Protein Biomarker Screening on Effect of Freeze Dried Crocodile Blood and Vitamin C in Iron-deficient Anemic Rats

Abstract

The effectiveness of combined treatment on Freeze-dried Crocodile Blood (FCB) and vitamin C on serum marker proteins in iron-deficient anemic (IDA) rats is essential for understanding of crocodile blood properties. High-resolution proteomic analysis, including SDS–PAGE, combined with MALDI-TOF mass spectrometry, were applied for the study of serum proteins in Sprague-Dawley rats by using 8 groups of 5 male rats. Normal group (G1) was given AIN93G+Fe diet but anemic group (G2) was received AIN93G-Fe diet. Six rat groups in the treated groups received AIN93G-Fe diet for 4 weeks, and followed by AIN93G -Fe + FCB 500 mg/kg body weight combined with vitamin C 0, 50, 100 and 150 mg/kg body weight, respectively (G3-G6), and followed by AIN93G -Fe + FCB 500 mg/kg body weight + 60 mg of ferrous sulphate (G7) and AIN93G -Fe + 60 mg of ferrous sulphate (G8). After treatment, the rat’s blood from the tail-vein was collected for hematological and serum investigations at 0, 4, 8 and 12 weeks of rat feeding. Haptoglobin (39.0 kDa) [or preprohaptoglobin (30.4 kDa)] and alpha-1-inhibitor III (165.0 kDa) were expressed in percentage of protein band on 7.5% SDS-PAGE gel since 4 weeks. Alpha-1-inhibitor III was conspicuously visualized at 8 weeks in normal group but this protein band was very low in IDA rats. Haptoglobin was markedly expressed in anemic group but its protein band was found to decrease in normal group. The combined treatment on the FCB and vitamin C that had a tendency to increase alpha-1-inhibitor III and to decrease haptoglobin was revealed efficient in a low dose of vitamin C (50 mg/day). Serum albumin (70.7 kDa) is difficult to assign as a protein biomarker because it could not correspond to anemic induction at 4 weeks. Results concluded that effects of FCB and vitamin C may lead to change on haptoglobin and alpha-1-inhibitor III screening.