Elsevier

Journal of Cereal Science

Volume 80, March 2018, Pages 150-157
Journal of Cereal Science

Proteomic analysis of two malting barleys (Hordeum vulgare L.) and their impact on wort quality

https://doi.org/10.1016/j.jcs.2018.02.004Get rights and content

Highlights

  • Comparison of proteome of two brewing malts was done by 2D-PAGE and Zymograms.

  • Quality parameters in worts were related with proteins and enzymatic activity.

  • Difference in hydrolytic proteins were observed using zymograms.

  • A small molecular size amylase was detected.

Abstract

Malted barley contributes directly to wort quality. Wort prepared with malt M1 had a higher free amino nitrogen content, whereas wort prepared with malt M2 had a higher diastatic power. The proteome analysis (pI 3–10) in 2D-PAGE gels showed significant differences, with M1 generating 246 ± 9.8 spots and M2 229 ± 9.8 spots. M1 had more hydrolytic enzymes, in particular an esterase with a size of 43.7 kDa and a pI of 9.4, whereas M2 had more proteins involved in stress tolerance and carbohydrate metabolism, particularly a β-glucosidase with a size of 58.7 kDa and a pI of 7.7, which coincided with its higher diastatic power. The biological activity of M1 showed a band at ca. 150 kDa with high proteolytic activity and another band at 27 kDa with amylase activity. This study is the first report of amylase activity of small molecular size in malts.

Introduction

Barley (Hordeum vulgare) is the fourth cereal most cultivated in Mexico with two grown cycles: Spring-summer cycle (rainfed lands) and Fall-winter cycle (irrigated lands) (Peñalva, 2011). Five varieties meet the quality criteria for brewing industry: Armida, Adabella, Esmeralda, Alina and Esperanza and correspond for the 70% of the total year production in the country (Peñalva, 2011). The use of one or another variety depends on the desired characteristics of the final beer. Malting is a process during which the barley grain is subjected to humid conditions to activate the germination process and then dried. This process induces the enzymes required to transform the grain stock (starch and proteins) into the components required for fermentation.

The qualities of the malt and the final beer are defined by the characteristics of the barley grains and the malting parameters. The barley malt chemical components, germination and viability are mainly determined by the grain characteristics, which are affected by the malting conditions and growing environments (Guo et al., 2016). Each malted barley variety generates unique flavor and body, which is exploited to produce different types of beer. Understanding the biochemical characteristics of the malt to predict the characteristics of the final beer is one of the major goals of the brewing industry.

Many studies have focused on the metabolic proteins present in the malt that influence its quality. The studied plant species have included Arabidopsis thaliana, which is used as a study model, barley (Hordeum vulgare), corn (Zea mays) and rice (Oryza sativa) (Li et al., 2014). Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) is the most popular proteome analysis method. When this method is combined with mass spectrometry identification procedures and enzymatic activity studies using zymography, comparisons can be performed between different malts to find proteins with potential as quality markers.

Some studies, such as the study of Zhao et al. (2013), have related metabolic proteins separated in a pH range of 4–7 with the characteristics of two types of malts with different quality levels for the brewing industry. Other examples of comparative analyses using proteomic techniques include the study of Guo et al. (2016) who compared the proteomes of two barley varieties used for the food and brewing industries with an aim toward identifying the protein content in each variety, and the study by Jin et al. (2013), in which two malts with different degrees of filterability were compared.

Various studies on the hydrolytic systems of malts have been conducted using electrophoretic separations with copolymerized substrates, which are better known as zymograms. These studies have found that distinct malt varieties can contain different band profiles with types of proteolytic activity at different pH values (Wrobel and Jones, 1992).

Due to the importance of studying variations in the proteome and its relationship to the quality of the malt and the beer, the differences in the proteomic profiles of two varieties of malted barleys commonly used in Mexico brewers were studied to obtain information at the proteomic level concerning the differences between the varieties and to relate the proteins found in the malts with wort quality parameters.

Section snippets

Biological samples

The malted barley grains (Hordeum vulgare) samples were provided by the Cuauhtémoc Moctezuma Brewery, Mexico. The malts were called M1 and M2 and were stored at −20 °C prior to use. Malts were produced using two different Mexican barley varieties by a malting company using the same conditions.

Protein extraction

The malts (10 g) were ground into a fine powder (<50 μm). The samples were homogenized in 1 ml of acetate buffer (50 mM, pH 5.0) and extracted by vortex for 30 min, resting in an ice bath every 5 min.

Malt quality

Brewing depends mainly on the biochemical characteristics of the wort, which are directly related to the type of malted barley used in the preparation. In this work, the proteome of two brewing malts and their relation to the wort quality are reported. Both malts are cultivated in different seasons of the year (winter and summer) and both are commonly used in the brewing industry in Mexico. Their use in the process produce different flavor profiles in beer (Cuauhtémoc Moctezuma Brewery). The

Funding source

We thank the National Council of Science and Technology (Consejo Nacional de Ciencia y Tecnología) (CONACYT) for financial support through project PEI 231412. JG. Herrera-Gamboa and CB. López-Alvarado received CONACYT scholarships.

Author contributions

JGHG and CBLA designed and performed the experimental work, BPA directed the project. JGHG wrote the paper with the supervision of BPA. All authors provided critical feedback and helped shape the project and paper. EPO, LCDB and JCCA gave the approval to the final version of the paper.

Conflicts of interest

All authors declare no competing financial interest.

Abbreviations used

M1, Malt 1; M2, Malt 2; 2D-PAGE, Two-dimensional polyacrylamide gel electrophoresis; kVh, kiloVolts per hour; HCl, Hydrochloric acid; SDS, Sodium dodecyl sulfate; DTT, Dithiothreitol; SDS-PAGE, sodium dodecyl sulfate polyacrylamide gel electrophoresis; mA, miliAmpere; kDa, kiloDalton; μg, micrograms; MgCl2, magnesium chloride; FAN, Free amino nitrogen; KI, Kolbach index.

Acknowledgments

We thank Rocío Ortiz López (CIDICS-UANL) and Víctor Aguirre (Facultad de Agronomía-UANL) for their support in conducting the 2D-PAGE technique.

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