Evaluation of offgel electrophoresis, electrothermal atomic absorption spectroscopy and inductively coupled plasma optical emission spectroscopy for trace metal analysis in marine plankton protein

摘要

Two-dimensional offgel and lab-on-a-chip (LOC) electrophoresis was performed for assessing proteins from marine plankton. Proteins were isolated from plankton samples by using a phenol/sodium dodecylsulfate (SDS) buffer (2:1) pH 8.0 mixture as an extracting solution. The SOS buffer consisted of 30% (m/v) sucrose, 2% (m/v) SDS, 0.1 M of Tris-HCl, pH 8.0,5% (v/v) beta-mercaptoethanol and protease inhibitor cocktail. Before protein extraction, the plankton were subjected to different washing steps involving 10% trichloroacetic acid (TCA)/acetone and methanol. Proteins exhibiting the highest molecular weights (MWs), within the 52.6-57.3 kDa range, were found in offgel fractions of isoelectric points (pis) ranging from 4.40 to 8.60; whereas, the highest protein concentrations were found in fraction 5 (100 mu g g(-1) for a protein of pI of 4.5 and MW of 22.0 kDa), fraction 16 (55 mu g g(-1), protein of pI/MW of 7.4/19.1 kDa), and fraction 8 (51 mu g g(-1), pI/MW of 53/22.2 kDa). Further studies were also developed for determining the amount of trace metals associated with the isolated proteins of similar pIs by electrothermal atomic absorption spectroscopy (ETAAS) and inductively coupled plasma optical emission spectroscopy (ICP-OES). Preliminary results indicate that Fe, Ni, Mn and Zn were found to be associated with proteins of variable MW (from 14 to 54 kDa) and pls (from 3.4 to 8.2). Otherwise, Cd was found to be associated with proteins of certain pls (6.0-6.5) and with MWs ranging from 14.1 to 53.1 kDa. (C) 2014 Elsevier B.V. All rights reserved,