In vitro digestion of soymilk using a human gastric simulator: Impact of structural changes on kinetics of release of proteins and lipids
Graphical abstract
Introduction
Soybeans have been part of the East Asian diet for centuries. Soybean seeds contain approximately 20% fat and 40% protein on a dry weight basis, providing the most inexpensive source of edible oils and high quality proteins (Kwok & Niranjan, 1995). Soymilk is a traditional beverage that is produced by grinding soybeans soaked in water, followed by cooking and removal of the insoluble material by filtration or centrifugation (Chen, 1988). Soymilk contains a variety of nutrients, such as high quality proteins and isoflavones; the consumption of soya-based foods has been shown to have beneficial effects in lowering blood pressure and improving lipid profile (Bricarello et al., 2004; Fukui et al., 2002; Rivas et al., 2002).
During the manufacture of soymilk, heating is an essential processing step to denature anti-nutritional compounds, such as trypsin inhibitors and lipoxygenase, and to modify the structure of the soymilk proteins to improve their colloidal stability (Nik, Tosh, Woodrow, Poysa, & Corredig, 2009). Soy proteins are known to denature, dissociate and subsequently aggregate to form particles during heating (Guo, Ono, & Mikami, 1997; Ono, Rak Choi, Ikida, & Odaoiri, 1991), which can influence the stability and the digestibility of soymilk products (Guo et al., 2006; Wallace, Bannatyne, & Khaleque, 1971). Soy lipids are contained in oil bodies, which have an average size of 300–400 nm (Ono, 2008). The surface of soybean oil bodies is made up of a monolayer of phospholipids, embedded with intrinsic proteins, called oleosins (Zhao, Chen, Cao, Kong, & Hua, 2013). Grinding of soybeans in the presence of water leads to the dispersion of oil bodies and extrinsic proteins, but some of these extrinsic proteins remain associated with the oil bodies. After heating (70–100 °C), some (but not all) of the extrinsic proteins dissociated from the oil bodies (Guo et al., 1997; Yan, Zhao, Kong, Hua, & Chen, 2016) and a few large oil droplets or oil body-protein aggregates (˃1 μm) were also observed (Chen, Zhao, Kong, Zhang, & Hua, 2014; Cruz et al., 2007). The surfaces of oil bodies in heated soymilk appeared to be coated by oleosins, phospholipids and some extrinsic proteins (Yan et al., 2016). Ding et al. (2020) reported that the oil bodies isolated from heated soymilk retained the oleosins at their surface.
Because of its proposed nutritional and health benefits, the digestibility of soymilk has been evaluated both in vivo and in vitro (Baglieri et al., 1994; Liu et al., 2019; Rui et al., 2016, 2019). Most current studies on the digestion of soymilk are focused on the bioaccessibility of bioactive compounds (Rodríguez-Roque, Rojas-Graü, Elez-Martínez, & Martín-Belloso, 2013; Wu et al., 2012). There is a lack of knowledge on the changes in colloidal behaviour and macronutrient delivery during gastrointestinal digestion. Moreover, no detailed studies on the digestion profile of soymilk in a dynamic digestive environment have been reported; for example, there have been no studies on the spatial distribution of macronutrients because of physical instability (e.g. coalescence or phase separation) in the stomach, or on structural changes of the proteins when experiencing pH changes. Using a dynamic human gastric simulator (HGS), our recent work on the behaviour of milk has revealed new insights into its coagulation (Ye, Cui, Dalgleish, & Singh, 2016b) and the consequences of this coagulation on the kinetics of the delivery of proteins and lipids to the small intestine (Ye, Cui, Dalgleish, & Singh, 2016a). Thus, the purpose of this study was to evaluate the structural changes during the gastric digestion of soymilk using a dynamic gastric digestion model.
Section snippets
Materials
Commercial dried soybeans were purchased from local grocery stores in Palmerston North, New Zealand. The chemical composition (per 100 g) as determined by standard methods was: protein (AOAC 991.20.II), 32.7 g; fat (AACC 30–10), 14.7 g; moisture (AOAC International, 2000), 10 g; ash (AOAC International, 2002), 4.4 g and total carbohydrate (by difference to achieve 100% of total content), 38.2 g. Pepsin from porcine gastric mucosa (EC 3.4.23.1; 695 units/mg solid) and pepstatin A (77170,
Changes in intragastric pH during dynamic digestion
The undigested soymilk contained 3.0% protein, 1.8% fat and 6.93% dry matter. The gastric digestion was performed using a dynamic in vitro digestion model, by the gradual addition of acid and enzyme to approximate the secretion process occurring in the stomach in vivo. The changes in the pH profile of soymilk during gastric digestion are shown in Fig. 2. The pH of the soymilk was significantly influenced by the digestion time (P < 0.001). It was approximately 6.65 initially, reduced to pH 4.5
Discussion
Soymilk is a turbid colloidal dispersion with majority of the particles between 50 and 1500 nm. (Fig. 4B and C, 0 min), representing protein particles and oil bodies. It is known that protein particles of greater than 40 nm in diameter constitute about 50% (including 14% of more than 100 nm) of the total protein in soymilk (Ono et al., 1991). Soybean oil bodies have an average particle size of 300–400 nm (Iwanaga et al., 2007; Ono, 2008).
Conclusions
The coagulation of soy proteins induced by both the action of pepsin and the acidic gastric pH was observed during dynamic in vitro gastric digestion. This resulted in a layer of coagulated protein particles at the bottom of the HGS. Because of relatively small size of these coagulated particles, a rapid emptying of proteins from the HGS to the next digestion step was seen in the early stages of gastric digestion. This observation is in stark contrast to the behaviour of cow's milk where
Funding
This research was supported by Tertiary Education Commission‒Centre of Research Excellence (CoRE) funding, New Zealand.
CRediT authorship contribution statement
Xin Wang: Methodology, Validation, Investigation, Formal analysis, Data curation, Writing - original draft, Visualization. Aiqian Ye: Conceptualization, Methodology, Writing - review & editing, Supervision. Anant Dave: Writing - review & editing, Supervision. Harjinder Singh: Conceptualization, Methodology, Resources, Writing - review & editing, Project administration, Supervision, Funding acquisition.
Declaration of competing interest
There are no conflicts of interest to declare.
Acknowledgements
We thank Dr Matthew Savoian (Manawatu Microscopy and Imaging Centre, Massey University, Palmerston North, New Zealand) for his technical assistance and useful advice on confocal imaging and Dr Wei Zhang (AgResearch Limited, Palmerston North, New Zealand) for her technical assistance with the statistical analysis.
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