聚乙烯聚吡咯烷酮吸附多酚物质性质研究及其在无甲醛酿造啤酒工艺中的应用

本文主要研究PVPP(聚乙烯聚吡咯烷酮)吸附多酚的性质,及其在无甲醛酿造工艺中的应用。结果表明,麦汁或啤酒中的敏感蛋白、酒精度、pH以及PVPP的添加量和添加方式等因素对PVPP吸附多酚都有一定影响。当pH在4.0左右,酒精度在3%(V/V)左右时PVPP的吸附效率最高。PVPP在添加量较低的情况下有一定的吸附专一性,主要吸附高分子的聚合多酚,但在添加量较高的情况下,不具备此性质,对单体酚的吸附迅速增加,会降低麦汁或啤酒的抗氧化能力。PVPP结合硅胶使用可以有效替代甲醛在啤酒酿造中的应用,在保证啤酒非生物稳定的条件下,改善啤酒风味稳定性和抗氧化能力。     

The Role of Malt and Hop Polyphenols in Beer Quality,Flavour and Haze Stability

Pilot‐scale brewing trials of a 12°P pale lager beer were conducted to look at the effect of a modified dose of hop and malt polyphenols on haze, flavour quality, and stability. Results confirmed that malt polyphenols, and particularly hop polyphenols, in the course of wort boiling, improved reducing activity values and the carbonyl content in fresh and stored beers. Hop polyphenols significantly increased reducing activity and decreased the formation of carbonyls (TBA value) in fresh and stored beer. Reduced content of malt polyphenols, combined with the use of hop CO₂ extract, caused an increase in the TBA value in beer. PVPP stabilized beers tended to be lower in reducing activity. Both malt and hop polyphenols affected the intensity of “harsh taste” in fresh beers and a significant influence from PVPP stabilization of beer was not observed. The staling degree of forced‐aged beers depended on the polyphenol content in the brewhouse. Both hop and malt polyphenols had a positive impact on flavour stability. PVPP treatment of beer had a positive effect on the flavour stability of heat‐aged beers. Polyphenols, especially hop polyphenols, slowed down flavour deterioration during the nine month storage period, but the primary effect was seen during the first four months of storage. Storage trials did not show any unambiguous effects for PVPP stabilization on beer flavour stability. Results confirmed the negative impact of malt and hop polyphenols on haze stability, and PVPP stabilization minimized differences in shelf life prediction values between beers prepared with the modified dose of polyphenols.     

Optimising Beer Stabilisation by the Selective Removal of Tannoids and Sensitive Proteins

Colloidal stabilisation of all malt and adjunct lager beers through selective removal of haze sensitive glycoproteins using silica gel and tannoid polyphenols using polyvinylpolypyrrolidone (PVPP) or a polyvinylpyrrolidone (PVP)‐modified silica gel has been demonstrated during full brewery production. At this scale PVPP and the PVP‐modified silica gel exhibited equivalent binding capability for tannoid polyphenols although PVPP removed additional polyphenolic species. Characterisation of proanthocyanidins following treatment at 4°C with PVPP and the PVP‐modified silica gel silica co‐product confirmed that PVPP removes a wider range of polyphenolics. Both products exhibited poorer polyphenolic binding capability at 4°C as would be expected for a physical adsorption process.     

Turbidity and Haze Formation in Beer — Insights and Overview

Beer is a complex mixture of over 450 constituents. In addition, it contains macromolecules such as proteins, nucleic acids, polysaccharides and lipids. Proteins influence the entire brewing process with regard to enzymes, which degrade starch, β‐glucans and proteins; with protein‐protein linkages that stabilize foam and are responsible for mouthfeel and flavour stability; and in combination with polyphenols, thought to form haze. With this complexity, problems in processability are as various as the constituents. Several substances in beer are responsible for haze formation. Organic components such as proteins, polyphenols and carbohydrates (α‐glucans, β‐glucans) are known to form haze. In addition, inorganic particles such as filter aids and label remains can cause increased turbidity. In this article only non‐microbiological induced hazes are described. Many studies have been conducted on the identification of haze and foam active components in beer. Hence the aim of this work was to survey the different possibilities of haze formation and for haze identification. A summary is provided on methods for haze identification including dyeing methods, microscopic analyses and size exclusion chromatography.     

Beer Polypeptides and Silica Gel Part I. Polypeptides Involved in Haze Formation

Beer contains approximately 500 mg/L protein depending on the brewing procedures employed. This protein is in the form of polypeptides, the majority of which lie within the 10–40 kD size range. Some of these polypeptides are responsible for causing colloidal haze, others enhance foam stability and the remainder appear to have no function in beer except to contribute to mouth‐feel. The polypeptides responsible for haze formation are those that can combine with polyphenols to produce a visible cloudy haze. This is undesirable as it can have a negative effect on the beer's shelf life. One way to reduce this effect is to remove these polypeptides using silica gels. It is important that this removal is selective, and the desirable foam enhancing polypeptides are not removed. Data will be presented to show that beer polypeptides are glycosylated and that silica preferentially adsorbs glycoproteins, particularly those with protein components rich in the amino acid proline. The molecular size and composition of glycoproteins recovered from untreated beer, cooked adjunct, silica exposed to beer and beer aged for one year are presented. Glycoproteins involved in foam, and the apparently functionless polypeptides, will be discussed in a subsequent paper.     

A Discussion of Polyphenols in Beer Physical and Flavour Stability

Generally referred to as polyphenols (PPs), beer flavonoids such as the flavan‐3‐ols and their condensed products, the proantho‐cyanidins, represent a class of readily oxidizable compounds capable of hindering or preventing the oxidation of other molecules present in beer. Flavan‐3‐ol and proanthocyanidin capacity to improve oxidative stability has been well established in other food systems, and thus these antioxidants have recently gained significant consideration as potential beer flavour modifiers and/or stabilizers. The duality of their presence in beer is that PPs complex with proteins in the beer matrix to form temporary and permanent hazes. Undesirable physical instability caused by PP‐protein interactions can be resolved via use of adsorptive resins such as polyvinylpyrrolidine. While there is no doubt that polyphenol removal increases beer shelf stability in terms of haze formation, the impact of polyphenol removal on beer flavour remains unresolved. This review discusses the sources, content and impact of polyphenol presence and removal on beer physical and flavour stability.     

应用新技术提高啤酒的胶体稳定性

引起啤酒胶体非稳定性混浊的主要物质为蛋白质和多酚物质。可采取在麦汁煮沸阶段加入胶体物质和在贮酒过程中加入硅胶，使啤酒中的高分子蛋白质沉淀而过滤除去。利用PVPP作为多酚吸附剂对啤酒进行二级过滤，可降低啤酒中的多酚物质，也可将PVPP同硅藻土、硅胶联用过滤，降低啤酒的多酚物质。     

啤酒中浑浊敏感蛋白的分离与鉴定

啤酒非生物稳定性是影响啤酒质量的重要指标之一,非生物浑浊主要是啤酒中的浑浊蛋白质与多酚结合体组成的,浑浊敏感蛋白质主要来自大麦醇溶蛋白,是一种富含脯氨酸的蛋白质.主要阐述了浑浊敏感蛋白质的分离,以及利用蛋白免疫印迹法(Immunobloting)鉴定啤酒中的浑浊敏感蛋白质,此外还研究了硅胶对啤酒的浑浊蛋白质的影响.     

PVPP对啤酒中多酚类物质和蛋白质的吸附作用比较

为了比较PVPP对啤酒中多酚类物质和蛋白质的吸附作用,以牛血清蛋白为例,采用PVPP分别处理原啤酒、牛血清蛋白(BSA)溶液以及BSA原啤酒混合溶液。实验发现,在相同条件下,尽管PVPP对牛血清蛋白具有较强的吸附作用,但对啤酒中多酚类物质的吸附作用更强;而在BSA原啤酒混合溶液中,多酚类物质被吸附量远远大于牛血清蛋白。结果表明,多酚类物质通过竞争吸附占据PVPP的吸附活性点。     

The chemical components in malt associated with haze formation in beer

In bright beer, haze formation is a serious quality problem, which reduces beer storage and shelf life. In this study, haze‐active proteins, alcohol chill haze formation ability, α‐amylase activity, the contents of total polyphenol, protein and its fractions and amino acids were analysed using 23 barley accessions to investigate the relationship between the quality components in the malt and the haze character in beer. The results showed that there were largely genotypic variations for all examined traits among the 23 barley accessions. However, there was no significant correlation between the haze character and α‐amylase activity. All haze characteristics were significantly and positively correlated with total protein content, albumin, globulin and the hordein content, as well as the glutamic acid (glutamine), proline and phenylalanine content, and were not correlated with total polyphenols. A model describing the relationship between the chill haze in the beer and the protein content in the malt was developed. Copyright © 2016 The Institute of Brewing & Distilling     

Beer Polypeptides and Silica Gel Part II. Polypeptides Involved in Foam Formation

Beer contains approximately 500 mg/L protein depending on the brewing procedures employed. This protein is in the form of polypeptides, the majority of which lie within the 10–40 kD size range. Some of these polypeptides are responsible for causing colloidal haze, some enhance foam stability and the remainder appear to have no function in beer except to contribute to mouth‐feel. Those polypeptides involved in haze formation were described in a previous paper. To continue these studies, data is presented to show that foam polypeptides are highly glycosylated and that purified foam glycoprotein contains low levels of the amino acid proline. As silica preferentially adsorbs polypeptides rich in proline, it is unlikely to adsorb this material and damage foam stability. The molecular sizes and composition of glycoproteins recovered from untreated beer, purified foam and beer from which the foam component has been removed are presented. These fractions include the polypeptides responsible for foam stability and those that appear to have no role in physical stability.     

Prediction of Colloidal Stability of Highly Stabilized Beers by a Modified Chapon Tannoid Content Test

The tannoid content test, introduced by Chapon, is accepted as a recommended MEBAK analytical method for the detection of haze‐forming polyphenols in beer and other intermediates. Under certain conditions, the results can be used directly for predicting the colloidal stability of beer. Its disadvantage is that it does not provide detectable results for beers stabilized by high doses of polyvinylpolypyrrolidone (PVPP). A test modification is proposed that increases the test sensitivity and gives non‐zero results, even for highly stabilized samples that are not measurable by the standard test. The modification is based on adding saturated ammonium sulphate solution (SASS) to the sample to reach 9% saturation (0.5 mL of SASS to 5 mL sample volume) before the actual test performance. The results of the modified test can simply be converted to results obtained by the standard method. Tannoid content correlates linearly with the content of haze‐forming polyphenols and with chill haze produced after defined accelerated aging. In a set of beer samples of one brand having different degrees of stabilization, a high correlation was determined between test results and long‐term colloidal stability. The modified test can be used as a rapid and simple test for checking the stabilization process. It is particularly suitable for Pilsner type beer of long (more than 6 months) colloidal stability controlled by stabilization on the side of polyphenols.     

硅凝胶对提高啤酒非生物稳定性的应用研究

本试验研究是利用硅凝胶吸附高分子蛋白质的特点,以提高啤酒非生物稳定性的应用研究。试验筛选出适用于啤酒的,经特定制备方法制得的C-Ⅰ型硅凝胶,其对啤酒中高分子含氮物质具有较强的选择性吸附作用,可有效地提高啤酒的非生物稳定性,且不改变啤酒的其它纽分和外观质量。试验结果表明:适宜的吸附处理时间为5～30分钟,吸附温度为0℃,起到了提高啤酒非生物稳定性和不改变啤酒其它质量指标的作用。     

THE REACTIONS BETWEEN POLYPHENOLS AND ALDEHYDES AND THE INFLUENCE OF ACETALDEHYDE ON HAZE FORMATION IN BEER

The reactions and interactions between proteins and polyphenols are, among other phenomena, responsible for haze formation in beer. The participation of aldehydes in the polymerisation of polyphenols is considered. The formation of phenolic Baeyer-type condensation products containing phenolic residues linked by CH₃CH-bridges through reaction with acetaldehyde is possible at the pH of beer (4.0). These and other reactions with acetaldehyde in beer participate in beer haze formation.     

啤酒混浊蛋白组分的分离鉴定

采用双向电泳结合基质辅助激光解吸/ 电离飞行时间质谱(MALDI-TOFMS)技术分析确定引起啤酒混浊的蛋白组分。结果表明,仅有少量的蛋白组分参与啤酒混浊的形成。聚丙烯酰胺凝胶电泳分析发现MW 40kD 左右,25～29kD 和6.5～17kD 作为啤酒混浊蛋白组分,可能主要来自麦芽中的水溶蛋白,小部分来自麦芽醇溶蛋白。双向电泳分析证实了大部分的麦芽蛋白在酿造过程中发生降解和变性,并结合质谱技术鉴定得到BTI-CMe、germin E(Hordeum vulgare)和 protein Z 3 种组分可以抵抗酿造过程的热变性和水解作用,将成为啤酒混浊形成重要的促进因子。     

苹果汁中活性蛋白的性质及其稳定化处理研究进展

苹果汁中活性蛋白质 (HAP)是澄清苹果汁产生混浊沉淀的重要原因之一 ,疏水性氨基酸易于和多酚聚集 ,形成混浊。苹果汁中不同浓度的活性蛋白质与低分子的多酚以及简单酚所引起混浊的反应机理不同 ;活性蛋白质的种类、分子结构、反应的时间、温度、反应体系的 pH、无机离子及其离子强度对浑浊的产生有重要影响。单宁酸、膨润土、硅溶胶和聚乙烯聚吡咯烷酮 (PVPP)是有效的稳定化处理措施。     

用浊度法预测香蕉汁中混浊活性蛋白质和多酚

本研究采用浊度仪测香蕉汁混浊活性(HA)蛋白和多酚含量。为减少内源性蛋白和多酚的影响,分别用聚乙烯吡咯烷酮(polyvinyl polypyrrolidone, PVPP)和膨润土吸附多酚和蛋白质。将单宁酸加入PVPP处理与未处理香蕉汁中,果汁浊度变化与蛋白质含量呈线性相关,在蛋白质含量相同情况下,PVPP处理果汁的浊度较未处理样品小。热处理添加适量明胶的香蕉汁可反映果汁中HA多酚的含量,膨润土处理与未处理系列样品的浊度与其HA多酚含量呈线性相关。浊度法可用于测定香蕉汁中HA蛋白和HA多酚。     

Influence of (+)-Catechin and Ferulic Acid on Formation of Beer Haze and Their Removal through different Polyvinylpolypyrollidone-Types

The influence of (+)-catechin and ferulic acid on the formation of haze was investigated. Both phenolic compounds were added in different concentrations to beer before ageing through a force test. The removal of these compounds was observed by measuring the difference before and after forcing the beer. (+)-Catechin and ferulic acid showed haze-decreasing as well as haze-increasing properties. The removal of (+)-catechin and ferulic acid from beer through three types of polyvinylpolypyrollidone (PVPP) at several concentrations was investigated. The removal of (+)-catechin and ferulic acid by PVPP-filtration was totally different. The removal of (+)-catechin can be used as a basis to calculate the doses for a desired stability, whereas ferulic acid is removed in irregular amounts.     

国外啤酒混浊及其预测方法

啤酒是一种成分复杂、稳定性不强的胶体溶液,在贮存过程中易产生混浊沉淀现象。啤酒中多种物质均能引起混浊,最觉的是多酚,蛋白质复合引起的混浊。对国外在混浊类型、蛋白多酚混浊、混浊敏感蛋白、混浊敏感多酚、蛋白多酚作用模式、保质期的新型方法６个方面的最新研究进行论述和介绍。     

THE USE OF 14C-LABELLED POLYPHENOLS TO STUDY HAZE FORMATION IN BEER

Haze formation in beer is claimed to involve the gradual polymerization of polyphenols and their subsequent reaction with proteins to form insoluble complexes. The mechanism of haze formation has now been studied using ¹⁴C-labelled epicatechin and dimeric catechin. Epicatechin does not polymerize to form dimeric or trimeric polyphenols when beer is stored and is only incorporated into beer haze to a small extent. Some of the epicatechin combines with nitrogen-containing compounds during storage to form soluble complexes. However, dimeric catechin does not form soluble complexes with nitrogen-containing compounds and instead there is a substantial incorporation of this dimer into beer haze. Dimeric polyphenols are important haze precursors which form insoluble complexes with one or more of the polypeptides of beer, probably by an oxidative coupling mechanism.