Benefits and Limitations of Lab-on-a-Chip Method over Reversed-Phase High-Performance Liquid Chromatography Method in Gluten Proteins Evaluation

Abstract

RP-HPLC (reversed-phase high-performance liquid chromatography) is widely used to determine the amounts of the different gluten protein types. However, this method is time-consuming, especially at early stages of wheat breeding, when large number of samples needs to be analyzed. On the other hand, LoaC (Lab-on-a-Chip) technique has the potential for a fast, reliable, and automatable analysis of proteins. In the present study, benefits and limitations of Lab-on-a-Chip method over RP-HPLC method in gluten proteins evaluation were explored in order to determine in which way LoaC method should be improved in order to make its results more compliant with the results of RP-HPLC method. Strong correlation ( ≤ 0.001) was found between numbers of HMW glutenin peaks determined by LoaC and RP-HPLC methods. Significant correlations ( ≤ 0.05) were obtained between percentages of HMW and LMW glutenin subunits calculated with regard to total HMW + LMW area. Even more significant correlation ( ≤ 0.001) was found when percentages of individual HMW areas were calculated with regard to total HMW. RP-HPLC method showed superiority in determination of gliadins since larger number and better resolution of gliadin peaks were obtained by this method.

Rapid and Easy High-Molecular-Weight Glutenin Subunit Identification System by Lab-on-a-Chip in Wheat (Triticum aestivum L.)

“…However, the HMW-GS identification system by SDS-PAGE was shown to be time-consuming and required a large gel system to clearly separate HMW-GS. Many research groups have tried to apply the Lab-on-a-chip system for HMW-GS identification, due to the fast and effective protein separation and quantification that offers the Lab-on-a-chip electrophoresis system [31,40]. However, the latter system is not commonly used for HMW-GS identification due to particular reasons [31].…”

“…Therefore, the upper marker in the standard cultivars harboring 1Dx2.2, such as Uri and Seodun, was adjusted before analyzing the HMW-GS composition (Figure 1). The recorded molecular weights of HMW-GS on Lab-on-a-chip were above 120 kDa [31]. Table 1.…”

“…Reverse-phase high-performance liquid chromatography (RP-HPLC) was developed and used to discriminate the allelic variation of HMW-GS [5,27]. In addition, several other methods, such as high-performance capillary electrophoresis (HPCE), matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), and Lab-on-a-chip capillary electrophoresis (Lab-on-a-chip) were applied to identify the composition of HMW-GS [28][29][30][31]. Of these methods, MALDI-TOF-MS is said to be appropriate for detecting new allelic variations of HMW-GS [29].…”

“…In contrast, Lab-on-a-chip has been recognized for being fast and easy to identify and quantify HMW-GS. However, this method is not properly used due to the fact that Lab-on-a-chip has not been completely established, specifically for HMW-GS identification [31]. Interestingly, the protein weight of HMW-GS on Lab-on-a-chip was shown to be higher than that observed on SDS-PAGE, and the electrophoresis pattern of HMW-GS on Lab-on-a-chip was different from SDS-PAGE.…”

Prediction of the genetic similarity of wheat and wheat quality by reversed-phase high-performance liquid chromatography and lab-on-chip methods

“…Furthermore, specifi c composition of HMW-GS in wheat cultivars is one of the most important genetic factors that infl uence the rheological properties of dough (PAYNE et al, 1987). It has been recently demonstrated that composition and quantity of HMW-GS could be determined by Lab-on-Chip (LOC) technique (ŽIVANČEV et al, 2013;2015), whereas for accurate determination of their amounts a combined extraction -Reversed-Phase High-Performance Liquid Chromatography (RP-HPLC) procedure developed by WIESER and coworkers (1998) is still used.…”