In the past, one of the major problems in gluten analysis has been the unavailability of an efficient, universal, extraction procedure of gliadins - the alcohol-soluble proteins of gluten - from both heat processed and unprocessed products.
Dr Garcia and colleagues designed a study to develop a universal, extraction procedure capable of extracting the totality of gliadins from both unprocessed and heat processed foods for celiac patients.
The research team developed a simple quantitative extraction solution containing 250 mM 2-mercaptoethanol and 2 M guanidine hydrochloride ('cocktail'), to extract gliadins from heated foods.
The researchers mentioned that the diluted reducing and disaggregating agents reaching the micro plate at low concentration do not affect the ELISA system based on the R5 monoclonal antibody.
|The recovery of gliadins with 2 M guanidine hydrochloride was 96 % versus 44 % with ethanol|
|European Journal of Gastroenterology and Hepatology|
The invesitigators found that the recovery of gliadins extracted by the cocktail from spiked samples was nearly complete, with a mean value of 96 %, which is clearly superior to 44 % obtained with conventional 60 % aqueous ethanol.
The cocktail always yielded either slightly similar or higher values than 60 % aqueous ethanol depending on the type of foods:
The research team reported that the cocktail yielded values 1.1 fold in unheated foods, 1.4-fold in wheat starches and 3-fold in heated foods and false positives or negatives were never observed using the cocktail solution.
Dr Garcia concludes, “We present a general complete gliadin extraction procedure based on reducing and disaggregating agents for both heated and unheated foods as a crucial tool for gliadin analysis.”
“The new extraction solution is used for corresponding proteins from rye (secalins) and barley (hordeins).”
“The cocktail was employed as the extraction method in the international ring trial evaluation of sandwich R5-ELISA as proposed by the Codex Alimentarius and organized by the Working Group on Prolamin Analysis and Toxicity.”