Hypoxia is a condition where a region of tissue has less than adequate oxygen. It is of particular importance in tumor biology. The hypoxic core of tumors has been shown to impede the effectiveness of various therapies, such as photodynamic therapy, radiotherapy and chemotherapy. The determination of oxygen saturation levels in blood and other tissues in vivo is critical for ensuring proper body functioning, monitoring status of diseases, e.g. cancer, and predicting the efficacy of therapy. Previously spectroscopic photoacoustic imaging technology has been used for in vivo blood oxygenation measurements by exploiting the distinct optical absorption spectra of oxygenated- and deoxygenated-hemoglobin. However, the accuracy of such measurements is affected by the heterogeneous light fluence in biological samples. Hemoglobin based spectroscopic measurements are also ineffective in blood devoid tissues such as tumors, especially in the necrosis core. Here we demonstrate, for the first time, a lifetime based photoacoustic technique for the measurement of oxygen saturation in vivo, using a porphyrine based oxygen sensitive dye. This molecular probe aided and lifetime based oxygen sensing technique overcomes the problems mentioned above. The experimental results derived from the main artery in the rat tail indicated that the lifetime of the probe, quantified by the photoacoustic measurement, shows a good linear relationship with the blood oxygenation level in the targeted artery.