Review on the extraction, characterization and application of soybean polysaccharide

Abstract

Soybean polysaccharide (SPS) is a class of soluble polysaccharide derived from soybean cotyledon, soybean meal or okara, and has broadly been used in the food industry. In recent decades, due to its attractive physicochemical properties, SPS has been developed into various emulsifiers or stabilizers for beverages. Additionally, studies have emerged to reveal its potential in biomaterial and biological applications. In this review, we critically appraise the latest literature on the extraction and the structural features of SPS, and provide a perspective on the biological applications of SPS. We focus on the current strategies for the extraction of this unique polysaccharide, specific structural features, and functional utilization of SPS. Notably, SPS-based food additives have been demonstrated to add value in biological applications due to their anticancer and immunoregulatory effects, encouraging us to use SPS directly in the area of biomedicine. Lastly, we suggest some potential directions for the development of SPS for extensive utilization in biomedicine.

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Xuejing Jia

Xuejing Jia received his BS degree (2011) in Biology and his MS degree (2014) in Botany from Sichuan Agricultural University, Ya’an, Sichuan, China. He is currently a PhD student in the State Key Laboratory of Quality Research in Chinese Medicine and Institute of Chinese Medical Sciences at University of Macau, Taipa, Macau, China. His research interests focus on the structural characterization and biological activities, including the anticancer, anti-inflammatory, and immunomodulatory activities, of polysaccharides from medicinal herbs.

Meiwan Chen

Meiwan Chen is an Assistant Professor at the University of Macau. She received her PhD degree from Sun Yat-sen University in 2010. She worked as a visiting scholar from 2009 to 2010 at the University of Mississippi (US). Her research focuses on biomaterials and nanomedicine, especially on the development of targeted drug delivery systems for cancer therapy. She has published more than 70 peer-reviewed SCI papers in top-tier pharmaceutical and biomaterial journals.

Jian-Bo Wan

Jian-Bo Wan is currently an Assistant Professor at the Institute of Chinese Medical Sciences (ICMS) and State Key Laboratory of Quality Research in Chinese Medicine, University of Macau (UM). He received his Ph.D. in Biomedical Sciences from the University of Macau in 2008, and completed his post-doctoral program at the Laboratory for Lipid Medicine and Technology in Massachusetts General Hospital, an affiliated hospital of the Harvard Medical School, from 2009–2011. His main research interests are the pharmacological study of Chinese Medicines for the prevention of metabolic diseases, including alcoholic fatty liver and atherosclerosis, and UPLC/Q-TOFMS-based metabolomic studies on Chinese Medicines and metabolic diseases.

Huanxing Su

Huanxing Su is an Associate Professor at the Institute of Chinese Medical Sciences, University of Macau. He obtained a PhD from the University of Hong Kong (HKU) and worked as a postdoctoral fellow at HKU. His research mainly focuses on using in vitro models to elucidate the molecular mechanisms for controlling the neural differentiation of human embryonic stem cells and human induced pluripotent stem cells (iPSCs); using stem cells, especially patient-derived iPSCs, as drug screening models for neurodegenerative diseases; and using nanomaterials and natural products to treat neurological disorders. He has published more than 65 peer-reviewed research articles in well-reputed journals.

Chengwei He

Chengwei He has been an Assistant Professor at the State Key Laboratory of Quality Research in Chinese Medicine and Institute of Chinese Medical Sciences at University of Macau since 2011. Dr He received his PhD degree in Biochemistry and Molecular Biology from Sun Yat-sen University of Medical Sciences in 2000. He completed his post-doctoral research at Harvard University-Massachusetts General Hospital in 2011. He then temporarily worked in the EMD Serono Research Institute at Merck KGaA in 2011. Dr He’s main research interests are the antitumor activities of traditional Chinese medicinal herbs and cellular oxidative stress adaptive responses.

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1. Introduction

Soybeans (Glycine max) are traditionally used to provide protein or oil for human consumption. A famous food made from soybean, tofu, has attracted much attention because it can constantly provide both calcium and protein for our diet. During the formation of tofu, the soybean curd residue, namely okara, is the dominating surplus material and it is always regarded as waste.¹ Okara has abundant active substances, such as protein, dietary fibre, mineral matter and oligosaccharides.² To reduce the cost and energy waste, okara is widely reused as a resource for polysaccharides, similarly as for soybean meal and soybean cotyledon. Intake of soybean polysaccharide (SPS) is likely to decrease human plasma cholesterol levels.³ So far, no research has found that SPS may cause adverse biological effects to human beings. Therefore, SPS is perfect for applications in the food service industry, such as enhancing the stability of beverages, increasing the emulsifying property of an acidic solution, and utilization as a biodegradable film. Additionally, SPS is increasingly used for the purposes of anticancer treatment or immunoregulation. However, the above mentioned properties of SPS are barely reviewed in recent literature. In order to enable better development and more convenient use of this unique, natural polysaccharide in the broader fields of the food industry and pharmaceutical chemistry, there is a pressing demand to comprehensively and efficiently analyze the properties of SPS from various angles.

Hence, in this concise review, we will firstly summarize the practical extraction methods for the preparation of SPS, followed by the introduction of the major structural characteristics which are desirable in food applications, and the detailed emulsification, anticancer, and immunoregulatory properties of this special polysaccharide. Finally, we will highlight the current potential applications that employ the advantages of SPS and its possible biological functions.

2. Extraction of SPS

SPS can be extracted from various resources, including common soybean seed, soybean meal, and okara. Among them, okara, the residue after oil or protein extraction from soybeans, is the most economic raw material for the extraction of SPS, particularly in East Asia. Okara contains various nutrients, including protein, dietary fiber, and some oligosaccharides.⁴ Owing to its valuable nutrients, especially the polysaccharide component, okara is increasingly used in food production. Black soybean (Glycine max (L.) Merr.), another species of legume commonly used in the oriental diet, is also an important source of SPS.⁵ Consequently, these rich resources solidly guarantee the sustainable production of SPS.

The optimization of the extraction solutions and procedures in the various extraction processes is critical for the yield of SPS. The methods for extracting SPS have been increasingly diversified over the last two decades. Fig. 1 shows a major schematic illustration of the technological processing of SPS. Many options can be selected for maximizing the yield of SPS. Besides the mentioned factors in Fig. 1, there are others that can also profoundly affect the yield of SPS, such as pH, the origin of the materials and the freshness of the raw materials.

Fig. 1 Schematic technical route for the effective extraction and refining of SPS from soybeans. BBD, Box–Behnken design; CCD, central composite design.

Many attempts have been made to improve the yield of SPS. Yamaguchi et al.⁶ extracted the polysaccharide from okara using a hexametaphosphate solution as the extractant, and then the polysaccharide was purified using DEAE cellulose column chromatography with a carbonate buffer. When using alkaline water as the extractant, the optimal extraction conditions are: pH 11.0, extraction temperature 120 °C, ratio of solid to liquid 1 : 20 (g : mL), and extraction time 2 h; the polysaccharide yield is 16.24%.⁷ However, for acidic water, the best parameters are: pH 4.0, extraction temperature 118 °C, ratio of material to liquid 1 : 30, and extraction time 2.5 h; the final yield is 37.88%.⁸ In order to reduce the dissolution rate of protein, an organic acid solution is used to extract SPS from soybean dregs. The optimal process parameters are determined, including a tartaric acid aqueous solution, pH 3.8, an extraction temperature of 110 °C and an extraction time of 1.5 h; resulting in a yield of 27.65%.⁹

To maximize the yield of SPS, an ultrasonic-assisted extraction method is applied. The optimum ultrasonic extraction parameters are: ultrasonic treatment time 20 min, ultrasonic power 200 W, bath temperature 90 °C, and solid to liquid ratio 1 : 25 (g : mL); the extraction yield is 1.87%.¹⁰ When some of the extraction parameters are modified, such as the extraction pH (4.5), solid to liquid ratio (1 : 20), and the ultrasonic treatment time (40 min), the yield is 8.82%.¹¹ Multiple approaches can be integrated in one assisted extraction. Enzymatic hydrolysis is used as well, and the optimal crucial technological parameters are: ultrasonic treatment time 30 min, ultrasonic power 200 W, and solid to liquid ratio 1 : 25 (g : mL); hydrolysis temperature 50 °C, hydrolysis duration 40 min, enzyme dosage 1.5%, and pH 5.0. Under such an environment, the polysaccharide yield is 12.23%.¹²

On the other hand, soybean meal is degraded using double enzymes in combination, an acidic protease and a flavor enzyme. The optimum conditions are: pH 6.0, enzymatic hydrolysis time 6 h, solid to liquid ratio 1 : 20, and the amounts of the protease and flavor enzyme are 10% and 8%, respectively; the yield of SPS is 9.28%.¹³ The microwave-assisted extraction of SPS from soybean meal has been optimized. The optimum conditions are: pH 8.0, ratio of water to raw material 1 : 6 (g : mL), microwave time 2.6 min, and microwave power 380 W; finally, the yield of SPS is 5.86%.¹⁴ Soybean dregs are hydrolyzed with a cellulase preparation, using microwave assistance. The optimal procedure for the extraction uses: cellulase dosage 1.5%, pH 5.0, hydrolysis temperature 50 °C, hydrolysis time 40 min, material to water ratio 1 : 15 (g : mL), microwave power 600 W, microwave time 7 min; the maximum yield is up to 15.85%.¹⁵

Another extraction solution, sub-critical water, is well employed; its best conditions are: water temperature 150 °C, solid mass to water ratio 1 : 35 (g : mL), extraction time 11 min. Under these conditions, the yield is 22.8%.¹⁶ To maximize the yield of the black soybean polysaccharides, Box–Behnken design is applied during the process of extraction. Liu et al.¹⁷ obtained the optimal extraction conditions: ratio of water to material 20 mL g⁻¹; extraction time: 6.4 h; and extraction temperature: 92 °C. Under these optimal conditions, the yields of crude SPS reach 2.56%. Taken together, various assisted extraction methods are truly benefiting both the enhancement of the yield of SPS and the reduction of the processing time.

3. Structural characterization of SPS

The SPS structure is gradually being understood by researchers. The fundamental properties of SPS are listed in Table 1. SPS was extracted with water at 60 °C for 4 h, fractionated with a series of solutions with different concentrations of sodium hydroxide, and six constituents of fucose, rhamnose, xylose, arabinose, galactose, and galacturonic acid, respectively, were identified.²⁵ Arabinogalactan, the major component of soybean seed polysaccharides, consists of arabinose and galactose residues and has an average molecular weight of 330 kDa.²⁶ Moreover, arabinogalactan, derived from defatted and deproteinized soybean cotyledon meal, has a backbone chain of 1 → 4 linked β-D-galactopyranose residues and a side chain containing, in general, two L-arabinofuranose residues with a 1 → 5 linkage.²⁷ Furthermore, an arabinan, found from the previous polysaccharides, is methylated and forms alditol acetates. Analysis with gas chromatography mass spectrometry (GC-MS) reveals a similar structure to other arabinans.²⁸

Table 1 The fundamental characterization of SPS isolated from soybean meal, soybean cotyledon, okara, and black soybean

Source	Extraction	Fraction name	Molecular weight	Uronic acid content (%)	Monosaccharide composition (molar ratio)							References
					Arabinose	Rhamnose	Galactose	Glucose	Mannose	Xylose	Fucose
a Chelating agent: 0.05 mol L^−1 1,2-diaminocyclohexane-N,N,N′,N′-tetraacetic acid (CDTA) and 0.05 mol L^−1 NH4-oxalate in 0.05 mol L^−1 NaAc-buffer; ND, not detected.
Soybean meal	Chelating agent^a, 70 °C pH 5.2 for 1 h	ChSS	About 10^6 Da	53%	16.65	1.10	20.54	0.56	0.56	ND	ND	18
	0.05 mol L^−1 NaOH 2 °C for 1 h	DASS	About 10^6 Da	10%	15.99	1.10	21.09	0.56	0.56	ND	ND
Soybean cotyledon	120 °C pH 5 for 1.5 h	SSPS	1.14 × 10^5 Da	23.4%	14.25	1.37	23.04	1.17	ND	3.73	2.13	19
	100 °C for 1 h	A1-β	2 × 10^6 Da	ND	1.00	0.05	1.47	0.04	0.01	0.03	0.02	20
Okara	120 °C pH 3 for 2 h	SSPS-L	ND	27.5%	10.19	2.36	26.81	0.89	ND	1.00	0.91	21–23
	130 °C pH 4–5 for 3 h	SSPS-H	ND	25.6%	10.39	3.08	26.48	1.11	ND	1.47	0.79
	120 °C pH 4–5 for 2 h	SSPS-M	ND	23.9%	13.39	2.25	26.20	0.61	ND	0.80	1.46
	0.05 mol L^−1 NaOH	0.05 MSF	ND	14.7%	18.00	1.30	26.30	3.40	1.50	5.50	2.30
	1 mol L^−1 KOH	1 MSF	ND	4.4%	16.90	ND	16.40	10.20	15.00	28.50	1.50
	4 mol L^−1 KOH	4 MSF	ND	5.1%	13.90	ND	18.60	12.90	5.00	36.70	2.20
	90 °C pH 13 for 3 h	1	3.96 × 10^5 Da	ND	18.3	3.2	53.6	6.4	ND	1.6	1.5
	80 °C pH 12 for 3 h	2	4.11 × 10^5 Da	ND	18.2	2.5	54.3	7.2	ND	1.9	1.1
	70 °C pH 12 for 2 h	3	4.37 × 10^5 Da	ND	19.4	2.7	54.9	6.8	ND	1.3	0.6
	60 °C pH 12 for 2 h	4	4.62 × 10^5 Da	ND	17.2	3.4	55.2	7.3	ND	0.9	1.8
	60 °C pH 11 for 1.5 h	5	4.68 × 10^5 Da	ND	18.1	3.3	55.7	6.5	ND	2	0.9
	50 °C pH 9 for 1.5 h	6	4.89 × 10^5 Da	ND	18.8	3.6	56.6	6.6	ND	1.4	1.1
Black soybean	60–100 °C for 3–7 h	BSPS-1	1.95 × 10^5 Da	0.14%	1.79	1.00	2.59	26.54	1.01	ND	ND	17, 24
	60–100 °C for 3–7 h	BSPS-2	ND	2.98%	8.10	4.80	9.15	13.38	1.00	ND	ND
	60–100 °C for 3–7 h	BSPS-3	1.88 × 10^5 Da	9.13%	16.80	3.60	33.66	ND	1.00	ND	ND

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Soybean pectic polysaccharides consist of two types of regions: galacturonan and rhamnogalacturonan. The galacturonan regions appear at both the reducing and non-reducing ends of the chains, while the latter regions link to the side chains.⁶ SPS from okara has a pectin-like structure. Its core backbone contains L-rhamnose and D-galacturonate residues equally, consisting of a -4)-α-D-GalA-(1 → 2)-α-L-Rha-(1- and -4)-α-D-GalA-(1- repeating units, respectively (Fig. 2a).²⁹ The SPS of soybean cotyledons contains the acidic polysaccharides galacturonan (GN), rhamnogalacturonan (RG), and xylosyl oligosaccharides with (β-D-Xyl)₇ or (β-D-Xyl)₄ residues at the C-3 site.³⁰ The side chain of β-1,4-galactans is branched with fucose and arabinose residues. For GN there are about 4–10 residues at the C-3 side of the galacturonates, while for RG there are about 43–47 residues on the C-4 side (Fig. 2b and c).³¹

Fig. 2 The possible structure residues of SPS in alkaline conditions (a),^19,32 and the structure model of SPS that possesses a globular form with arabinan and/or galactan chains that can be digested with RGase, AFase, and GPase (b and c).³³ GalUA, galacturonic acid; Rham, rhamnose; Ara, arabinose; Gal, galactose; pectinases (polygalacturonase (PGase) and rhamnogalacturonase (RGase)) or hemicellulases (galactosidase (GPase) and arabinosidase (AFase)). Reproduced with permission from ref. 19. Copyright 1999, Elsevier.

Soybean meal, the byproduct of oil extraction, is rich in proteins and polysaccharides. Two similar soybean meal polysaccharides, ChSS (chelating agent soluble solids), extracted with a chelating agent, and DASS (dilute alkali soluble solids), extracted with dilute alkali, were sequentially fractionated using anion exchange chromatography.¹⁸ To explore the detailed characterization of ChSS, the degradation of the cell wall by enzymes, endo-galactanase, endo-arabinanase, rhamnogalacturonan hydrolase, rhamnogalacturonan acetylesterase and polygalacturonase-1, is performed in a rather specific way, which indicates that ChSS are likely to have a highly substituted pectic structure.³⁴

The SPS from soybean cotyledons that have been digested by five enzymes, with a side chain of arabinan and galactan and a backbone mainly of polygalacturonase and rhamnogalacturonase, enhances the stability of acidic beverages.³³ Compared with the SPS treated with GPase (galactosidase), the authors indirectly confirm that the native SPS has a galactan side chain and presents a branched globular conformation.³⁵ Combining the analysis from the NMR spectra and methylation, the HSQC (heteronuclear single quantum coherence) spectra of BSPS-1 (purified fraction of black soybean polysaccharide 1) and BSPS-3 (purified fraction of black soybean polysaccharide 3) is shown in Fig. 3a and b as an example of compositional structural analysis. Liu et al.²⁴ identified two novel soluble polysaccharides (BSPS-1 and BSPS-3) from black soybeans. BSPS-1 is a linear (1 → 6)-α-D-glucan of 195 kDa, while BSPS-3 is a type II arabinogalactan of 188 kDa (Fig. 3c and d). In conclusion, SPS possesses special structures that contain galacturonan and rhamnogalacturonan, suggesting its promising applications in the food industry and biomedical areas.

Fig. 3 HSQC (heteronuclear single quantum coherence) spectra of BSPS-1 (a) and BSPS-3 (b) in D₂O at 25 °C. H1–C1 represents the cross peak between H-1 and C-1 of the →6)-α-D-Glcp-(1→ residue, etc. A1 represents the cross peak between H-1 and C-1 of residue A, etc. A, B, C, D, E, F, and G represent the residues of α-L-Araf-(1→, →5)-α-L-Araf-(1→, →3,6)-β-D-Galp-(1→, →3)-β-D-Galp-(1→, 4-O-Me-β-D-GlcAp-(1→, →2)-α-L-Rhap-(1→, and →6)-α-D-Glcp-(1→, respectively. Possible structures of BSPS-1 (c) and BSPS-3 (d).²⁴ Reproduced with permission from ref. 24. Copyright 2015, American Chemical Society.

4. Potential applications of SPS

4.1 SPS in the food industry

As a food additive, SPS shows excellent stabilization and emulsification behaviors and is mainly used by food researchers to improve the stability of beverages and increase the emulsifying properties of oil droplets in response to diverse environmental challenges. Notably, SPS-based formulations could have extensively enhanced health benefits. All of these promising applications of SPS may be attributable to its known physicochemical features, as summarized in Table 1 and elaborated below.

4.1.1 Emulsifying properties of SPS. In an aqueous environment, SPS, one of the most abundant components of soybean byproducts, strongly endures the usual sterilization and acidic conditions.¹⁹ SPS is a perfect candidate for an interfacial film because of its high water solubility, low bulk viscosity and excellent thermostability.²¹ Even in acidic and hot water conditions, within the pH range of 2–6 and the temperature range of 40–120 °C, water-soluble polysaccharides that mainly consist of rhamnogalacturonan remain as a fluid.³⁶ Under the conditions of 4 °C for 24 h, 0.5% SPS increases the rate constant of 5% starch retrogradation, and meanwhile declines the saturated dynamic storage modulus of the composite system.³⁷ After alkali treatment and subsequent acidic extraction, the SPS with a lower degree of esterification exhibits highly emulsifying properties for oil-in-water and stabilizing abilities in acidic milk beverages.²³

SPS has been demonstrated to increase the emulsifying properties and then to form a unique film, which can prevent aggregation caused by steric or electrostatic repulsion among the various oil droplets.²¹ To understand which chains of polysaccharides are responsible for these strong emulsifying properties, SPS was digested by pectinases and hemicellulases. It was found that sugar chains, β-galactan and α-arabinan, play a notable role in the emulsifying capabilities and stabilities, which provides a promising utilization for SPS in beverages.³⁸ The similar findings also proved that SPS could mutually prevent the aggregation of casein micelles.³⁹ In comparison with the necessary concentration of sugar beet pectin (1.5%) and gum arabic (10%), SPS is required in a moderate amount (4%) to surround the oil droplets and stabilize the emulsion of oil-in-water.⁴⁰ To broaden its function, the SPS was phosphorylated and formed a high molecular mass complex, leading to a functional stabilization of acid particle dispersion within the pH range of 4–4.8.⁴¹

The fractions of SPS, HMF (high molecular weight) and LMF (low molecular weight), possess diverse functions. The HMF is used to emulsify oil–water droplets and stabilize α-casein dispersions while the LMF is better to protect emulsified lipids from oxidative aggression.⁴² Compared with the stabilization of the LMF of soybean cotyledons, the HMF, with larger electrostatic and steric repulsive force, can clearly disperse milk proteins.⁴³ The presence of impure proteins in the LMF would increase its particle size and then change its functional performance after heating at 90 °C for 30 minutes.⁴⁴

4.1.2 Interactions between SPS and other substances. SPS, absorbed on the droplet surface, can improve the emulsions of lactoferrin-coated oil and thus prevent lipid oxidation.⁴⁵ In addition, coats made from SPS, based on the layer by layer electrostatic deposition of the orange oil, have enhanced resistance against environmental stresses such as various ions, pH, and light.⁴⁶

Adding polysaccharides to a soybean protein emulsion can decrease the initial droplet size, thereby improving their stability.⁴⁷ A combination of SPS and dSWP (denatured soy whey protein) at a ratio of 1.5 : 1, forming a dSWP–SPS layer and covering the oil droplets, promotes emulsion stability and prevents the coalescence and phase separation of oil-in-water.⁴⁸ Conjugating β-lactoglobulin, a whey protein of 18 kDa, with SPS in a special way would enhance the emulsifying property of this complex.⁴⁹ SPS fractions, a mixture of low and high-molecular-weight components, encapsulated with linoleic acid can increase the antioxidative capacity of these microcapsules and retard the oxidation process.⁵⁰ Cross-linking SPS with sodium hexametaphosphate via an esterification reaction under acidic conditions can improve the stability of isolated soy protein when stored at 4 °C.⁵¹

Dietary fiber in food has important health benefits, such as reducing blood cholesterol, decreasing the risk of diabetes, and improving bowel movement. SPS extracted and refined from okara is incorporated into thickened milkshake-style beverages. This popular beverage containing 0.015% κ-carrageenan, namely a 4% SPS-fortified dairy beverage, is favored by ordinary consumers because it increases their soluble dietary fiber intake.⁵² The combination of SPS and sodium carboxymethyl cellulose at a ratio of 3 : 1 effectively prevents the aggregation of casein and exhibits strong stabilization in acidified skimmed milk drinks.⁵³

SPS can be used as an additive to improve the quality and value of food. The presence of SPS from soybean cotyledons can reduce the viscosity of gelatinized starch, therefore, it is used to cook rice or noodles, which prevents them from adhering to each other.⁵⁴ Lactose, as a food additive, is widely used in infant formulas, protein powders, and candies. However, lactose can easily absorb moisture and crystallize. Mixing with soluble soybean polysaccharide at 10 g/100 g, the crystallization of spray-dried lactose powder can be remarkably delayed.⁵⁵ Anionic SPS-coated droplets and SPS-coated β-carotene droplets, are stabilized in oil–water emulsions with an improved viscosity and consistency index.⁵⁶ A fraction of SPS from okara, glycosidoprotein with molecular weight of 14–370 kDa, is better than that from the soybean hull, acidic heteropolysaccharides with molecular weight of 45–150 kDa, in terms of its emulsifying performance and in vitro bile acid binding activity.⁵⁷

4.2 SPS in biomedicine

The antioxidant capacity and stabilization of SPS is positively related to its concentration. SPS can scavenge hydroxyl radicals and keep stable for a long time. The inhibitory rate of 0.08% SPS against oxidation is above 95% within 200 s, surprisingly, and 0.2% SPS could keep this status for 20 d.⁵⁸ The soluble polysaccharide fractions of okara, namely 0.05 MSF (0.05 mol L⁻¹ NaOH soluble polysaccharides), 1 MSF (1 mol L⁻¹ KOH soluble polysaccharides) and 4 MSF (4 mol L⁻¹ KOH soluble polysaccharides), supposedly possess a β-glycosidic linkage, strongly scavenge ABTS (2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid)) radicals, and potently reduce Fe(III) to Fe(II).²² However, crude polysaccharide from black soybeans possesses higher superoxide anion and DPPH (2,2-diphenyl-1-picrylhydrazyl) radical scavenging abilities than the purified fractions (BSPS-1, BSPS-2 and BSPS-3).¹⁷ Soybean polysaccharide degraded with hydrogen peroxide (DPS), with a smaller molecular weight about 10.2 kDa, efficiently inhibits the formation of calcium oxalate crystals, therefore, it highly reduces the risk of kidney stone formation. In addition, DPS can distinctly weaken the external oxidative damage of the renal epithelial cells of the Africa green monkey, resulting in increased cell viability.⁵⁹

Black soybean polysaccharides, purified by column chromatography, stimulated the production of granulocyte colony-stimulating factor in peripheral blood mononuclear cells, mediated via the activation of the PI3K (phosphoinositide 3-kinase), ERK (extracellular signal-regulated protein kinase), PKC (protein kinase C), and NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) signaling pathways.⁶⁰ PSBS, the polysaccharides of the black soybean, accelerate myelopoiesis in vitro and increase the levels of various hematopoietic growth factors from spleen cells, and, in vivo, reconstitute bone marrow after 5-flurouracil- and irradiation-induced damage.⁶¹ SPS as the excipient is added in Epimedium granules with the proportion of 1 : 3.5, endowing this type of granule with greater granulation, dissolubility, and applicability.⁶²

Pre-treating mouse spleen lymphocytes with SPS for 2 h before X-ray radiation protects the cells from DNA damage and increases cell viability.⁶³ SPS shows antitumor activity via regulating the immune functions of S₁₈₀-bearing mice, namely improving the phagocytosis and the production of NO in macrophages, and greatly increasing the number of B-lymphocytes.⁶⁴ Additionally, SPS could increase the CD₄⁺ T cell numbers and the ratio of CD₄⁺/CD₈⁺ cells, and the level of IL-2 in serum. Obviously, SPS could notably stimulate T-lymphoid cell proliferation and IL-2 secretion.⁶⁵ A combination of cyclophosphamide and SPS shows a better inhibitory effect on tumor growth and improves the thymus and spleen indices and IL-2 secretion, suggesting a synergistic anticancer effect and reduction of the toxicity of cyclophosphamide.⁶⁶

At pH 6, SPS exhibits strong inhibitory effects on Escherichia coli, Staphylococcus aureus, Aspergillus niger and Penicillium chrysogenum, and the minimal inhibitory concentrations are 8, 6, 1, and 1 mg mL⁻¹, respectively.⁶⁷ Treatment with 5% SPS in vitro promotes the growth of Bifidobacterium longum, Bifidobacterium and Lactobacillus. Similar results can be seen with fructooligosaccharides.⁶⁸ Emulsifying both thyme oil and soluble soybean polysaccharide results in better antimicrobial activities against Listeria monocytogenes Scott A, Salmonella enteritidis and Escherichia coli O157:H7 versus thyme oil alone.⁶⁹

4.3 SPS in biomaterials

A film based on SPS, a new biodegradable edible biomaterial, is a promising raw material commercially utilized for food package.⁷⁰ It has been successful gelatinized, as shown in Fig. 4. Essential oils from Zataria multiflora Boiss or Mentha pulegium are incorporated with SPS to form a sandwich-like film, which promotes the polysaccharide interaction, reduces water solubility, and increases elongation at break remarkably.⁷¹ On the basis of these properties, this active edible film, in addition, inhibits the growth of Gram negative and positive bacteria in a dose dependent manner, and potently scavenges free radicals, especially for Zataria multiflora Boiss.⁷² This composite film is intensively recommended for use in food packaging.

Fig. 4 Scanning electron microscopy (SEM) images of the surface and cross-section of SSPS films plasticized with 0% and 30% (w/w) glycerol. Atomic force microscopy (AFM) topographic images of SSPS films plasticized with 0% and 30% (w/w) glycerol. An image of the appearance of a biodegradable edible film based on the formation of 30% (w/w) glycerol.⁷⁰ Reproduced with permission from ref. 70. Copyright 2013, Elsevier.

To highly reduce nutrient cost and to utilize its maximum direct value, soybean curd residue is reused as the nutrient source for the solid state fermentative production of polysaccharides.⁷³ In comparison with normally submerged fermentation, it is not only more time efficient to ferment polysaccharides from okara but also lower in cost. Li et al.⁷³ reported that polysaccharides derived from Wolfiporia extensa (Peck) Ginns, fermented from okara, showed positive antioxidant abilities against DPPH and ABTS radicals.

5. Perspectives

Based on the physicochemical properties discussed above, SPS shows remarkable advantages in potential applications as a food additive, biomedicine and biomaterial, which motivates us to explore more possible applications in these fields. Future studies may be primarily focused on the following directions.

Firstly, the application of SPS in the food industry could be largely broadened owing to its unique structural features and chemical properties. Actually, SPS has already been extensively applied as a modifying agent. For example, it has been demonstrated that under acidic conditions SPS can disperse a stabilized protein solution. In this case, adding a small amount of SPS to a favorable milk beverage can greatly lower its viscosity.⁷⁴ Carboxymethyl SPS, which dissolves in an alkali solution but not in an acidic solution, inhibits the growth of Bacillus subtilis and Bacillus cereus.⁷⁵ However, little attention is paid to the adverse biological effects of modified SPS. We do not know whether these refined natural or modified SPS derivatives are harmful to the health of human beings and livestock. Therefore, more comprehensive studies are highly in demand to investigate the potential influences on health.

Secondly, it is interesting to devise novel biodegradable or edible materials based on SPS, which has been initially achieved and has shown the possibilities for films. For instance, the composite films, containing 12.5% SPS, show good water solubility, incredible tensile strength and an elongation rate at breaking, and they are supposedly non-toxic and eco-friendly as well, which are extraordinary features for food packaging.⁷⁶ The preliminary findings have provided possible practical information for the utilization of SPS-based biodegradable or edible films. However, additives, such as essential oils, sucrose, etc., need to be further optimized to achieve the optimal gelation abilities, including the gel strength, gel elasticity and adhesion strength.⁷⁷

Thirdly, the application of SPS has exhibited its huge potential in the area of biomedicines, particularly for the treatment of cancer and immunoregulation. However, only a few studies (as mentioned in this paper) have been involved in these interesting fields. One of the most distinct functions of polysaccharides is immunomodulation, which might be closely related to their anticancer activity. At a dosage of 50–400 μg mL⁻¹, SPS exhibits great immunomodulatory activity in vitro, dramatically stimulating spleen lymphocyte proliferation, observably increasing the phagocytosis of macrophages, and enhancing macrophage NO production.⁷⁸ Moreover, SPS could potentially attenuate the toxicity of anticancer chemical compounds. Injecting a dose of SPS into S₁₈₀ sarcoma mice could significantly improve the parameters of immune functions, including the number of leukocytes, the level of TNF-α (tumor necrosis factor alpha), and the ratio of CD₄⁺/CD₈⁺, compared to cyclophosphamide used alone.⁷⁹ Clearly, these attempts shed light on the use of SPS for a wide range of biomedical applications. More studies are required to investigate the details of the anticancer and immunoregulatory activities of SPS in animal models, and particularly in clinical trials. Whether SPS is beneficial to the prevention or treatment of oxidation- or inflammation-related diseases, such as neurodegenerative diseases, diabetes mellitus, or renal diseases, is highly interesting to be explored since SPS shows strong antioxidative activities as well.

In summary, a growing number of studies have indicated that SPS is a promising candidate for food industry, biomedical, and biomaterial applications, in which further potential is under exploration. However, the risk evaluation from a scientific perspective is still absent, especially for cross-linked SPS. In this regard, substantial systemic toxicological investigations, both in vitro and in vivo, are highly in demand.

6. Conclusions

In conclusion, SPS could be effectively isolated from soybeans or okara using various extraction methods, including ultrasonic assistance, microwave assistance, enzymatic treatment, and subcritical water extraction, which all show better extraction efficiencies than hot water alone. Meanwhile, SPS, a linear chain of galacturonan and rhamnogalacturonan, is widely accepted as a food additive, showing its advantages in emulsifying and stabilizing an oil–water system. Moreover, SPS has potential in the area of biomedicine, with antioxidant activity, antimicrobial activity, and anticancer activity. Indeed, SPS can inhibit the growth of a tumor via regulating the immune function, such as increasing the level of NO and IL-2. Another promising application is to use SPS as a biodegradable material for food packaging and preservation. However, the potential risk or toxicity of SPS and its derivatives have not been reported yet. Thus, to better use SPS and its derivatives, comprehensive toxicological studies or risk assessments, both in vivo and in vitro, within the standard guidelines are highly in demand.

Conflict of interest

The authors declare no conflict of interest.

Acknowledgements

This study was supported by the Macao Science and Technology Development Fund (074/2013/A) and the Research Fund of the University of Macau (MYRG107(Y1-L3)-ICMS13-HCW, MYRG2015-00081-ICMS-QRCM).

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