Insight into konjac glucomannan-retarding deterioration of steamed bread during frozen storage: Quality characteristics, water status, multi-scale structure, and flavor compounds 了解魔芋葡甘露聚糖在冷冻贮藏过程中的延缓劣变质:品质特性、水分状态、多尺度结构和风味化合物
Fen Zhao , Jinying Guo , Gege Zhang , Lantian Zhang 赵芬 , 郭金英 , 张 格格 , 张 蓝天 College of Food and Bioengineering, Henan University of Science and Technology, Luoyang 471023, PR China 河南科技大学食品与生物工程学院, 中国 洛阳 471023 Hebei Food Safety Key Laboratory, Key Laboratory of Special Food Supervision Technology for State Market Regulation, Hebei Engineering Research Center for Special 河北省食品安全重点实验室, 国家市场监管专用食品监管技术重点实验室, 河北省特种食品工程研究中心Food Safety and Health, Hebei Food Inspection and Research Institute, Shijiazhuang 050227, PR China 河北省食品检验研究院, 中国石家庄市 050227 食品安全与健康
A B S T R A C T Konjac glucomannan (KGM), a water-soluble hydrocolloid, holds considerable potential in the food industry, especially for improving the quality and nutritional properties of frozen products. This study explored the alleviative effect of KGM on the quality characteristics, water status, multi-scale structure, and flavor compounds of steamed bread throughout frozen storage. KGM significantly improved the quality of steamed bread by slowing down the decrease in water content and the increase in water migration while maintaining softness and taste during frozen storage. Notably, KGM also delayed amylopectin retrogradation and starch recrystallization, thus preserving the texture and structure of the steamed bread. At week 3, the microstructure of the steamed bread with KGM remained intact, with the lowest free sulfhydryl content. Additionally, heat map analysis revealed that KGM contributed to flavor retention in steamed bread frozen for 3 weeks. These results indicate that KGM holds promise as an effective cryoprotectant for improving the quality of frozen steamed bread. A B S T R A C T 魔芋葡甘露聚糖 (KGM) 是一种水溶性亲水胶体,在食品工业中具有相当大的潜力,特别是在提高冷冻产品的质量和营养特性方面。本研究探讨了 KGM 对馒头整个冷冻贮藏过程中品质特性、水分状态、多尺度结构和风味化合物的缓解作用。KGM 通过减缓水分含量的降低和水迁移的增加,同时在冷冻储存过程中保持柔软度和味道,显着提高了馒头的品质。值得注意的是,KGM 还延迟了支链淀粉的回生和淀粉重结晶,从而保留了馒头的质地和结构。第 3 周,KGM 馒头的微观结构保持完整,游离巯基含量最低。此外,热图分析显示,KGM 有助于冷冻 3 周的馒头保持风味。这些结果表明,KGM 有望成为提高冷冻馒头质量的有效冷冻保护剂。
1. Introduction 1. 引言
Steamed bread holds a special place as a cherished fermented staple across various cultures, offering a multitude of health benefits. Beyond simply satisfying hunger, it supports gastrointestinal health and aids digestion, facilitating the absorption of vital nutrients such as calcium, magnesium, and iron. In response to evolving consumer preferences, functional variations of steamed bread fortified with probiotics and dietary fiber have emerged. However, despite its nutritional richness, traditional steamed bread faces challenges in large-scale industrial production due to its perishable nature, stemming from its high water content (Qian, Gu, Sun, & Wang, 2021). Balancing the preservation of its nutritional value with extending its shelf life presents a complex challenge for manufacturers. As such, innovative solutions are continually sought to meet consumer demand for both healthy and convenient food options. 馒头在各种文化中作为珍贵的发酵主食,具有特殊的地位,具有多种健康益处。除了满足饥饿感外,它还支持胃肠道健康并帮助消化,促进钙、镁和铁等重要营养物质的吸收。为了响应不断变化的消费者偏好,出现了用益生菌和膳食纤维强化的馒头的功能变化。然而,尽管营养丰富,传统的蒸面包由于其高含水量而易腐烂,在大规模工业生产中面临挑战,这源于其高水分含量(Qian, Gu, Sun, & Wang, 2021)。在保持其营养价值与延长其保质期之间取得平衡,对制造商来说是一个复杂的挑战。因此,不断寻求创新解决方案,以满足消费者对健康和方便食品选择的需求。
To address this challenge, freezing technology has been embraced to manufacture frozen products. This innovation has elevated frozen steamed bread to a prominent position among supermarket offerings due to its extended shelf life, convenience, and adherence to stringent safety standards (Zhu, 2021; Qian et al., 2021). However, freezing and frozen storage can lead to quality deterioration in frozen steamed bread, manifesting as increased hardness, reduced specific volume, and damage to its microstructure (Qian et al., 2022; Gao, Zeng, Qin, Zeng, & Wang, 2023). Frozen storage significantly altered the aggregation and structural properties of heat-denatured gluten proteins in steamed bread (Qian et al., 2021). Another study reported that amylopectin extracted from frozen steamed bread exhibited decreased fractions of B1 and B2 chains, while the long B3 chains and average chain length increased (Deng, Zhang, Ren, Song, & Zhao, 2023). Additionally, the addition of gluten alleviated the deterioration of specific volume, hardness, and microstructure qualities of fast-frozen steamed bread during frozen storage (Qian et al., 2022). The decline of steamed bread quality is primarily due to alterations in water content, starch properties, and gluten networks throughout frozen storage (Qian et al., 2022; Deng et al., 2023). Water undergoes migration and redistribution within the dough, while recrystallization of ice crystals disrupts the starch-gluten matrix (Gao et al., 2023). Moreover, starch degradation and recrystallization of branched starch further contribute to these changes (Kou et al., 2019). To maintain a competitive edge in the market, continual 为了应对这一挑战,冷冻技术已被用于制造冷冻产品。这项创新使冷冻馒头在超市产品中占据了突出地位,因为它延长了保质期、方便并遵守严格的安全标准(Zhu,2021 年;Qian et al., 2021)。然而,冷冻和冷冻储存会导致冷冻馒头的质量下降,表现为硬度增加、比容降低和微观结构受损(Qian et al., 2022;Gao, Zeng, Qin, Zeng, & Wang, 2023)。冷冻储存显着改变了馒头中热变性面筋蛋白的聚集和结构特性(Qian et al., 2021)。另一项研究报告称,从冷冻馒头中提取的支链淀粉表现出 B1 和 B2 链的分数减少,而长 B3 链和平均链长增加(邓、张、任、宋和赵,2023 年)。此外,面筋的添加缓解了速冻馒头在冷冻储存过程中比体积、硬度和微观结构品质的恶化(Qian et al., 2022)。馒头质量的下降主要是由于整个冷冻储存过程中水分含量、淀粉特性和面筋网络的变化(Qian 等人,2022 年;邓等人,2023 年)。水在面团内发生迁移和重新分布,而冰晶的再结晶破坏了淀粉-面筋基质(Gao等人,2023 年)。此外,淀粉降解和支链淀粉的再结晶进一步促进了这些变化(Kou et al., 2019)。为了在市场上保持竞争优势,持续
improvement of frozen steamed bread quality is essential to meeting the evolving expectations of consumers. 提高冷冻馒头的质量对于满足消费者不断变化的期望至关重要。
Significant research efforts have been dedicated to enhancing frozen product quality in recent years. Hydrocolloids have emerged as a promising solution, effectively mitigating the deterioration of frozen dough and its resultant product (Tekle, Ozulku, Bekiroglu, & Sagdic, 2023; He et al., 2020). Their notable water-holding capacity plays a crucial role in inhibiting water migration within frozen flour products during storage. This is achieved by strengthening the coactions between starch and gluten granules while simultaneously reducing the water activity of the products by competing for water with proteins and starch. Studies have demonstrated that guar gum (Hejrani, Sheikholeslami, Mortazavi, & Davoodi, 2017), carboxymethyl cellulose sodium (Xin, Nie, Chen, Li, & Li, 2018), and chitooligosaccharides (Jiang, Guo, Xing, & Zhu, 2023) can enhance the specific volume of frozen dough products while reducing their hardness. In one study, hydroxypropyl methyl cellulose stabilized the gluten network, thereby inhabiting the transition from unfreezable to freezable water (Xuan et al., 2017). In summary, these investigations collectively underscore the robust protective effects of these hydrocolloids on frozen dough. Their primary mechanisms involve mitigating the impact of frozen treatments on water mobility and stabilizing the structure of the gluten network. 近年来,人们致力于提高冷冻产品质量的大量研究工作。亲水胶体已成为一种有前途的解决方案,有效地减轻了冷冻面团及其结果产品的恶化(Tekle, Ozulku, Bekiroglu, & Sagdic, 2023;He et al., 2020)。它们显着的持水能力在抑制冷冻面粉产品在储存过程中的水分迁移方面起着至关重要的作用。这是通过加强淀粉和面筋颗粒之间的相互作用来实现的,同时通过与蛋白质和淀粉竞争水分来降低产品的水分活度。研究表明,瓜尔豆胶(Hejrani、Sheikholeslami、Mortazavi 和 Davoodi,2017 年)、羧甲基纤维素钠(Xin、Nie、Chen、Li 和 Li,2018 年)和壳寡糖(江、郭、邢和朱,2023 年)可以提高冷冻面团产品的比体积,同时降低其硬度。在一项研究中,羟丙基甲基纤维素稳定了面筋网络,从而存在于从不可冻结到可冻结水的过渡中(Xuan et al., 2017)。总之,这些研究共同强调了这些亲水胶体对冷冻面团的强大保护作用。它们的主要机制包括减轻冷冻处理对水流动性的影响和稳定面筋网络的结构。
Konjac glucomannan (KGM) is a primary constituent of hemicellulose found in its cell walls (Ye, Zongo, Shah, Li, & Li, 2021). This heteropolysaccharide consists of D-glucose and D-mannose units, characterized by a distinctive hydrogen bond network, spiral structure, and topological arrangement (Shen & Li, 2022; He et al., 2020). KGM has been approved as a food additive by the U.S. Food and Drug Administration and has been recognized as safe for consumption since 1994 (Behera & Ray, 2017). While KGM is not digestible by human enzymes, it can be fermented by specific gut bacteria, particularly bifidobacteria and lactobacilli. These bacteria break down KGM into shortchain fatty acids, such as butyrate, propionate, and acetate, which are beneficial for gut health (Devaraj, Reddy, & Xu, 2019). KGM has also been shown to have protective effects against colitis by regulating the intestinal immune response and modulating the gut microbiota (Xia et al., 2023; Li, Liu, Fu, Yang, & Wu, 2024). Beyond its health benefits, KGM also enhances the quality of steamed bread. In one study, KGM significantly increased specific volume, improved whiteness, and reduced the hardness of steamed bread by influencing the state and distribution of water in frozen dough (He et al., 2020). Additionally, the incorporation of KGM not only improved the sensory qualities of steamed bread but also provided a valuable source of dietary fiber (Guo et al., 2022). Moreover, adding 0.8 % KGM dramatically reduced the staling rate by delaying the increase in hardness during storage (Sim, Aziah, & Cheng, 2011). In our research, KGM increased the bound water (He et al., 2020) and enhanced the structural stability of dough (Guo et al., 2022). Additionally, it aided in preserving the integrity of wheat starch (Guo, Wang, Liu, & Wang, 2021b), improving thermally stable properties of frozen gluten and gliadin (Guo et al., 2021a). These findings suggest that KGM holds promise for enhancing the quality of wheatbased foods. Building upon this foundation, we hypothesize that KGM may alter water distribution and improve protein and starch properties in frozen steamed bread, thereby augmenting its overall quality. 魔芋葡甘露聚糖(KGM)是其细胞壁中发现的半纤维素的主要成分(Ye, Zongo, Shah, Li, & Li, 2021)。这种异多糖由 D-葡萄糖和 D-甘露糖单元组成,其特征是独特的氢键网络、螺旋结构和拓扑排列(Shen & Li,2022 年;He et al., 2020)。KGM已被美国食品和药物管理局批准为食品添加剂,并且自1994年以来一直被认为是安全的食用产品(Behera & Ray,2017)。虽然 KGM 不能被人类酶消化,但它可以被特定的肠道细菌发酵,特别是双歧杆菌和乳酸杆菌。这些细菌将KGM分解成短链脂肪酸,如丁酸盐、丙酸盐和乙酸盐,这些对肠道健康有益(Devaraj, Reddy, & Xu, 2019)。KGM 还被证明通过调节肠道免疫反应和调节肠道微生物群对结肠炎具有保护作用(Xia et al., 2023;Li, Liu, Fu, Yang, & Wu, 2024)。除了健康益处外,KGM 还可以提高馒头的品质。在一项研究中,KGM 通过影响冷冻面团中水分的状态和分布,显着增加比容、提高白度并降低馒头的硬度(He et al., 2020)。此外,KGM 的加入不仅改善了馒头的感官品质,而且还提供了宝贵的膳食纤维来源(Guo et al., 2022)。此外,添加0.8% KGM可以显著降低老化率,从而延迟储存过程中硬度的增加(Sim, Aziah, & Cheng, 2011)。在我们的研究中,KGM 增加了结合水(He et al., 2020)并增强了面团的结构稳定性(Guo et al., 2022)。 此外,它还有助于保持小麦淀粉的完整性(Guo, Wang, Liu, & Wang, 2021b),改善冷冻面筋和醇溶蛋白的热稳定性(Guo et al., 2021a)。这些发现表明,KGM 有望提高小麦食品的质量。在此基础上,我们假设 KGM 可能会改变冷冻馒头中的水分分布并改善蛋白质和淀粉特性,从而提高其整体质量。
Up to now, research has primarily concentrated on evaluating the influence of KGM on the attributes of frozen dough and the resulting steamed bread quality. However, there remains a gap in understanding how KGM comprehensively influences the frozen storage characteristics of steamed bread. This study aims to bridge this gap by investigating the impact of KGM on various quality parameters of steamed bread, including water content, starch retrogradation, and overall quality characteristics. Additionally, we examine the microstructural changes in steamed bread incorporating KGM during frozen storage. Furthermore, PCA analysis is performed to elucidate the relationship between KGM and the behavior of steamed bread throughout frozen storage. These findings not only provide a deeper understanding of KGM's role in frozen steamed bread but also highlight its potential applications in the industrial manufacture of frozen steamed bread. 到目前为止,研究主要集中在评估 KGM 对冷冻面团属性和由此产生的馒头质量的影响。然而,在理解 KGM 如何全面影响馒头的冷冻储存特性方面仍然存在差距。本研究旨在通过调查 KGM 对馒头各种质量参数的影响来弥合这一差距,包括含水量、淀粉回生和整体质量特性。此外,我们研究了冷冻储存过程中含有 KGM 的馒头的微观结构变化。此外,进行 PCA 分析以阐明 KGM 与馒头在整个冷冻储存过程中的行为之间的关系。这些发现不仅提供了对 KGM 在冷冻馒头中的作用的更深入理解,还突出了它在冷冻馒头工业制造中的潜在应用。
2. Material and methods 2. 材料和方法
2.1. Material 2.1. 材料
Plain wheat flour with a composition of water, 100 g ash, protein, and starch (dry basis) was supplied by Xiangnian Flour Co., Ltd. (Nanyang, Henan, China). Konjac glucomannan (Model: KJ36, viscosity 36,000 Pas, and purity 95 %) was obtained from Hubei Jansen Konjac Technology Co., Ltd. (Wuhan, Hubei, China). Active dry yeast was presented from Angel Yeast Co., Ltd. (Yichang, Hubei, China). Glycine, trichloroacetic acid, and mercaptoethanol were obtained from Aladdin Biotechnology Co., Ltd. (Xi'an, Shaanxi, China). All chemicals were of analytical grade in the study. 由 水、 100 克灰分、 蛋白质和 淀粉(干基)组成的普通小麦粉由湘年面粉有限公司(中国河南省南阳市)提供。魔芋葡甘露聚糖(型号:KJ36,粘度 36,000 Pas,纯度 95%)购自湖北詹森魔芋科技有限公司(中国湖北省武汉市)。活性干酵母由 Angel Yeast Co., Ltd.(中国湖北省宜昌市)推出。甘氨酸、三氯乙酸和巯基乙醇购自阿拉丁生物科技有限公司(中国陕西省习安市)。在研究中,所有化学品均属于分析级。
2.2. Farinograph properties 2.2. Farinograph 属性
The flour ( 300 g ) was substituted with varying proportions of KGM ( , and w/w), determined through preliminary trials. The resulting blends were subjected to farinograph analysis using a model JFZD Farinograph-E (Dong Fu Jiu Cheng Instrument Technology Co., Ltd., Beijing, China) following the standard AACC Method (AACC, 2010a). Water was added incrementally until the dough reached its maximum consistency of . The water absorption, dough development time, and dough stabilization time were obtained. Three replications of the test were performed. 面粉 ( 300 g ) 用不同比例的 KGM ( 和 w/w) 代替,通过初步试验确定。按照标准 AACC 方法 (AACC, 2010a),使用模型 JFZD Farinograph-E(东福久程仪表科技有限公司,中国北京)对所得混合物进行粉蛋白分析。逐渐加水,直到面团达到其最大稠度 。获得吸水率、面团发展时间和面团稳定时间。进行了三次重复测试。
2.3. Preparation of samples 2.3. 样品的制备
The foundational recipe for the steamed bread comprised 100 g of wheat flour, 0.8 g of dry yeast, and deionized water. Based on the water absorption, , and 62.0 mL of water were added to the flour with , and KGM content, respectively. Before mixing, the dry yeast was dispersed in distilled water and activated at for 4 min . All ingredients in each formulation were combined in a mode HM740 kneader (Qingdao Hanshang Electric Co., Ltd., Qingdao, Shandong, China) at 50 rpm for 4 min and at 100 rpm for 5 min until gluten formation was complete. The formed dough was cut into 50 g pieces, molded, and proofed in a model FJX-16 fermentation bin (Demashi Instrument Co., Ltd., Guangzhou, Guangdong, China) at for 45 min . Next, the primary fermented dough was deflated, reshaped, and allowed to ferment for an additional 15 min . Finally, the secondary fermented dough was steamed for 18 min and cooled at for 1 h . One portion of the steamed bread was directly analyzed, while the other was placed in self-sealing bags and rapidly cooled for 1 h in a refrigerator at (Qingdao Haier Zhijia Co., Ltd., Qingdao, Shandong, China). Subsequently, samples were frozen at in a model BCD-218S DA refrigerator (Qingdao Haier Co., Ltd., Qingdao, Shandong, China) for 1, 2, and 3 weeks, respectively. At designated intervals during frozen storage, one portion of the frozen samples was thawed at for 2 h , and the other was re-steamed for sensory evaluation. The sample without KGM was the control. 馒头的基本配方包括 100 克小麦粉、0.8 克干酵母和去离子水。根据吸水率, 向面粉中加入 和 62.0 mL 水,分别含有 和 KGM 含量。混合前,将干酵母分散在蒸馏水中并在 4 min 下活化 。将每种制剂中的所有成分以 HM740 捏合机(青岛汉尚电气有限公司,中国山东青岛)以 50 rpm 混合 4 分钟,并以 100 rpm 混合 5 分钟,直至面筋形成完成。将成型的面团切成 50 克小块,成型,并在 FJX-16 型发酵仓(Demashi Instrument Co., Ltd.,中国广东广州) 中醒发 45 分钟。接下来,将初级发酵面团放气、重塑并再发酵 15 分钟。最后,将二次发酵的面团蒸 18 min,冷却 1 h。直接分析馒头的一部分,而另一部分放入自密封袋中,并在冰箱中快速冷却 1 小时 (青岛海尔智佳有限公司,中国山东省青岛市)。随后,将样品 在 BCD-218S DA 型冰箱(青岛海尔股份有限公司,中国山东青岛)中分别冷冻 1、2 和 3 周。在冷冻贮藏期间,按指定时间间隔将 一部分冷冻样品解冻 2 h,另一部分重新蒸煮进行感官评价。没有 KGM 的样品是对照。
2.4. Quality characteristics 2.4. 质量特性
2.4.1. Textural properties 2.4.1. 纹理属性
The texture of steamed bread was examined as previously reported (Duan et al.,2024). Samples ( ) were extracted from steamed bread and meticulously wrapped in cling film to prevent water loss. Each sample was placed on the glass plate and evaluated by a model TA. XT Express Texture Analyzer (Stable Micro Systems, London, UK) with a probe (P/36R). The pre-test and test speeds were , the compression degree was , the trigger force was 0.05 N , and the post- 如前所述检查了馒头的质地(Duan et al.,2024)。从馒头中提取样品 ( ),并用保鲜膜精心包裹以防止水分流失。将每个样品放在玻璃板上,并通过 TA 模型进行评估。带探头 (P/36R) 的 XT Express 质构仪(Stable Micro Systems,英国伦敦)。前测速和测试速度为 ,压缩度为 ,触发力为 0.05 N ,后测速为
test speed was . The hardness, chewiness, springiness, and resilience were recorded. Three replications of the test were performed. 测试速度为 。记录了硬度、咀嚼性、弹性和弹性。进行了三次重复测试。
2.4.2. Cell structure 2.4.2. 细胞结构
The steamed bread was cut into 12 mm thick slices and photographed for documentation. The steamed bread images were transformed to 8-bit greyscale by Image J software (NIH, Bethesda, USA) to calculate cell numbers and average gas cell sizes (Lu, Xing, Yang, Guo, & Zhu, 2021). 将蒸面包切成 12 毫米厚的薄片并拍照以备记录。通过 Image J 软件(NIH,Bethesda,USA)将馒头图像转换为 8 位灰度,以计算细胞数量和平均气体细胞大小(Lu、Xing、Yang、Guo 和 Zhu,2021 年)。
2.4.3. Specific volume 2.4.3. 比容
The specific volume of steamed bread frozen for 0 and 3 weeks was carried out following the previous method (Li, Guo, Lu, Yue, & Wang, 2024) and calculated by dividing the volume by the mass ( g ). The experiment was replicated three times. 按照前一种方法(Li, Guo, Lu, Yue, & Wang, 2024)进行冷冻0周和3周的蒸面包的特定体积,并通过将体积 除以质量(g)来计算。该实验重复了 3 次。
2.4.4. Sensory evaluation 2.4.4. 感官评价
The sensory evaluation of steamed bread was performed by 20 trained assessors from Henan University of Science and Technology, following the method (Zhao et al., 2021) with some modifications. Before sensory assessment, the samples were re-steamed for 10 min . During evaluation, both whole steamed bread and approximately 10 g portions ( ) were presented to the panelists on disposable white plates, each labeled with a random 3-digit code to ensure anonymity. Samples were presented in a random order for duplicate quantification, and panelists individually scored various sensory attributes on a scale from 0 to 10 , utilizing predefined criteria standards (Table S1). To minimize the cross-sample influence, a oneminute interval was observed between tastings, and panelists were provided with filtered water to cleanse their palates. All evaluations took place in sensory laboratories maintained at suitable temperatures. The total score comprised assessments across five aspects: appearance, structure, toughness, stickiness, and aroma. 馒头的感官评价由河南科技大学的 20 名训练有素的评估员按照该方法(Zhao et al., 2021)进行,并进行了一些修改。在感官评估之前,将样品重新蒸 10 分钟。在评估过程中,整个馒头和大约 10 克的份量 ( ) 都放在一次性白色盘子上呈现给小组成员,每个盘子都标有随机的 3 位代码以确保匿名。样品以随机顺序呈现以进行重复量化,小组成员使用预定义的标准标准(表 S1)以 0 到 10 的等级对各种感官属性进行单独评分。为了尽量减少交叉样品的影响,在品酒之间间隔一分钟,并为小组成员提供过滤水以清洁他们的味觉。所有评估均在保持适当温度的感官实验室中进行。总分包括外观、结构、韧性、粘性和香气五个方面的评估。
2.5. Water status 2.5. 水状态
2.5.1. Water content 2.5.1. 含水量
About 1.5 g of sample was cut from steamed bread, placed on empty plates, and dried in a model DHG-9040C drying oven (Hangzhou Blue Sky Instrument Co., Ltd., Hangzhou, Zhejiang, China) at until a constant weight was reached, following the standard method (AACC, 2010b). The water content ( , w/w) was calculated by dividing the mass change ( g ) of the sample after drying by the mass of the sample before drying. The average value for the three subsamples was recorded. 从馒头上切下约 1.5 g 样品,放在空盘子上,并在 DHG-9040C 型干燥箱(杭州蓝天仪器有限公司,中国浙江省杭州市)中干燥, 直至达到恒定重量,遵循标准方法(AACC,2010b)。水分含量 ( , w/w) 的计算方法是将干燥后样品的质量变化 ( g ) 除以干燥前的样品质量 。记录 3 个子样本的平均值。
2.5.2. Water distribution 2.5.2. 配水
The water distribution within steamed bread was assessed as previously described (He et al., 2020). About 1 g of sample was cut from steamed bread, carefully placed into an NMR bottle, and tightly compacted to remove any voids. Subsequently, the NMR bottle containing the sample was inserted into NMR test tubes and analyzed using a model NM120-015V-1 low-field NMR spectrometer (Neumay Electronic Technology Co., Ltd., Shanghai, China). The relaxation time was measured by specific parameters: a frequency of 200 kHz , an interval time of 3500 ms , and a cumulative number of times of 16 . The percentages of bound water, weakly bound water, and free water were recorded. The average of three measurements was taken. 如前所述评估了馒头内的水分分布(He et al., 2020)。从蒸面包上切下约 1 g 样品,小心地放入 NMR 瓶中,并紧密压实以去除任何空隙。随后,将装有样品的 NMR 瓶插入 NMR 试管中,并使用 NM120-015V-1 型低场 NMR 波谱仪(Neumay Electronic Technology Co., Ltd.,中国上海)进行分析。弛豫时间由特定参数测量:频率 200 kHz,间隔时间为 3500 ms,累积次数为 16 次。记录结合水、弱结合水和游离水的百分比。取了三次测量的平均值。
2.6. X-ray diffraction 2.6. X 射线衍射
X-ray tests were conducted using a model D-8 Advance diffractometer (Bruker Corporation, Billerica, USA) with 40 mA current and 40 kV voltage (Lalush, Bar, Zakaria, Eichler, & Shimoni, 2005). The steamed bread was first lyophilized, ground, and sifted through a 200-mesh sieve to obtain a fine powder. Steamed bread powder was carefully placed into the measurement cell, leveled using a slide, and then subjected to X- ray analysis over a range of . The X-ray diffraction was analyzed using MDI Jade 6.0 software (Materials Data Inc., Livermore, California, USA). 使用D-8型高级衍射仪(Bruker Corporation,Billerica,USA)进行X射线测试,电流为40 mA,电压为40 kV(Lalush,Bar,Zakaria,Eichler和Shimoni,2005年)。首先将馒头冻干、磨碎并通过 200 目筛子过筛以获得细粉。将蒸面包粉小心地放入测量池中,使用载玻片调平,然后在 . 使用 MDI Jade 6.0 软件(Materials Data Inc.,Livermore,California,USA)分析 X 射线衍射。
The infrared spectra of steamed bread were recorded as previously described (Pan et al., 2019). The KBr was combined with steamed bread powder in a mortar at a ratio of , ground for 10 min , and then compressed by a tablet press at 16 MPa pressure for 60 s to produce transparent tablets. The peak spectra of the steamed bread powder were analyzed utilizing a model VERTEX70 FT-IR spectroscopy (Bruker Corporation, Lucken, Germany) with 64 scans at a resolution of . The and 995/1022 were performed by OMNIC software (version 8.2, Thermo Nicolet Corp.). 如前所述记录馒头的红外光谱(Pan等人,2019 年)。将 KBr 与蒸面包粉在研钵中按比例混合 ,研磨 10 min,然后用压片机在 16 MPa 压力下压制 60 s,制成透明片剂。使用 FT-IR 光谱模型(Bruker Corporation,Lucken,Germany)分析蒸面包粉的峰光谱,VERTEX70 64 次扫描,分辨率为 。和 995/1022 由 OMNIC 软件(8.2 版,Thermo Nicolet Corp.)执行。
2.8. Free sulfhydryl content 2.8. 游离巯基含量
The free sulfhydryl content of samples was analyzed as previously described with minor modifications (Guan, Zhang, Wu, Yang, & Bian, 2023). Approximately 75 mg of steamed bread and 4.7 g of guanidine hydrochloride were added to a 1 mL Tris-glycine buffer ( Tris, 6.9 g glycine, 1.2 g EDTA) and fixed to 10 mL with buffer solution. Subsequently, 2 mL of the prepared solution, 1 mL of Ellman's reagent, and 8 mL of urea-guanidine hydrochloride solution were mixed. This mixture was reacted for 1 h at . Next, the sample solution was transferred to a cuvette, and the absorbance of the sample was measured at 412 nm by a model UV-2600 spectrophotometer (Shimadzu Corporation, Kyoto, Japan). Free sulfhydryl content ( ) was calculated using Eq. (1): 如前所述,对样品的游离巯基含量进行了分析,并进行了微小的修改(Guan, Zhang, Wu, Yang, & Bian, 2023)。将大约 75 mg 馒头和 4.7 g 盐酸胍加入 1 mL Tris-甘氨酸缓冲液( Tris、6.9 g 甘氨酸、1.2 g EDTA)中,并用缓冲溶液固定至 10 mL。随后,将 2 mL 制备的溶液、1 mL Ellman 试剂和 8 mL 尿素-盐酸胍溶液混合。该混合物在 中反应 1 小时。接下来,将样品溶液转移至比色皿中,并通过 UV-2600 型分光光度计(Shimadzu Corporation,日本京都)在 412 nm 处测量样品的吸光度。游离巯基含量 ( ) 使用方程 (1) 计算:
Where, 73.53 is the extinction coefficient; is the absorbance at is the dilution factor; is the sample concentration ( ). 其中,73.53 是消光系数; 是吸光度 是稀释因子; 是样品浓度 ( )。
2.9. Microstructure 2.9. 微观结构
The steamed bread sample was fractured into pieces 1 cm ) to expose the interior structure after being frozen for 0 or 3 weeks (Qian et al., 2022). These fractured pieces were then dehydrated using a model LGJ-10D freeze dryer (Sihuan Scientific Instrument Co., Ltd., Beijing, China). Subsequently, the dehydrated samples were sprayed with gold about 12 nm in thickness using a model EM ACE200 gold sprayer (Leica Camera Corporation, Hesse, Germany). The microstructure of samples was measured by a model TM3030 Plus SEM (Hitachi Corporation, Mito, Japan) with magnifications of and , respectively. 将馒头样品冷冻 0 或 3 周后碎成 1 厘米的碎片 ),露出内部结构(Qian et al., 2022)。然后使用 LGJ-10D 型冷冻干燥机(四环科学仪器有限公司,中国北京)对这些断裂的碎片进行脱水。随后,使用 EM ACE12 型金喷雾器(徕卡相机公司,黑森州,德国)用约 200 nm 厚的金喷涂脱水样品。样品的微观结构由型号 TM3030 Plus SEM (Hitachi Corporation, Mito, Japan) 测量,放大倍数分别为 和 。
2.10. Volatile flavor compounds 2.10. 挥发性风味化合物
The volatile compounds of the samples were measured using a model TSQ 9000 gas chromatography-mass spectrometry (Thermo Scientific, Massachusetts, USA) as previously described (Ma, Mu, & Zhou, 2021) with some modifications. About 2 g of steamed bread and 2-methyl3-heptanone were put in a 20 mL headspace. A Solid Phase Microextraction fiber was inserted into the headspace and extracted volatiles at a water bath of for 55 min . Next, the fiber was promptly put into the injection port of the gas chromatograph to desorb the compounds at for 5 min . Volatiles were separated on an HP-5MS capillary column ( ) with a helium flow rate of . The GC oven temperature was programmed from (hold for 4 min ) to (hold for 12 min ) at a rate of . The transfer line and ion source temperatures were 280 and , respectively. The scan mode was between 35 and . Compound identification was performed by comparing mass spectrometry data with the National Institute of Standards and Technology 如前所述(马、Mu 和 周,2021 年),使用型号 TSQ 9000 气相色谱-质谱法(Thermo Scientific,美国马萨诸塞州)测量样品的挥发性化合物,并进行了一些修改。将约 2 g 馒头和 2-甲基 3-庚酮放入 20 mL 顶部空间。将固相微萃取纤维 头插入顶部空间,并在 55 分钟的水浴 中萃取挥发物。接下来,立即将纤维头放入气相色谱仪的进样口,以 5 min 解吸化合物 。挥发物在 HP-5MS 毛细管柱 ( ) 上分离,氦流速为 。GC 柱温箱温度以 的速率从 (保持 4 分钟) 编程为 (保持 12 分钟)。传输线和离子源温度分别为 280 和 。扫描模式介于 35 和 之间。通过将质谱数据与美国国家标准与技术研究院 (National Institute of Standards and Technology) 进行比较来进行化合物鉴定
database. The content ( ) of volatile flavor compounds was calculated using Eq. (2): 数据库。挥发性风味化合物的含量 ( ) 使用方程 (2) 计算:
Where is the content ( ) of the compound; is the mass concentration of the internal standard content; is the volume ( ) of internal standard injection; is the peak area of the compound; is the peak area of the internal standard substance; is the sample quality . 其中 是化合物的含量 ( ); 是内标含量的质量浓度 ; 是内标进样的体积 ( ); 是化合物的峰面积 ; 是内标物质的峰面积 ; 是样本质量 。
2.11. Statistical analysis 2.11. 统计分析
The results were expressed as means standard deviation. Statistical analysis was performed by a one-way analysis of variance followed by Duncan's multiple range test ( ) and independent-samples -test using SPSS 20.0 software (SPSS Inc., Chicago, IL, USA). Graphs were drawn using Origin Software 2021 (Origin Lab Corp., MA, USA). 结果表示为均值 标准差。通过使用 SPSS 20.0 软件 (SPSS Inc., Chicago, IL, USA) 进行单因素方差分析,然后进行 Duncan 多范围检验 ( ) 和独立样本 检验。图表是使用 Origin Software 2021(Origin Lab Corp.,MA,USA)绘制的。
3. Results and discussion 3. 结果和讨论
3.1. Farinographic properties 3.1. 粉质学性质
Farinographic properties are indeed crucial for evaluating wheat flour quality. Flour's strength and water absorption capacity are key factors in determining its suitability for various applications. These properties provide valuable insights into the behavior of flour during processing, helping manufacturers optimize formulations and processes to achieve desired product attributes. The farinographic properties of flour with KGM are illustrated in Fig. 1. Water absorption (WA) indicates the amount of water needed to achieve the desired dough consistency of 500 FU . The WA of the flour with different KGM additions increased in the order of control (Fig. 1a). At the replacement level of , the WA of the dough increased by compared with the control. This indicates that more water is absorbed in the presence of KGM to achieve the desired dough consistency of 500 FU . This is because KGM, with its polyhydroxy structure, has a higher water-binding capacity than gluten (Sim et al., 2011). In one study, the water absorption of dough increased with the rising substitution levels and viscosity of KGM (Guo et al., 2022). Similarly, another study found that dough containing KGM exhibited a higher level of water absorption compared with the control (Meng et al., 2021). 粉学特性对于评估小麦粉质量确实至关重要。面粉的强度和吸水能力是决定其适用于各种应用的关键因素。这些特性为面粉在加工过程中的行为提供了有价值的见解,帮助制造商优化配方和工艺以实现所需的产品属性。含 KGM 的面粉的粉学特性如图 1 所示。吸水率 (WA) 表示达到所需的面团稠度 500 FU 所需的水量。添加不同 KGM 的面粉的 WA 按 对照顺序增加(图 1a)。在 的更换水平 ,与对照 相比,面团的 WA 增加。这表明在 KGM 存在下吸收了更多的水,以达到所需的 500 FU 面团稠度。这是因为 KGM 具有多羟基结构,比面筋具有更高的水结合能力(Sim et al., 2011)。在一项研究中,面团的吸水率随着 KGM 取代水平和粘度的升高而增加(Guo et al., 2022)。同样,另一项研究发现,与对照组相比,含有 KGM 的面团表现出更高水平的吸水率(Meng et al., 2021)。
Dough development time (DDT) refers to the period from the addition of water until the farinograph curve reaches its peak consistency, marking the time required for water to fully interact with the flour and for the dough to form. The DDT values in wheat-KGM blends were obviously higher than those in the control (Fig. 1b), indicating an extended time to reach maximum dough consistency. This finding implies that KGM molecules compete against gluten for water, and therefore affect the development of gluten network due to the swelling of hydrocolloids (Li et al., 2019). In one study, adding bran increased the DDT in low-gluten flour but decreased it in medium and high-gluten flour (Li et al., 2023). This inconsistency could be partially 面团形成时间 (DDT) 是指从加水到粉质曲线达到其峰值稠度的时间,标志着水与面粉充分相互作用和面团形成所需的时间。小麦-KGM 混合物中的 DDT 值明显 高于对照中的 DDT 值(图 1b),表明达到最大面团稠度的时间延长。这一发现意味着 KGM 分子与面筋竞争水,因此由于亲水胶体的膨胀而影响面筋网络的发展(Li et al., 2019)。在一项研究中,添加麸皮会增加低筋面粉中的 DDT,但会降低中高筋面粉中的 DDT(Li 等人,2023 年)。这种不一致可能是部分的
Fig. 1. Water absorption (a), dough development time (b), and stability time (c) of wheat flours added with different additions of konjac glucomannan (KGM). Different lower case letters represent significant ( ) differences among steamed bread with different KGM additions. 图 1.添加不同添加量魔芋葡甘聚糖 (KGM) 的小麦粉的吸水率 (a)、面团形成时间 (b) 和稳定时间 (c)。不同的小写字母表示添加不同 KGM 的馒头之间的显著 ( ) 差异。
attributed to variations in gluten content within the flour. Another reason might be the diverse starch compositions, which influence the mixing properties of the dough (Li et al., 2021). 归因于面粉中面筋含量的变化。另一个原因可能是淀粉成分的多样性,这会影响面团的混合特性(Li et al., 2021)。
The stability time (ST) characterizes its resistance to kneading, with higher values indicating stronger gluten and more elastic dough ( Yu et al., 2019). Adding KGM dramatically ( ) reduced the ST value in the dough, and this decline in ST corresponded with the escalation in KGM levels (Fig. 1c). This is because KGM causes steric hindrance, dilutes the gluten, or even punctures gas cells, thereby impacting gluten development. Another reason is that KGM decreases the overall rigidity of the dough. 稳定时间 (ST) 表征了其抗揉捏性,较高的值表示面筋更强,面团更有弹性(Yu et al., 2019)。添加 KGM 显着降低了面团中的 ST 值 ( ),ST 的下降与 KGM 水平的升高相对应(图 1c)。这是因为 KGM 会引起空间位阻,稀释面筋,甚至刺穿气室,从而影响面筋的形成。另一个原因是 KGM 降低了面团的整体刚度。
In summary, as the amount of KGM increases, water absorption increases, dough development time initially rises and then declines, while stabilization time decreases in wheat-KGM blends. 综上所述,随着 KGM 量的增加,吸水率增加,面团形成时间最初上升后下降,而小麦-KGM 共混物的稳定时间缩短。
3.2. Specific volume and cell structure 3.2. 比容和细胞结构
High-quality steamed bread is generally associated with a larger specific volume and a more uniform cell structure, which directly influences product shape and consumer preferences. The impact of KGM on specific volume of steamed bread frozen at 0 and 3 weeks is presented in Table 1. At week 3, the specific volume of samples significantly ( 0.05 ) increased and then displayed a slight decrease as the KGM increased. The highest specific volume, which was higher than the control, was achieved with the KGM steamed bread. This increase in specific volume is related to KGM's interaction with gluten, leading to better gas retention in the gluten network (Guo et al., 2022). Meanwhile, the enhanced gas retention capacity also leads to smaller average gas cell sizes and higher cell numbers in samples, resulting in a more uniform cell structure (Table 1). 高质量的馒头通常与更大的比容和更均匀的细胞结构有关,这直接影响产品形状和消费者偏好。KGM 对 0 周和 3 周冷冻馒头比体积的影响如表 1 所示。第 3 周时,样品比体积显著增加 ( 0.05 ),然后随着 KGM 的增加而略有减少。最高的比体积 高于对照,使用 KGM 馒头实现。这种比容的增加与 KGM 与面筋的相互作用有关,导致面筋网络中更好的气体保留(Guo et al., 2022)。同时,增强的气体保留能力还导致样品中的平均气体池尺寸更小,细胞数量更高,从而获得更均匀的细胞结构(表 1)。
Here, the pore structure of the KGM steamed bread was similar to that of the control, with large pores and a non-uniform size and distribution (Fig. 2). In contrast, steamed bread with and KGM additions exhibited no obvious large pores, and their internal structure displayed more homogeneous and dense pores compared with the control. The reason might be that KGM helps to form the desired starchgluten matrix, physically stabilizing gas cells and ensuring homogeneous expansion of air cells (Janssen, Wouters, & Delcour, 2021; Škara, Novotni, Čukelj, Smerdel, & Ćuri, 2013). At a molecular level, the mechanism through which KGM controls the stability of gas cells reduces the likelihood of coalescence among individual gas cells by forming a liquid film at the gas-liquid interface (Janssen et al., 2021; Sroan, Bean, & MacRitchie, 2009). The size and density of the gas cells can lead to significant differences in the texture and sensory characteristics of the end product (Turbin-Orger et al., 2012). 在这里, KGM 馒头的孔结构与对照相似,具有大孔和不均匀的大小和分布(图 2)。相比之下,添加 KGM 的馒头没有明显的大孔隙,其内部结构与对照相比表现出更均匀和致密的孔隙。原因可能是KGM有助于形成所需的淀粉面筋基质,物理稳定气室并确保气室的均匀膨胀(Janssen, Wouters, & Delcour, 2021; Škara, Novotni, Čukelj, Smerdel, & Ćuri, 2013)。在分子水平上,KGM 控制气室稳定性的机制通过在气液界面形成液膜来降低单个气室之间聚结的可能性(Janssen等人,2021 年;Sroan, Bean, & MacRitchie, 2009)。气室的大小和密度会导致最终产品的质地和感官特性出现显着差异(Turbin-Orger et al., 2012)。
During frozen storage, the specific volume significantly decreased in all steamed bread groups due to the damage to the gluten network induced by the formation of ice crystals (Table 1). It may also be related to the destruction of the aggregated and molecularly ordered structure of starch (Gao et al., 2023). Notably, the decrease in a specific volume of , and KGM steamed bread was 0.15 , , and from 0 to 3 weeks, respectively. The decrease in specific volume in all KGM groups was observed to be smaller than that in the control group. The delay in the specific volume decrease might be due to less damage to the gluten network in the presence of KGM during frozen storage. 在冷冻储存过程中,由于冰晶的形成对面筋网络的破坏,所有馒头组的比容都显著降低 (表 1)。这也可能与淀粉聚集和分子有序结构的破坏有关(Gao等人,2023 年)。值得注意的是, 比容 和 KGM 馒头的比容分别减少 0.15 、 和 0 至 3 周。观察到所有 KGM 组的比容下降幅度均小于对照组。比体积减少的延迟可能是由于在冷冻储存过程中 KGM 存在的情况下对面筋网络的损害较小。
Overall, KGM demonstrates positive effects on increasing the specific volume and improving the cell structure of steamed bread. Moreover, KGM delays the decline in the specific volume, suggesting that KGM contributes to the formation or maintenance of a larger specific volume throughout frozen storage. 总体而言,KGM 对增加馒头的比容和改善馒头的细胞结构表现出积极作用。此外,KGM 延缓了比容的下降,表明 KGM 有助于在整个冷冻储存过程中形成或维持更大的比容。
3.3. Texture properties 3.3. 纹理属性
Texture analysis is a useful tool for evaluating the popularity of wheat products, representing vital sensory attributes. Among them, hardness is regarded as the primary indicator of textural properties, and a smaller value of hardness usually indicates that steamed bread is fluffy and more easily accepted by consumers. The influence of KGM on the textural characteristics of steamed bread frozen at different time is shown in Table 2. At the same frozen storage time, the hardness significantly decreased and then slightly increased as the KGM increased. The samples with KGM addition had the lowest hardness ( 777.01 g , week 3), which was reduced by compared with the control. This is consistent with our previous findings that the hardness of steamed bread decreased when KGM was added (Guo et al., 2022; He et al., 2020). This is because KGM alters the conformations of gluten and gliadin, affecting the rearrangement of polypeptide chains' spatial structure by enhancing hydrophobic interactions (He et al., 2020). Additionally, KGM's polyhydroxyl structures may compete with starch and gluten for water to form hydrogen bonds, which can limit starch swelling. This competition may lead to changes in the starchgluten network and result in reduced hardness (Fu et al., 2021). 质构分析是评估小麦产品受欢迎程度的有用工具,代表了重要的感官属性。其中,硬度被视为质地特性的主要指标,硬度值越小,通常表明馒头蓬松,更容易被消费者接受。KGM 对不同时间冷冻的馒头质地特性的影响如表 2 所示。在相同的冷冻贮藏时间内,硬度随着 KGM 的增加而显著降低 后略有增加。添加 KGM 的样品硬度最低 ( 777.01 g,第 3 周),与对照 相比降低。这与我们之前的发现一致,即添加 KGM 后馒头的硬度降低(Guo et al., 2022;He et al., 2020)。这是因为 KGM 改变了面筋和麦醇溶蛋白的构象,通过增强疏水相互作用来影响多肽链空间结构的重排(He et al., 2020)。此外,KGM 的多羟基结构可能与淀粉和面筋竞争水形成氢键,从而限制淀粉溶胀。这种竞争可能导致淀粉面筋网络发生变化并导致硬度降低(Fu et al., 2021)。
Here, the hardness of all steamed bread groups increased as the frozen time increased, further confirming that the specific volume decreased (Section 3.2). The hardness of the control group increased from 758.42 to 951.37 g after being frozen for 3 weeks. This is primarily related to molecular aggregates of amylose (Deng et al., 2023) and the destruction of the gluten network structure (Gao et al., 2023). From 0 to 3 weeks, it is noticeable that the hardness in the control group increased by , while in the , and KGM groups it increased by , and , respectively. This result implies that the steamed bread with KGM addition reduces the growth rate of hardness and delays starch retrogradation. This is because KGM restricts water migration and ice recrystallization by interacting with water molecules to form intermolecular hydrogen bonds, thereby 在这里,所有馒头组的硬度都随着冷冻时间的增加而增加,进一步证实了比容减小(第 3.2 节)。对照组冷冻 3 周后硬度由 758.42 g 增加到 951.37 g。这主要与直链淀粉的分子聚集体(邓等人,2023 年)和面筋网络结构的破坏(Gao等人,2023 年)有关。从 0 到 3 周,很明显,对照组的硬度增加了 ,而在 和 KGM 组中,硬度分别增加了 、 和 。这一结果表明,添加 KGM 的馒头降低了硬度的增长速度并延缓了淀粉的回生。这是因为 KGM 通过与水分子相互作用形成分子间氢键来限制水迁移和冰再结晶,从而
Table 1 表 1
Specific volume, cell parameters, and FT-IR peak ratio characteristics of steamed bread added with different additions of konjac glucomannan (KGM) and frozen for 0 and 3 weeks. 添加不同添加量魔芋葡甘露聚糖 (KGM) 并冷冻 0 和 3 周的馒头的比体积、细胞参数和 FT-IR 峰比特征。
千克甘基姆 (%)
KGM
(%)
比容
(
周)
Specific volume
( weeks)
比容
(
周)
Specific volume
( weeks)
Cell parameters Cell 参数
R1047/1022 (0 周)
R1047/1022
(0 weeks)
R1047/1022 (3 周)
R1047/1022
(3 weeks)
R995/1022 (0 周)
R995/1022 (0
weeks)
R995/1022 (3 周)
R995/1022 (3
weeks)
平均气体池尺寸
Average gas cell
size
每个 单元格数
Number of cells
per
0
0.5
1.5
All data are expressed as mean standard deviation ). Different lower case letters represent significant ( ) differences among steamed bread with different KGM additions at the same frozen time. Different upper case letters indicate significant ( ) differences among steamed bread with the same KGM addition at different frozen time. 所有数据均表示为平均 标准差 )。不同的小写字母表示在同一冷冻时间内添加不同 KGM 的馒头之间的显著 ( ) 差异。不同的大写字母表示相同 KGM 添加量的馒头在不同冷冻时间存在显著 ( ) 差异。
Fig. 2. Profiles and thresholding images of cell structure from 0 -week steamed bread added with different additions of konjac glucomannan (KGM). 图 2.添加不同添加魔芋葡甘露聚糖 (KGM) 的 0 周馒头的细胞结构剖面和阈值图像。
protecting the gluten network structure of samples (Jekle & Becker, 2012; Gao et al., 2023). Thus, adding KGM could alleviate the decline in hardness and delay the staling of steamed bread to a certain extent during frozen storage. Meanwhile, the chewiness of steamed bread showed a similar variation trend to their hardness as the KGM increased and the extension of frozen time. 保护样品的麸质网络结构(Jekle & Becker,2012;Gao等人,2023 年)。因此,添加 KGM 可以在一定程度上缓解馒头在冷冻贮存过程中的硬度下降并延缓馒头的老化。同时,随着 KGM 的增加和冷冻时间的延长,馒头的咀嚼性与其硬度呈相似的变化趋势。
As presented in Table 2, at the same frozen storage time, springiness and resilience decreased slightly as the KGM increased. However, the effects on the springiness and resilience of steamed bread were insignificant ( ), suggesting that KGM does not lead to a deterioration in springiness and resilience. For the control and KGM groups, springiness and resilience both decreased as the frozen time increased. This is primarily attributed to the mechanical damage to the gluten network (Zhu, 2021). Higher chewiness, hardness, and lower resilience indicate a worse quality of steamed bread. 如表 2 所示,在相同的冷冻贮藏时间内,随着 KGM 的增加,弹性和弹性略有下降。然而,对馒头的弹性和弹性的影响微不足道 ( ),表明 KGM 不会导致弹性和弹性的恶化。对照组和 KGM 组的弹性和弹性均随着冻结时间的增加而降低。这主要归因于面筋网络的机械损伤(Zhu,2021 年)。较高的咀嚼性、硬度和较低的弹性表明馒头的质量较差。
Combining the changes observed in specific volume, cell structure (Table 1), and texture (Table 2), we conclude that the addition of KGM results in steamed bread with the maximum specific volume, a more uniform cell structure, and the minimum hardness. This suggests that steamed bread with KGM displays high freeze stability. Furthermore, compared with the control, the increase in hardness of KGM-added samples was lower throughout frozen storage. These findings indicate that KGM delays the deterioration of steamed bread quality, suggesting that the internal network structure is less susceptible to damage by ice crystals. Additionally, it can be inferred that the quality degradation of steamed bread with KGM is alleviated to the greatest extent during frozen storage. 结合观察到的比容、细胞结构(表 1)和质地(表 2)的变化,我们得出结论,添加 KGM 可产生具有最大比容、更均匀的细胞结构和最小硬度的馒头。这表明用 KGM 制成的 馒头表现出很高的冷冻稳定性。此外,与对照相比,在整个冷冻储存过程中,添加 KGM 的样品的硬度增加较低。这些发现表明 KGM 延缓了馒头品质的恶化,表明内部网络结构不易受到冰晶的破坏。此外,可以推断,KGM 蒸煮馒头在冷冻贮藏过程中质量下降得到了最大程度的缓解。
3.4. Sensory evaluation 3.4. 感官评价
The sensory evaluation of steamed bread with KGM addition for different frozen storage time is depicted in Fig. 3. Across all steamed bread groups, sensory evaluation indicators decreased with increasing frozen storage time. Notably, blistering occurred on the surface of steamed bread, visibly reducing its appearance and quality. Additionally, the texture of the frozen sample appeared less homogeneous compared with the unfrozen sample. In addition, the taste and aroma of the frozen, reheated sample were not as satisfactory as those of the unfrozen sample. Except for aroma, sensory evaluation indicators in the KGM groups were generally higher compared with the control group at the same frozen storage time, except for stickiness at 0 weeks and appearance at 1 week. Steamed bread with or KGM addition showed the highest sensory evaluation scores, followed by and . The aroma score of steamed bread decreased at 0 or 1 week with the KGM increase, while the aroma score increased as the KGM increased at 2 or 3 weeks. This suggests that the negative impact of freezing on the aroma of the sample is more significant than the effect of KGM on the aroma. This is because adding KGM protects the gluten network, resulting in the retention of more volatile substances. 图 3 描述了添加 KGM 的蒸包在不同冷冻贮存时间下的感官评价。在所有馒头组中,感官评价指标均随着冷冻贮藏时间的增加而降低。值得注意的是,蒸面包表面出现起泡,明显降低了其外观和质量。此外,与未冷冻样品相比,冷冻样品的质地似乎不那么均匀。此外,冷冻、再加热样品的味道和香气不如未冷冻样品令人满意。除香气外,在相同冷冻贮藏时间下,KGM 组的感官评价指标普遍高于对照组,除 0 周时粘性外,1 周时出现粘性。添加 或 KGM 的馒头感官评价得分最高,其次是 和 。馒头香气评分在 0 或 1 周随着 KGM 的增加而降低,而香气评分在 2 或 3 周随着 KGM 的增加而升高。这表明冷冻对样品香气的负面影响比 KGM 对香气的影响更显着。这是因为添加 KGM 可以保护面筋网络,从而保留更多挥发性物质。
Overall, the presence of KGM mitigates the damage inflicted by 总体而言,KGM 的存在减轻了
Table 2 表 2
Texture of steamed bread added with different additions of konjac glucomannan (KGM) at different frozen time. 在不同冷冻时间添加不同魔芋葡甘露聚糖 (KGM) 的蒸面包质地。
Texture 质地
KGM
Frozen storage time (week) 冷冻储存时间(周)
0
1
2
3
硬度 (g)
Hardness
(g)
0
0.5
1.0
1.5
Chewiness 咀嚼性
0
0.5
1.0
1.5
Springiness 弹性
0
0.5
1.0
1.5
Resilience 达观
0
0.5
1.0
1.5
All data are expressed as mean standard deviation ). Different lower case letters represent significant ( ) differences among steamed bread with different KGM additions at the same frozen time. Different upper case letters indicate significant ( ) differences among steamed bread with the same KGM addition at different frozen time. 所有数据均表示为平均 标准差 )。不同的小写字母表示在同一冷冻时间内添加不同 KGM 的馒头之间的显著 ( ) 差异。不同的大写字母表示相同 KGM 添加量的馒头在不同冷冻时间存在显著 ( ) 差异。
frozen treatment on the main components of steamed bread, thereby preserving its quality characteristics. The addition of or KGM appears to be more favorable for frozen storage, as it helps maintain the sensory qualities and overall integrity of frozen steamed bread. 对馒头的主要成分进行冷冻处理,从而保持其质量特性。添加 或 KGM 似乎更适合冷冻储存,因为它有助于保持冷冻馒头的感官品质和整体完整性。
3.5. Water content 3.5. 含水量
The interplay between water and food components significantly impacts the storage stability of frozen foods. The changes in the water content of steamed bread are presented in Fig. 4a. The water content showed a notable ( ) decrease in all steamed bread groups as the frozen storage time increased. This is attributed to ice recrystallization, which can disrupt the gluten network and further cause the separation of water molecules (Zeng, Shi, Liang, Wang, & Gao, 2023). The resulting damage to starches and gluten networks reduces their ability to hold water, ultimately causing water migration and decrease (Deng et al., 2023). 水和食品成分之间的相互作用会显著影响冷冻食品的储存稳定性。馒头含水量的变化如图 4a 所示。随着冷冻储存时间的增加,所有馒头组的含水量均显著 ( ) 降低。这归因于冰的再结晶,它可以破坏面筋网络并进一步导致水分子的分离(Zeng, Shi, Liang, Wang, & Gao, 2023)。由此对淀粉和面筋网络的损害降低了它们的保水能力,最终导致水迁移和减少(邓等人,2023 年)。
Here, adding KGM significantly ( ) influenced the water content of steamed bread. The water content increased with the rise in KGM content at the same frozen storage time. This suggests that KGM enhances the water absorption capacity of the dough, aligning with Section 3.1. From 0 to 3 weeks, the water content in the control group decreased from to , while the KGM group ranged from to . The reduction in water content in the KGM groups was less than in the control group. This is because the KGM molecule readily forms hydrogen bonds with water, thereby mitigating the decrease in water content. 在这里,添加 KGM ( ) 显著影响了馒头的含水量。在相同的冷冻贮藏时间内,含水量随着 KGM 含量的增加而增加。这表明 KGM 增强了面团的吸水能力,与第 3.1 节一致。从 0 到 3 周,对照组的含水量由 到 下降,而 KGM 组的含水量从 到 不等。KGM 组含水量的降低幅度小于对照组。这是因为 KGM 分子很容易与水形成氢键,从而减轻水分含量的降低。
3.6. Water distribution 3.6. 配水
LF-NMR is applied further to monitor water distribution and migration during frozen storage (Cotrim, Minim, Felix, & Minim, 2020). Three different water populations were named according to their relaxation times from the shortest to the longest: T21 ( ( ), and T23 ( ), representing bound water, weakly bound water, and free water, respectively. Moreover, T22 was the dominant population, comprising about of the total proton number, while T21 accounted for approximately of the total proton number, and T23 was the smallest population, representing only 1-2 %. The H T2 distributions are illustrated in Fig. 4b. When frozen from 0 to 3 weeks, for all steamed bread groups, the T21 and T23 shifted towards shorter (left shift) and the T22 shifted towards longer (right shift), except for the KGM group, possibly due to water redistribution in the amorphous regions of steamed bread. Additionally, the T23 decreased from 151 to 114 ms throughout frozen storage, suggesting a decrease in overall exchangeable proton mobility. LF-NMR进一步应用于监测冷冻储存过程中的水分布和迁移(Cotrim, Minim, Felix, & Minim, 2020)。根据它们的弛豫时间从最短到最长命名三种不同的水种群: T21 ( ( ) 和 T23 ( ),分别代表束缚水、弱束缚水和游离水。此外,T22 是优势种群,约 占总质子数,而 T21 约 占总质子数,T23 是最小的种群,仅占 1-2%。 H T2 分布如图 4b 所示。当冷冻 0 到 3 周时,对于所有馒头组,T21 和 T23 向较短(左移)移动,T22 向较长(右移)移动,除了 KGM 组,这可能是由于水在馒头的无定形区域中的重新分布。此外,在整个冷冻存储过程中,T23 从 151 毫秒下降到 114 毫秒,表明整体可交换质子迁移率降低。
The left shift of T23 confirmed the reduced overall water mobility in the sample. T23 的左移证实了样品中整体水迁移率降低。
A21, A22, and A23 correspond to the proportion of bound water, weakly bound water, and free water, respectively. At the same frozen storage time, a lower A23 was found in the KGM groups compared with the control group (Fig. 4c), indicating that KGM is beneficial in decreasing water fluidity. Several hydrophilic groups existed in KGM molecule, which bound the water in a certain space and reduced the water mobility and the ability of water vapor to escape (He et al., 2020). Meanwhile, A21 showed a descending tendency, while A22 displayed an increasing trend as the KGM increased. This suggests that KGM increases the amount of the weakly bound water in samples due to its high waterholding capacity. A21、A22 和 A23 分别对应于结合水、弱结合水和自由水的比例。在相同的冷冻储存时间内,与对照组相比,KGM 组的 A23 含量较低(图 4c),表明 KGM 有利于降低水流动性。KGM 分子中存在几个亲水基团,它们将水结合在一定空间内,降低了水的流动性和水蒸气的逃逸能力(He et al., 2020)。同时,A21 呈下降趋势,而 A22 则随着 KGM 的增加呈上升趋势。这表明 KGM 由于其高持水能力而增加了样品中弱结合水的量。
Here, as frozen storage time increased, the A21 of all steamed bread groups decreased, while the A22 and A23 increased, indicating that the bound water gradually migrated to weakly bound and free water. It was found that the frozen inevitably led to bound water desorbing from starch-gluten networks. For the control group, A21 decreased from 22.6 % to , and A22 increased from to from 0 to 3 weeks, indicating that water migrates from the gluten domain to the starch domain, in agreement with Curti, Bubici, Carini, Baroni, & Vittadini (2011). However, the A22 increment in the control group was 3.06 % during frozen storage, whereas that in different KGM groups was only . The A22 increment in all KGM groups was less compared with the control group, suggesting that KGM might decrease the binding of water molecules from the gluten to gelatin-starch matrix (Yu et al., 2019; Curti et al., 2011). This is because the hydroxyl groups in KGM interact with water through hydrogen bonding, which reduces the mobility of the adsorbed water via proton exchange (He et al., 2020; Silva et al., 2013). Additionally, the hydroxyl groups on the KGM polysaccharide chain form hydrogen bonds with the hydroxyl groups on the starch chain, thereby decreasing the incorporation of water molecules into the starch domain (Chen, Long, Ruan, & Labuza, 1997; Gómez, Jiménez, Ruiz, & Oliete, 2011). The A21 reduction of the control group was , while that of different KGM groups was 2.49-3.36 % from 0 to 3 weeks. This indicates that the increment in A23 for all KGM groups (except for the KGM group) is less compared with the control group. The reason for these phenomena may be that in the presence of KGM, there is less damage to the gluten protein, resulting in a reduction in water migration (Qian et al., 2022). 在这里,随着冷冻贮藏时间的增加,所有馒头组的 A21 均降低,而 A22 和 A23 增加,表明结合水逐渐迁移到弱结合水和游离水。研究发现,冷冻不可避免地导致结合水从淀粉-面筋网络中解吸。对于对照组,A21从22.6%下降到 ,A22从 0增加到3周,这表明水从面筋领域迁移到淀粉领域,这与Curti, Bubici, Carini, Baroni和Vittadini (2011)的观点一致。然而,对照组在冷冻贮藏期间的 A22 增量为 3.06%,而不同 KGM 组的 A22 增量仅为 。与对照组相比,所有 KGM 组的 A22 增量都较小,这表明 KGM 可能会减少面筋中水分子与明胶淀粉基质的结合(Yu等人,2019 年;Curti et al., 2011)。这是因为 KGM 中的羟基通过氢键与水相互作用,这通过质子交换降低了吸附水的迁移率(He等人,2020 年;Silva et al., 2013)。此外,KGM多糖链上的羟基与淀粉链上的羟基形成氢键,从而减少水分子在淀粉结构域中的结合(Chen, Long, Ruan, & Labuza, 1997;Gómez, Jiménez, Ruiz, & Oliete, 2011)。对照组的 A21 降低为 ,而不同 KGM 组从 0 到 3 周的 A21 降低为 2.49-3.36 %。这表明所有 KGM 组( KGM 组除外)的 A23 增量都小于对照组。 造成这些现象的原因可能是,在 KGM 存在的情况下,对麸质蛋白的损伤较小,导致水迁移减少(Qian et al., 2022)。
In summary, frozen storage may trigger heightened water migration, potentially leading to a deterioration in the sensory attributes and texture of steamed bread. However, the incorporation of KGM 总之,冷冻储存可能会引发水分迁移加剧,可能导致蒸面包的感官特性和质地变差。然而,KGM 的成立
Fig. 3. Sensory evaluation of steamed bread added with different konjac glucomannan (KGM) additions at different frozen time. (a), (b), (c), and (d) represent steamed bread frozen for , and 3 weeks, respectively. The red line, blue line, green line, and purple line are steamed bread added with , and KGM, respectively. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.) 图 3.不同冷冻时间添加不同魔芋葡甘聚糖 (KGM) 的馒头的感官评价。(a)、(b)、(c) 和 (d) 分别表示冷冻 和 3 周的馒头。红线、蓝线、绿线和紫线分别是添加了 和 KGM 的馒头。(有关此图例中对颜色的引用的解释,读者可参考本文的网络版本。
significantly bolsters the interaction between KGM and water, thereby impeding water migration throughout the entire frozen storage duration. Consequently, this attenuates the physical harm inflicted by ice crystals on the gluten network, ultimately enhancing the resilience of steamed bread during frozen storage. 显著加强了 KGM 与水之间的相互作用,从而在整个冷冻储存期间阻碍了水的迁移。因此,这减轻了冰晶对面筋网络造成的物理伤害,最终增强了馒头在冷冻储存过程中的弹性。
3.7. Crystal structure 3.7. 晶体结构
XRD can be utilized to determine the structure of crystalline substances at the atomic level. The crystal structure of starch granules, formed due to the aggregation of amylose and branched starch, disappears upon gelatinization but can spontaneously rearrange and recrystallize during the cooling process. The relative crystallinity, as determined by XRD, provides insights into the degree of starch retrogradation. The XRD patterns and relative crystallinity of steamed bread with KGM at 0 and 3 weeks are displayed in Fig. 5a and b. A distinct diffraction peak was identified at (20) in steamed bread (Fig. 5a), which corresponds to V-type crystals and reflects the amylose-lipid complexes (Lalush et al., 2005). After 3 weeks of frozen storage, a crystalline B-type emerged with a prominent diffraction peak near (20) (Fig. 5b). In one study, both the steamed bread with and without inulin exhibited two prominent peaks at and after being stored for 5 days (Kou et al., 2019). This is attributed to the gelatinization of starch by the formation of double helices with amylose and amylopectin chains, resulting in more structural crystallization (Kang, Reddy, Park, Choi, & Lim, 2018), further leading to an increase in hardness. The difference in XRD patterns between 0 and 3 weeks is partly due to the difference in the water content (Hong et al., 2023). Additionally, the relative crystallinity of all treatments increased during frozen storage, indicating that starch retrogradation occurred. XRD 可用于在原子水平上确定晶体物质的结构。淀粉颗粒的晶体结构是由于直链淀粉和支链淀粉的聚集而形成的,在糊化后消失,但在冷却过程中可以自发重排和重结晶。通过 XRD 测定的相对结晶度有助于了解淀粉的回生程度。图 5a 和 b 显示了 0 周和 3 周时 KGM 蒸面包的 XRD 图谱和相对结晶度。在蒸面包的 (20) 处 发现了一个明显的衍射峰(图 5a),它对应于 V 型晶体并反映了直链淀粉-脂质复合物(Lalush et al., 2005)。冷冻储存 3 周后,出现结晶 B 型,在 (20) 附近 有一个突出的衍射峰(图 5b)。在一项研究中,含菊粉和不含菊粉的馒头在储存 5 天时 和 储存 5 天后都表现出两个突出的峰值(Kou et al., 2019)。这归因于淀粉的糊化,通过形成双螺旋与直链淀粉和支链淀粉,导致更多的结构结晶(Kang, Reddy, Park, Choi, & Lim, 2018),进一步导致硬度增加。XRD 图谱在 0 周和 3 周之间的差异部分是由于含水量的差异(Hong et al., 2023)。此外,在冷冻储存过程中,所有处理的相对结晶度均增加,表明发生了淀粉回生。
Here, at 0 or 3 weeks, the diffraction peaks in the KGM groups exhibited a similar trend to the control group (Fig. 5a and b), suggesting that adding KGM did not alter the crystal structure of starch. However, obvious differences were observed in the relative crystallinity of samples with varying levels of KGM addition. As the KGM increased, the relative crystallinity initially decreased and slightly increased at the same frozen storage time. Particularly at week 3, the relative crystallinity in KGM groups ( ) was lower than that in the control group ( ). Meanwhile, the increment of relative crystallinity in the control group was , while in the KGM groups, it ranged from to from 0 to 3 weeks. This indicates that the addition of KGM inhibits the recrystallization of amylopectin, thereby delaying the staling of samples. The reason was that KGM prevents the aggregation of amylose molecules by forming hydrogen bonds with the hydroxyl groups on the starch chain, reducing the ability of amylose to form a V-type complex with lipids (Gómez et al., 2011). In one study, the presence of mulberry leaf polysaccharides could retard the recrystallization of starch after storage (Xie et al., 2022). 在这里,在 0 或 3 周时,KGM 组的衍射峰表现出与对照组相似的趋势(图 5a 和 b),表明添加 KGM 不会改变淀粉的晶体结构。然而,在不同 KGM 添加量水平下,观察到样品的相对结晶度存在明显差异。随着 KGM 的增加,相对结晶度在相同的冷冻贮藏时间内最初下降并略有增加。特别是在第 3 周,KGM 组 ( ) 的相对结晶度低于对照组 ( )。同时,对照组相对结晶度的增加为 ,而 KGM 组的相对结晶度增量为 0 至 3 周。这表明KGM的添加抑制了支链淀粉的重结晶,从而延缓了样品的老化。原因是 KGM 通过与淀粉链上的羟基形成氢键来阻止直链淀粉分子的聚集,从而降低了直链淀粉与脂质形成 V 型复合物的能力(Gómez et al., 2011)。在一项研究中,桑叶多糖的存在可以延缓淀粉储存后的再结晶(Xie等人,2022 年)。
3.8. Short-range ordered structure 3.8. 短程有序结构
FT-IR is a useful tool used to study the vibrational frequencies and energy levels of chemical bonds within a sample. In the scanning range of , the changes in the peak width of the wave number can indicate the staling degree of the samples. The FT-IR spectrum of samples with various KGM added at 0 and 3 weeks is presented in FT-IR 是一种有用的工具,用于研究样品中化学键的振动频率和能级。在 的扫描范围内,波数峰宽的变化可以指示样品的老化程度。在 0 周和 3 周添加各种 KGM 的样品的 FT-IR 光谱如
Fig. 4. Water content (a), T2 distributions of relaxation times (b), peak area proportion of proton populations (c), and free sulfhydryl (SH) content (d) of steamed bread added with different amounts of konjac glucomannan (KGM) and frozen at different time. Different lower case letters represent significant ( ) differences among steamed bread with different KGM additions at the same frozen time. Different upper case letters indicate significant ( ) differences among steamed bread with the same KGM addition at different frozen time. 图 4.添加不同量魔芋葡甘聚糖 (KGM) 并在不同时间冷冻的馒头的含水量 (a)、 弛豫时间的 T2 分布 (b)、质子群的峰面积比例 (c) 和游离巯基 (SH) 含量 (d)。不同的小写字母表示在同一冷冻时间内添加不同 KGM 的馒头之间的显著 ( ) 差异。不同的大写字母表示相同 KGM 添加量的馒头在不同冷冻时间存在显著 ( ) 差异。
Fig. 5c. The band at represents the complex stretching vibration of starch and characterizes the strength of hydrogen bonds (Pan et al., 2019; Zheng et al., 2021). The band at is associated with water absorbed in the amorphous regions, while the spectral peak near corresponds to the backbone mode vibrations of starch and glycosidic bonds (Pan et al., 2019; Zheng et al., 2021). No new peaks were observed in any of the samples (Fig. 5c), indicating that neither the addition of KGM nor the frozen treatment alters the chemical composition of the steamed bread molecules or introduces new chemical groups. After being frozen for 3 weeks, the bandwidth of the steamed bread at became narrower, and the stretching vibration peak position shifted to a higher wavenumber compared with the 0 -week samples, indicating that the frozen storage destroys part of the hydrogen bonds. Conversely, at 0 or 3 weeks, the KGM-added samples had wider bands at compared with the control sample. This suggests that starch added with KGM could form hydrogen bonds much more easily than wheat starch, disrupting the interaction between starch chains and thus causing changes in starch structure (Pan et al., 2019; Zheng et al., 2021). 图 5c.带 at 代表淀粉的复杂 拉伸振动并表征氢键的强度(Pan等人,2019 年;Zheng et al., 2021)。带 at 与非晶区吸收的水有关,而附近的 光谱峰对应于淀粉和糖苷键的骨架模式振动(Pan等人,2019 年;Zheng et al., 2021)。在任何样品中均未观察到新的峰(图 5c),表明添加 KGM 和冷冻处理都不会改变蒸面包分子的化学成分或引入新的化学基团。冷冻 3 周后,馒头的 带宽变窄, 拉伸振动峰值位置与 0 周样品相比向更高的波数移动,表明冷冻储存破坏了部分氢键。相反,在 0 或 3 周时,与对照样本 相比,KGM 添加的样本具有更宽的条带。这表明添加 KGM 的淀粉比小麦淀粉更容易形成氢键,破坏淀粉链之间的相互作用,从而导致淀粉结构发生变化(Pan等人,2019 年;Zheng et al., 2021)。
The deconvoluted FT-IR spectrum in the wavelength range of is presented in Fig. 5d. The absorbance ratio of R1047/ 1022 reflects the starch crystalline fraction over a short range, and R995/1022 represents the degree of short-range order of molecules (Xie et al., 2022). The R1047/1022 value decreased and then increased as KGM increased at the same frozen storage time, with showing the lowest value (Fig. 5d and Table 1). During frozen storage, the R1047/1022 significantly ( ) increased in all KGM groups. From 0 to 3 weeks, R1047/1022 of samples with different KGM additions ( , and ) increased from , and 0.804 to , and 0.814 , respectively. The increment of R1047/1022 in samples with KGM was less than in the control sample during frozen storage, suggesting that adding KGM reduces the crystallinity of starch. This is because KGM with strong hydrophilicity reduces water loss, thus retarding cross-linking between gluten and starch (Xie et al., 2022). Conversely, in one study, adding KGM to rice starch increased R1047/1022. This is attributed to the structural differences among different starches (He, Zhang, Liao, & Shen, 2023). At 0 or 3 weeks, the steamed bread added with KGM showed a decrease in the R995/1022 values compared with the control, indicating a reduction in the double helical molecular order of starch. Moreover, as the frozen storage time extended, the R995/1022 values displayed a similar trend to R1047/1022. The results from XRD and FT-IR confirmed these findings. Thus, it can be inferred that KGM is an effective additive in frozen steamed bread, maintaining its quality. 波长范围内的去卷积 FT-IR 光谱 如图 5d 所示。R1047/1022 的吸光度比反映了短范围内的淀粉结晶分数,R995/1022 代表分子短程有序的程度(Xie等人,2022 年)。在相同的冷冻贮藏时间内,R1047/1022 值随着 KGM 的增加而降低后增加,显示出 最低的值(图 5d 和表 1)。在冷冻贮藏期间,R1047/1022 在所有 KGM 组中均显著 ( ) 增加。从 0 到 3 周,添加不同 KGM ( 、 和 ) 的样品的 R1047/1022 分别从 、 和 0.804 增加到 和 0.814。在冷冻储存过程中,含有 KGM 的样品中 R1047/1022 的增加小于对照样品,表明添加 KGM 会降低淀粉的结晶度。这是因为具有强亲水性的 KGM 减少了水分流失,从而延缓了面筋和淀粉之间的交联(Xie et al., 2022)。相反,在一项研究中,在大米淀粉中添加 KGM 会增加 R1047/1022。这归因于不同淀粉之间的结构差异(He, Zhang, Liao, & Shen, 2023)。在 0 或 3 周时,添加 KGM 的馒头与对照相比,R995/1022 值降低,表明淀粉的双螺旋分子顺序降低。此外,随着冷冻储存时间的延长,R995/1022 值显示出与 R1047/1022 相似的趋势。XRD 和 FT-IR 的结果证实了这些发现。因此,可以推断 KGM 是冷冻馒头中的有效添加剂,保持其质量。
3.9. Free sulfhydryl content 3.9. 游离巯基含量
Characterizing free sulfhydryl groups is crucial for understanding the structural integrity and stability of proteins, especially in the context of protein aggregation and matrix stability. The changes in free sulfhydryl 表征游离巯基对于了解蛋白质的结构完整性和稳定性至关重要,尤其是在蛋白质聚集和基质稳定性的背景下。游离巯基的变化
Fig. 5. XRD pattern of steamed bread frozen for 0 weeks (a) and 3 weeks (b), original FT-IR spectra (c), and deconvoluted spectra (d) of steamed bread. The lines a, b, c, d, e, f, g, and h represent , and , respectively. 图 5.冷冻 0 周 (a) 和 3 周 (b) 的馒头 XRD 图谱、馒头的原始 FT-IR 光谱 (c) 和去卷积光谱 (d)。行 a、b、c、d、e、f、g 和 h 分别表示 和 。
content in steamed bread are displayed in Fig. 4d. Frozen samples presented an obvious ( ) increase in free sulfhydryl content compared with the 0 -week samples, with a tendency to increase further over frozen storage time. This is attributed to the disruption of crosslinking disulfide bonds between gliadin and glutenin during frozen storage. Additionally, physical damage to proteins in frozen products due to ice recrystallization could contribute to these changes. 馒头中的含量如图 4d 所示。与 0 周样品相比,冷冻样品的游离巯基含量明显增加 ( ),并且随着冷冻储存时间的增加,有进一步增加的趋势。这归因于冷冻储存过程中麦醇溶蛋白和谷蛋白之间的交联二硫键的破坏。此外,冰重结晶对冷冻产品中蛋白质的物理损伤也可能导致这些变化。
At the same frozen storage time, the free sulfhydryl content of KGM groups was significantly lower than that of the control group, indicating that KGM efficiently facilitates the strengthening of the gluten network. While KGM itself lacked the chemical structure required for disulfide bond formation, it likely interacted with gluten proteins through non-covalent interactions, altering the protein's spatial conformation. This proximity of sulfhydryl groups made them more susceptible to oxidation into disulfide bonds (Guan et al., 2023). Interestingly, compared with the 1.0 % KGM group, increasing the KGM concentration to led to a slight increase in free sulfhydryl content, suggesting that excess KGM acts as a barrier to protein aggregation via disulfide crosslinking. These findings imply that optimal KGM addition facilitates protein polymerization. Moreover, the increment of free sulfhydryl content was lower in the KGM groups than in the control, suggesting that KGM reduces disulfide bond cleavage. KGM may interact with gluten proteins to inhibit water redistribution and ice recrystallization, thus delaying disulfide bond breakage and increasing free sulfhydryl content (Guo et al., 2021a). 在相同冷冻贮藏时间内,KGM 组的游离巯基含量显著 低于对照组,表明 KGM 有效地促进了面筋网络的强化。虽然 KGM 本身缺乏形成二硫键所需的化学结构,但它可能通过非共价相互作用与面筋蛋白相互作用,从而改变蛋白质的空间构象。巯基的这种接近性使它们更容易氧化成二硫键(Guan等人,2023 年)。有趣的是,与 1.0% KGM 组相比,增加 KGM 浓度 导致游离巯基含量略有增加,这表明过量的 KGM 通过二硫键交联成为蛋白质聚集的障碍。这些发现表明最佳 KGM 添加有助于蛋白质聚合。此外,KGM 组游离巯基含量的增加低于对照,表明 KGM 减少了二硫键裂解。KGM 可能与面筋蛋白相互作用以抑制水的再分布和冰的重结晶,从而延缓二硫键断裂并增加游离巯基含量(Guo等人,2021a)。
3.10. Microstructure 3.10. 微观结构
Steamed bread is characterized by a soft and porous solid microstructure, reminiscent of a sponge-like texture (Jekle & Becker, 2012). The microstructure of KGM-added samples frozen for 0 or 3 weeks was examined using SEM, as depicted in Fig. 6. At 0 weeks, all samples displayed intact, continuous pores and smooth gluten network structures. Furthermore, the gluten network structure in samples with KGM addition appeared more uniform and smooth compared with the control, particularly at KGM addition. However, after 3 weeks of frozen storage, numerous fractures were observed in the pore structure, and the gluten structure became less cohesive for all samples. This is attributed to water migration and ice recrystallization, which cause significant mechanical damage to the continuous gluten structure of samples, consequently increasing the free sulfhydryl content. Similar disruptions in microstructure were reported in frozen steamed bread (Qian et al., 2021).In another study, starch particles were observed to separate from the gluten network following frozen storage (Gao et al., 2023). The weakened gluten network in frozen samples resulted in reduced fermentation gas, further leading to a decrease in specific volume and an increase in hardness. 蒸面包的特点是柔软且多孔的固体微观结构,让人联想到海绵状的质地(Jekle & Becker,2012)。使用 SEM 检查冷冻 0 或 3 周的 KGM 添加样品的微观结构,如图 6 所示。在 0 周时,所有样品均显示完整、连续的孔和光滑的面筋网络结构。此外,与对照相比,添加 KGM 的样品中的面筋网络结构显得更加均匀和光滑,尤其是在添加 KGM 时 。然而,冷冻储存 3 周后,孔隙结构中观察到大量裂缝,并且所有样品的面筋结构都变得不那么粘结。这归因于水迁移和冰再结晶,它们对样品的连续面筋结构造成了严重的机械损伤,从而增加了游离巯基的含量。在冷冻馒头中也报道了类似的微观结构破坏(Qian et al., 2021)。在另一项研究中,观察到淀粉颗粒在冷冻储存后与面筋网络分离(Gao et al., 2023)。冷冻样品中面筋网络减弱导致发酵气体减少,进一步导致比容降低和硬度增加。
However, after 3 weeks of freezing, less breakage was observed in KGM-added samples compared with the control. Interestingly, the least breakage to the gluten network structure occurred when the KGM addition was . Concurrently, KGM-added samples displayed the maximum specific volume (Table 1) and the minimum hardness 然而,冷冻 3 周后,与对照相比,在 KGM 添加的样品中观察到的破损更少。有趣的是,当 KGM 添加 时,面筋网络结构的破坏最小。同时, 添加 KGM 的样品显示最大比体积(表 1)和最小硬度 KGM
Fig. 6. Microstructure of KGM-added steamed bread frozen for 0 or 3 weeks. First and third rows: magnification of ; second and fourth rows: magnification of . KGM, konjac glucomannan. 图 6.添加 KGM 的馒头冷冻 0 或 3 周的微观结构。第一行和第三行:放大倍数 ;第二行和第四行:放大 。KGM,魔芋葡甘露聚糖。
(Table 2). This indicates that adding KGM provides sufficient gluten structural stability and hydrophilicity, thereby mitigating the adverse effects of freezing on steamed bread quality. This is because KGM prevents water migration and ice recrystallization, thus effectively enhancing frozen steamed bread quality. These results were in line with changes in water status (Fig. 4a, b, and c) and free sulfhydryl content (Fig. 4d). (表 2)。这表明添加 KGM 提供了足够的面筋结构稳定性和亲水性,从而减轻了冷冻对馒头品质的不利影响。这是因为 KGM 防止了水迁移和冰再结晶,从而有效地提高了冷冻馒头的品质。这些结果与水状态(图 4a、b 和 c)和游离巯基含量(图 4d)的变化一致。
3.11. Volatile flavor compounds 3.11. 挥发性风味化合物
The aroma profile is widely recognized as a crucial aspect of steamed bread quality. Volatile compounds in steamed bread are intricate, requiring sophisticated techniques such as gas chromatography and mass spectrometry for analysis and identification of specific changes. As illustrated in Fig. 7c, to depict the changes in volatile compounds across all steamed bread groups during frozen storage, a heat map analysis was conducted. In this analysis, a blue box indicates a high level of a volatile compound, while a red box suggests a low level. The volatile compounds within steamed bread comprise 6 alcohols (ethanol, phenylethyl alcohol, isoamyl alcohol, 2,3-dimethyl-2-pentanol, (Z)-2-methyl-3octen-2-ol, and cis-2-methyl-cyclopentanol), 4 aldehydes (hexanal, nonanal, benzaldehyde, and decanal), 4 ketones (4-octanone, 2-methyl3-octanone, 1-phenyl-2-butanone, and 7-tridecanone), and 3 esters (butyl isobutyrate, methyl benzoate, and octyl-oxirane) (Fig. 7c). 香气特征被广泛认为是馒头质量的一个重要方面。馒头中的挥发性化合物很复杂,需要复杂的技术,如气相色谱法和质谱法来分析和鉴定特定变化。如图 7c 所示,为了描述冷冻储存过程中所有馒头组中挥发性化合物的变化,进行了热图分析。在此分析中,蓝色框表示挥发性化合物含量高,而红色框表示低含量。馒头中的挥发性化合物包括 6 种醇类(乙醇、苯乙醇、异戊醇、2,3-二甲基-2-戊醇、(Z)-2-甲基-3辛烯-2-醇和顺-2-甲基-环戊醇)、4 种醛类(己醛、壬醛、苯甲醛和癸醛)、4 种酮类(4-辛酮、2-甲基 3-辛酮、1-苯基-2-丁酮和 7-十三烷酮)和 3 种酯类(异丁酸丁酯、苯甲酸甲酯和辛基环氧乙烷)(图 7c)。
Here, ethanol was a major volatile compound within all steamed bread frozen for 0 or 3 weeks. At 0 or 3 weeks, the ethanol content within KGM-added samples was lower than that within the control group. This is because of a lower starch composition within KGM-added steamed bread. Furthermore, compared with the control, the levels of all alcohols, 4-octanone, 2-methyl-3-octanone, methyl benzoate, and octyloxirane within KGM-added samples were lower at week 0 . Meanwhile, the levels of hexanal, nonanal, and benzaldehyde were higher in the steamed bread with KGM compared with the control. The levels of volatile flavor compounds within all samples decreased from 0 to 3 weeks. This is because the disruption of gluten structure is not conducive to the retention of volatile substances. 在这里,乙醇是所有冷冻 0 或 3 周的馒头中的主要挥发性化合物。在 0 或 3 周时,KGM 添加样品中的乙醇含量低于对照组内。这是因为 KGM 添加的馒头中的淀粉成分较低。此外,与对照相比,KGM 添加样品中所有醇、4-辛酮、2-甲基-3-辛酮、苯甲酸甲酯和辛氧醚的水平在第 0 周时较低。同时,与对照相比,KGM 蒸面包中己醛、壬醛和苯甲醛水平较高。所有样品中挥发性风味物质的水平从 0 到 3 周下降。这是因为面筋结构的破坏不利于挥发性物质的保留。
At week 3, the levels of all esters, 4-octanone, 1-phenyl-2-butanone, 7-tridecanone, 2-pentanol, 2,3-dimethyl-2-pentanol, and cis-2-methylcyclopentanol within KGM-added samples were higher compared with the control. This finding indicates that adding KGM effectively protects the gluten structure of frozen samples, thereby slowing down the decrease in volatile compounds. In one study, the addition of KGM resulted in the formation of smaller ice crystals and reduced gluten damage during freezing (Cui, Liu, Wu, Sui, & Zhang, 2019). This effect is attributed to KGM's strong water adsorption ability, which limits the conversion of freezable water into ice crystals. As a result, KGM helps protect the gluten structure from ice crystal damage (Xuan et al., 2017). Indeed, in sensory evaluations of wheat bread, elevated levels of alcohols, ketones, and esters are commonly associated with positive attributes (Pico, Bernal, & Gomez, 2015). The findings indicate a favorable impact of KGM on the flavor profile of frozen samples. 第 3 周时,KGM 添加样品中所有酯、4-辛酮、1-苯基-2-丁酮、7-十三烷酮、2-戊醇、2,3-二甲基-2-戊醇和顺-2-甲基环戊醇的水平高于对照。这一发现表明,添加 KGM 可有效保护冷冻样品的面筋结构,从而减缓挥发性化合物的减少。在一项研究中,添加KGM导致在冷冻过程中形成更小的冰晶并减少面筋损伤(Cui, Liu, Wu, Sui, & Zhang, 2019)。这种效果归因于 KGM 强大的吸水能力,它限制了可冷冻水转化为冰晶。因此,KGM 有助于保护面筋结构免受冰晶损伤(Xuan et al., 2017)。事实上,在小麦面包的感官评估中,醇、酮和酯水平的升高通常与积极的属性有关(Pico, Bernal, & Gomez, 2015)。研究结果表明,KGM 对冷冻样品的风味特征有有利影响。
Fig. 7. Score plot (a), loading plot (b) of principal component analysis, and heat map analysis (c) of volatile compounds in steamed bread. The score plot shows the steamed bread frozen for 0 and 3 weeks, respectively. Multiple variables of hardness, , relative crystallinity, water content, specific volume, free sulfhydryl content, A21, A22, and A23 are analyzed in the loading plot. 图 7.馒头中挥发性化合物的评分图 (a)、主成分分析的载荷图 (b) 和热图 (c)。分数图分别显示冷冻 0 周和 3 周的馒头。在加载图中分析了硬度、 相对结晶度、含水量、比容、游离巯基含量、A21、A22 和 A23 的多个变量。
3.12. PCA analysis 3.12. PCA 分析
Factor analysis was conducted on typical parameters to fully understand the impact of KGM on steamed bread quality during frozen storage (Fig. 7a and b). In the PCA plots, the first two main components accounted for of the variance, with PC1 and PC2 explaining 72.5 and of the variance, respectively. In the score plot, closer distances indicate higher similarity between sample points, while in the loadings plot, closer variables indicate stronger positive correlation coefficients. Steamed bread samples were clustered into four groups. The distribution of samples was close to , while 1.0 samples were close to , suggesting similarities between control samples and those with KGM, but noticeable differences from samples with or KGM (Fig. 7a). On the negative side of PC1 and PC2, steamed bread was characterized by higher specific volume and water content, indicating that or KGM significantly improved steamed bread quality. Water content had a positive correlation with specific volume and a negative correlation with hardness, 对典型参数进行因子分析,以充分了解 KGM 对冷冻贮藏期间馒头品质的影响(图 7a 和 b)。在 PCA 图中,前两个主要分量解释了 方差,PC1 和 PC2 分别解释了 72.5 和 方差。在分值图中,距离越近表示样本点之间的相似性越高,而在载荷图中,变量越近表示正相关系数越强。将馒头样品分为四组。样本分布 接近 ,而 1.0 样本接近 ,表明对照样本与具有 KGM 的样本之间存在相似性,但与具有 或 KGM 的样本之间存在显着差异(图 7a)。在 PC1 和 PC2 的负端,馒头具有较高的比容和含水量,表明 KGM 显著提高了馒头品质。含水量与比容呈正相关,与硬度呈负相关,
R1047/1022, relative crystallinity, and free sulfhydryl content (Fig. 7b), suggesting that these parameters could be used as indicators to evaluate steamed bread deterioration during frozen storage. On the positive side of PC1, was separated from , and by higher hardness, free sulfhydryl content, R1047/1022, relative crystallinity, and A23, indicating that KGM had a favorable effect on steamed bread, delaying the negative effects of frozen storage. R1047/1022、相对结晶度和游离巯基含量(图 7b),表明这些参数可以用作评估馒头在冷冻储存过程中变质的指标。在 PC1 的积极面, 与较高的硬度、游离巯基含量、R1047/1022、相对结晶度和 A23 分离 ,表明 KGM 对馒头有有利影响,延缓了冷冻储存的负面影响。
3.13. Mechanism 3.13. 机制
Based on the results, we propose the mechanism by which KGM mitigates the quality deterioration of steamed bread during frozen storage. (i) KGM's structure enhances water interaction through hydrogen bonding (as indicated by increased water absorption). The hydroxyl groups in KGM's molecular chain reduce the mobility of adsorbed water through proton exchange (as evidenced by LF-NMR). This reduces the number and volume of ice crystals by limiting water migration during frozen storage. Consequently, the physical damage of 基于研究结果,我们提出了 KGM 减轻馒头在冷冻贮藏过程中质量劣化的机制。(i) KGM 的结构通过氢键增强水相互作用(如吸水率增加所示)。KGM 分子链中的羟基通过质子交换降低吸附水的迁移率(如 LF-NMR 所证明的那样)。这通过限制冷冻储存期间的水迁移来减少冰晶的数量和体积。因此,物理损伤
ice crystals to the gluten network is minimized, thereby protecting the gluten network during frozen storage (as shown by SEM). (ii) The hydroxyl groups on the KGM polysaccharide chain form hydrogen bonds and hydrophobic interactions with amino acids on the gluten protein side chains. This interaction strengthens non-covalent bonds between gluten protein chains and increases cross-linking among gluten proteins (as indicated by free sulfhydryl content), ultimately providing additional support to the gluten network. (iii) KGM competes with starch for water, preventing water molecules from penetrating the non-crystalline regions of starch granules. This helps protect the hydrogen bonds in the crystalline regions and reduces the leaching of amylose molecules. Additionally, KGM molecules form hydrogen bonds with hydroxyl groups on the starch chains, entangling with leached amylose and competing with amylose-amylose interactions. This prevents amylose aggregation and inhibits starch retrogradation (as demonstrated by XRD and FT-IR). In summary, the addition of KGM protects the gluten network and inhibits starch retrogradation through these complex interactions in the steamed bread. This results in significant improvements in the quality of steamed bread, including specific volume (Table 1), texture (Table 2), sensory properties (Fig. 3), and flavor compounds (Fig. 7 c ). 面筋网络的冰晶被最小化,从而在冷冻储存过程中保护面筋网络(如 SEM 所示)。(ii) KGM 多糖链上的羟基与面筋蛋白侧链上的氨基酸形成氢键和疏水相互作用。这种相互作用加强了面筋蛋白链之间的非共价键,并增加了面筋蛋白之间的交联(如游离巯基含量所示),最终为面筋网络提供额外的支持。(iii) KGM 与淀粉竞争水,阻止水分子渗透淀粉颗粒的非结晶区域。这有助于保护结晶区域中的氢键并减少直链淀粉分子的浸出。此外,KGM 分子与淀粉链上的羟基形成氢键,与浸出的直链淀粉缠结并与直链淀粉-直链淀粉相互作用竞争。这可以防止直链淀粉聚集并抑制淀粉回生(如 XRD 和 FT-IR 所示)。总之,添加 KGM 可以保护面筋网络,并通过馒头中的这些复杂相互作用抑制淀粉回生。这导致馒头的质量得到显著改善,包括比容(表 1)、质地(表 2)、感官特性(图 3)和风味化合物(图 7 c)。
4. Conclusions 4. 结论
This study delved into the impact of KGM on the storage properties of steamed bread during frozen storage, along with a detailed analysis of potential mechanisms. The results underscored that the incorporation of KGM notably improved the specific volume, texture, and sensory attributes of steamed bread throughout frozen storage, with the most optimal effect observed at 1.0 % KGM addition. Moreover, KGM was found to mitigate the increase in water migration and decrease in water content in steamed bread frozen from 0 to 3 weeks. Crystal structure analysis unveiled that KGM could engage in hydrogen bonding with hydroxyl groups on the starch chain, influencing starch crystallization during frozen storage. This observation was corroborated by FT-IR analysis, which indicated retardation in crystallinity increase and reduction in short-range ordered structure with KGM addition. SEM analysis further revealed a reduction in gluten network damage and cracking with KGM addition, while the lowest free sulfhydryl content was observed in steamed bread containing KGM after 3 weeks of frozen storage. Additionally, KGM contributed to flavor retention in steamed bread during frozen storage. In summary, KGM emerges as a promising improver for maintaining the overall quality of steamed bread during frozen storage, influencing quality characteristics, water status, multi-scale structure, and flavor compounds. Future research should explore the effect of KGM on the nutritional properties of frozen steamed bread. 本研究深入探讨了 KGM 对馒头在冷冻储存过程中储存特性的影响,并详细分析了潜在机制。结果强调,KGM 的加入显着改善了整个冷冻储存过程中馒头的比体积、质地和感官属性,添加 1.0% KGM 时观察到最佳效果。此外,发现 KGM 可以减轻冷冻馒头中 0 至 3 周水分迁移的增加和水分含量的降低。晶体结构分析揭示了 KGM 可以与淀粉链上的羟基进行氢键,从而影响冷冻储存过程中的淀粉结晶。FT-IR 分析证实了这一观察结果,该分析表明添加 KGM 后结晶度增加延迟,短程有序结构减少。SEM 分析进一步显示,添加 KGM 后面筋网络损伤和开裂减少,而在含有 KGM 的馒头中观察到游离巯基含量最低,冷冻储存 3 周后。此外,KGM 有助于在冷冻储存过程中保持馒头的风味。综上所述,KGM 是一种很有前途的改进剂,可以维持馒头在冷冻贮藏过程中的整体品质,影响品质特性、水分状态、多尺度结构和风味化合物。未来的研究应探讨 KGM 对冷冻馒头营养特性的影响。
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. 作者声明,他们没有已知的竞争性经济利益或个人关系,这些利益或个人关系似乎可能会影响本文报告的工作。
Data availability 数据可用性
Acknowledgments 确认
This work was financially supported by the Higher Foreign Expert Introduction Program of Henan Province (HNGD2022054) and the National Study Abroad Fund of China (202208410256). 这项工作得到了河南省高等外国专家引进计划 (HNGD2022054) 和中国国家留学基金 (202208410256) 的财政支持。
Appendix A. Supplementary material 附录 A. 补充材料
Supplementary data to this article can be found online at https://doi. org/10.1016/j.foodres.2024.114962. 本文的补充数据可在 https://doi 在线找到。org/10.1016/j.foodres.2024.114962.
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