New Modified Method for Determination of Nitric oxide Synthase Activity in Plasma of Vitiligo Patients

Vitiligo is a non-contagious skin disorder that characterized by depigmentation of skin due to melanocyte impairment which may be caused to increase levels of free radicals (such as superoxide and nitric oxide) that causing an increase in oxidative stress. The purpose of this study was measured by the activity of oxide synthase (NOS) by our modified method and nitric oxide concentration in plasma of vitiligo patients. The activity of nitric oxide synthase was determined via a modified method by coupling two methods; the first method was based on converting L-arginine to L-citrulline and nitric oxide and the second was used to measure the concentration of nitric oxide. This modified method was applied to patients with vitiligo disease and healthy individuals who matched in age and gender with patients. The condition of this modified method was optimized and the results revealed the following: the activity of NOS was higher in a solution that contains: Tris buffer (50mM), arginine (100mM), calcium chloride (20mM), and NADPH (5mM) during 30 min. meanwhile the precision of this method was 2.03. In the current study, the results show that the levels of NOS activity and nitric oxide were affected by the disease in which both parameters appeared highly significant increases in vitiligo patients (p=0.000 and 0.002 respectively) in comparison with the healthy individuals. Results of the experiments proved that it is possible to depend on the modified method to measure the activity of nitric oxide synthase (NOS). Also, the increased levels of NOS activity and nitric oxide concentration in vitiligo patients support the autocytotoxic hypothesis which suggests that oxidative stress may have a role in melanocyte impairment.