chronotherapeutical release of drug in the form of bi layered tablet can be achieved for newer chronotherapeutical drug delivery system.