[논문]Fluorescence-labelling for analysis of protein in starch using asymmetrical flow field-flow fractionation (AF4)

Yoo, Yeongsuk; Choi, Jaeyeong; Zielke, Claudia; Nilsson, Lars; Lee, Seungho

doi:10.5806/ast.2017.30.1.1

Fluorescence-labelling for analysis of protein in starch using asymmetrical flow field-flow fractionation (AF4) 원문보기

분석과학 = Analytical science & technology, v.30 no.1, 2017년, pp.1 - 9

Yoo, Yeongsuk (Department of Chemistry, Hannam University) , Choi, Jaeyeong (Department of Chemistry, Hannam University) , Zielke, Claudia (Department of Food Technology, Engineering and Nutrition, Lund University) , Nilsson, Lars (Department of Food Technology, Engineering and Nutrition, Lund University) , Lee, Seungho (Department of Chemistry, Hannam University)

Abstract ▼ AI-Helper

Starch is a mixture of amylose (AMY) and amylopectin (AMP) which are different in physical properties such as molar mass (M), rms radius ($R_g$) and hydrodynamic diameter ($d_H$). The rheological and functional properties of starch are influenced by various factors including the molecular size, molar mass distribution (MD) and the concentration ratio of AMY and AMP. It is also important to analyze proteinaceous material in starch as they affect the flavor and texture of food to which starch is added. In this study, asymmetrical flow field-flow fractionation (AF4) was employed for separation and quantitation of AMY and AMP in starches (Amaranth, potato, taros and quinoa). AF4 was coupled with a multi-angle light scattering (MALS) and a refractive index (RI) detector for determination of the absolute M, MD and molecular structure. It was found that AMP has the M and $R_g$ ranging $3.7{\times}10^7{\sim}6.5{\times}10^8g/mol$ and 84 ~ 250 nm, respectively. Also the existence of branch was confirmed in higher M. In addition, proteinaceous material in starch was analyzed by AF4 coupled with a fluorescence detector (FS) after fluorescence-labeling. AF4-FS with fluorescence-labelling showed a potential for investigation on existence of proteinaceous material and the interaction between proteinaceous material and polysaccharide in starch.

주제어

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제안 방법

In this study, the molecular conformation as well as the molecular mass and size were analyzed using an AF4-MALS-RI for various types of starches.

대상 데이터

The AF4 system used in this study was an Eclipse 2 Separations System (Wyatt Technology Europe, Dernbach, Germany). It was connected with a RI detector (Shimadzu, Kyoto, Japan), a DAWN EOS MALS detector (Wyatt technology, Santa Barbara, CA, USA) operating at the wavelength of 690 nm, and a RL-10AXL fluorescence detector (Shimadzu, Kyoto, Japan). A refractive index increment, dn/dc, for starch of 0.
In this study, four starch varieties from botanical origin: potato, taros, amaranth and quinoa were used. Starch samples were received from Department of Food Technology, Engineering and Nutrition in Lund University (Lund, Sweden). Starch solutions were prepared by the so-called ‘DMSO method’.
The AF4 system used in this study was an Eclipse 2 Separations System (Wyatt Technology Europe, Dernbach, Germany). It was connected with a RI detector (Shimadzu, Kyoto, Japan), a DAWN EOS MALS detector (Wyatt technology, Santa Barbara, CA, USA) operating at the wavelength of 690 nm, and a RL-10AXL fluorescence detector (Shimadzu, Kyoto, Japan).

이론/모형

For analysis of proteinaceous material in starch using AF4, fluorescence labeling was tested using amine-reactive fluorescent dye method. The labeling solution was prepared by mixing 10 mM 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide (EDAC) and 10 mM 7-methoxycoumarin-3-carboxylic acid in the volume ratio of 1:1 for 3 h.
MALS data processing was performed using the Astra software. The Berry method was used for datafitting in this study.

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