2011 YÜksek lisans tez özetleri

Total Antıoxıdant Capacıty Assay Of Serum Usıng Modıfıed Cuprac (Copper(Iı) Reducıng Antıoxıdant Capacıty) Method

Although individual antioxidant compounds in serum are roughly known by means of studies performed, a need for devising analytical methods for the determination of total antioxidant capacity (TAC) has been described in literature rather than determining the contribution of each component. This is because the combined effect of antioxidants in certain circumstances may exceed the individual effects. Especially because of inadequacies of analytical methods the antioxidative contribution to TAC of serum proteins is essentially unknown. Because in most TAC assays, proteins are initially separated by precipitation from the main matrix. In this work TAC of serum was measured with standard reference methods and the modified CUPRAC (CUPric Reducing Antioxidant Capacity) method, and the contribution of serum proteins, especially thiol-containing proteins was identified, to the measured TAC. The oxidizing reagent of the modified CUPRAC method for thiol-containing protein assay is the Cu(II)-neocuproine reagent which was previously used for the simultaneous determination of cysteine and cystine. As opposed to the Ellman method capable of determining thiol compounds but not other common antioxidants, CUPRAC is most advantageous in regard to its strong response to both common antioxidants and thiol compounds. Other reference spectrophotometric methods used for comparison of assay results were ABTS radical scavenging and FRAP (ferric reducing antioxidant power) methods. The results were calculated in the units of cysteine equivalents. TAC values for serum were calculated as results showed that Ellman assay (known to be sensitive to only thiols) estimated nearly 71.2% of TAC coming from serum protins, whereas the contribution of proteins to TAC of serum was calculated as 45.8% with ABTS and 69.4% with modified CUPRAC. As FRAP can only be used for whole serum samples, it is not possible to compare results. These findings clearly demonstrate that the contribution of serum proteins to TAC is by no means negligible, and should necessarily be taken into consideration for further antioxidant measurements. These precise determinations incorporating the contribution of proteins to serum antioxidant capacity is believed to be potentially useful to biochemists investigating the diagnosis, treatment, prognosis, and follow-up of diseases utilizing TAC measurement.