WO2015101435A1

WO2015101435A1 - A method of making soymilk

Info

Publication number: WO2015101435A1
Application number: PCT/EP2014/073938
Authority: WO
Inventors: Jing Su; Fei Xue; Mo Li
Original assignee: Koninklijke Philips N.V.
Priority date: 2013-12-30
Filing date: 2014-11-06
Publication date: 2015-07-09

Abstract

This disclosure relates to a method of making soymilk comprising the step of subjecting the soymilk processing stream to a dialysis separation so as to separate Bowman- Birk inhibitor (BBI) from the soymilk processing stream, which consequently shortens the cooking time. The disclosure also relates to the equipment using said method and related food processor.

Description

A method of making soymilk

FIELD OF THE INVENTION

The present disclosure relates to a method of making soymilk comprising the step of subjecting the soymilk processing stream to a dialysis separation so as to separate Bowman-Birk inhibitor (BBI) from the soymilk processing stream, which consequently shortens the cooking time. The disclosure also relates to the equipment using said method and related food processor.

BACKGROUND OF THE INVENTION

Trypsin inhibitors (TI) are generally considered as the main anti-nutrients in soybeans. TI inhibits trypsin and chymotrypsin which are important digestive enzymes in animal to break down proteins into di- and tri-peptides. Unless thoroughly inactivated, TI could limit human and animal's growth as inferred from laboratory studies with animals [Phyllis, C. H.; Lyman, R. L., Relationship of Pancreatic Enzyme Secretion to Growth Inhibition in Rats Fed Soybean Trypsin Inhibitor. J. Nutrition, 1974, 61, 445-452]. Currently, it takes 15-20 min to inactivate 90% of TI activity at 95-100°C. This limits the speed of soymilk production at home, making it is impossible to produce soymilk in shorter time at this temperature.

There are two types of soybean trypsin inhibitors, Kunitz trypsin inhibitor (KTI) and Bowman-Birk inhibitor (BBI). BBI is comparatively small and rigid which is heat-stable in aqueous solution. It has a molecular weight of 7848 Daltons, 71 amino acids, and 7 disulfide bonds, with the tendency to self-associate. BBI possesses two independent sites of inhibition, one at Lys 16-Ser 17 binding trypsin and the other at Leu 43-Ser 44 binding chymotrypsin. KTI has a molecular weight of 20,083 Daltons, 181 amino acid residues, and two disulfide bridges. KTI only has one-active site (arginine at residue 63 and iso leucine at residue 64) which complexes with trypsin.

BBI is reported more difficult to be inactivated than KTI in aqueous solution. For example, when pure KTI and BBI solutions were heated at 100°C, it was found KTI lost its trypsin inhibitor activity (TIA) after 180 min while BBI retained over 75% of its TIA even after 360 min [DiPietro, C. M.; Liener, I. E., Heat inactivation of the Kunitz and Bowman-Birk soybean protease inhibitors. J. Agric. Food Chem. 1989, 37, 39-44]. As BBI inhibits both trypsin and chymotrypsin, Zhicun Xu et al. reported that chymotrypsin inhibitor activity (CIA) assay showed about 89% of the original CIA remained in soymilk after 100 °C for 15 min [Xu, Z. C, Chen, Y. M.; Zhang, C. M.; Kong, X. Z.; Hua, Y. F., The Heat- Induced Protein Aggregate Correlated with Trypsin Inhibitor Inactivation in Soymilk

Processing. J. Agric. Food Chem. 2012, 60, 8012-8019], which indicated that 89% of BBI retained after 15 min cooking. To sum up, BBI in soymilk is hard to be inactivated which results in the requirement of longer cooking time. There is a need in the art to separate or inactivate the BBI in a more efficient manner during the process of making soymilk so as to shorten the cooking time of soymilk.

SUMMARY OF THE INVENTION

The present disclosure provides a method of making soymilk comprising the step of subjecting the soymilk processing stream to a dialysis separation so as to separate BBI from the soymilk processing stream, which consequently shortens the cooking time of soymilk for TI A inactivation. The disclosure also relates to the equipment using said method and related food processor.

In one aspect, the present disclosure provides a method of making soymilk comprising the steps of:

(a) preparing a raw soymilk processing stream,

(b) subjecting the soymilk processing stream to a dialysis separation so as to separate

Bowman-Birk inhibitor (BBI) from the soymilk processing stream, and

(c) cooking the soymilk processing stream;

wherein step (b) and step (c) can be carried out at the same time, or step (b) can be carried out prior to or subsequent to step (c).

In one embodiment, the step (b) further comprises subjecting the soymilk processing stream to one or more chromatographic separation techniques which are performed prior to and/or subsequent to the dialysis separation. Preferably, the

chromatographic separation techniques are selected from ion exchange chromatography, affinity chromatography and hydrophobic interaction chromatography.

In one embodiment, during the method of making soymilk as mentioned above, the pore size of a dialysis membrane used in the dialysis is 8,000 Da-14,000Da.

In one embodiment, the step (b) is carried out via an equipment comprising a water tank and a dialysis device. In one embodiment, the equipment further comprises a device for one or more chromatographic separation techniques. Preferably, the chromatographic separation techniques are selected from ion exchange chromatography, affinity chromatography and hydrophobic interaction chromatography.

In one embodiment, the pore size of a dialysis membrane used in the dialysis device is 8,000 Da-14,000Da.

In one embodiment, the dialysis device further comprises an accelerator for dialysis.

In a further embodiment, the accelerator is selected from stirring system, ultrasonic system, heating system, water refreshing system and vibration system.

In one aspect, this disclosure provides a food processor comprising:

a machine head connected with a grinding blade;

a grinding container;

a solution switching control;

a cooking container comprising the equipment for separating Bowman-Birk inhibitor (BBI) from a soymilk processing stream as mentioned above;

a heat plate;

an electric connector;

wherein the solution switching control and the electric connector are fixed; the machine head can plug in the electric connector; the grinding container and the cooking container comprising the equipment for separating Bowman-Birk inhibitor (BBI) from a soymilk processing stream are detachable; the machine head, the solution switching control, the heat plate and the electric connector are connected, and wherein a food processing stream is cooked in the equipment for separating Bowman-Birk inhibitor (BBI) from a soymilk processing stream inside the cooking container and then transferred to a cup.

In one embodiment, the grinding container is configured to be inside the cooking container.

The food processor in the present disclosure can be selected from a soymilk maker, a tofu making machine and a soy food processor.

In one aspect, the present disclosure provides a method for separating

Bowman-Birk inhibitor (BBI) from a soy processing stream comprising the step of subjecting the soy processing stream to a dialysis separation, and optionally, one or more

chromatographic separation techniques which are performed prior to and/or subsequent to the dialysis separation, wherein the pore size of a dialysis membrane used in the dialysis separation is 8,000 Da-14,000 Da.

BRIEF DESCRIPTION OF THE DRAWINGS

The following drawings illustrate the embodiments, and do not limit the scope of invention defined in the claims.

Figure 1. Schematic representation of the principle of dialysis: only protein smaller than the pore size can pass the membrane.

Figure 2. Schematic flow sheet of three embodiments of soymilk making process.

Figure 3. Schematic representation of one embodiment of dialysis device for soymilk.

Figure 4. Schematic representation of one embodiment of soymilk maker. Figure 5. Schematic representation of another embodiment of soymilk maker. Figure 6. Trypsin inhibitor activity (TIA) of soymilk with different treatments (Raw soymilk and Raw soymilk with dialysis).

Figure 7. Chymotrypsin inhibitor activity (CIA) of soymilk with different treatments (Raw soymilk and Raw soymilk with dialysis).

Figure 8. Dissolved protein content in soymilk with different treatments (Raw soymilk, Dialysis raw soymilk, Dialysis raw soymilk with cooking, Cooked Soymilk by Philips HD2076, and Cooked Soymilk by Joyong DJ13B-D08). The weight percent of dissloved protein content was calculated based on the weight of soymilk.

DETAILED DESCRIPTION OF THE INVENTION

Before the present method, device and equipment are described in details, it is to be understood that this invention is not limited to the particular configurations, method steps, and arrangements disclosed herein as such configurations, steps and materials may vary somewhat. It is also to be understood that the terminology employed herein is used for the purpose of describing particular embodiments only and is not intended to be limiting since the scope of the present invention will be limited only by the appended claims and equivalents thereof.

When introducing elements of the present invention or the preferred embodiments(s) thereof, the articles "a," "an," "the" and "said" are intended to mean that there are one or more of the elements. The terms "comprising," "including" and "having" are intended to be inclusive and mean that there may be additional elements other than the listed elements.

As various changes could be made in the above methods, devices and equipments without departing from the scope of the invention, it is intended that all matter contained in the above description and in the examples given below, shall be interpreted as illustrative and not in a limiting sense.

In describing and claiming the device and method, the following terminology will be used in accordance with the definitions set out herein.

The term "processing stream" as used herein refers to the secondary or incidental product derived from the process of refining a whole legume or oilseed, including an aqueous stream, a solvent stream, or a reconstituted from dried (e.g., spray dried) stream, which includes, for example, an aqueous soy extract stream, an aqueous soymilk extract stream, an aqueous soy whey stream, an aqueous soy molasses stream, an aqueous soy protein concentrate soy molasses stream, an aqueous soy permeate stream, an aqueous tofu whey stream, and additionally includes soy whey protein, for example, in both liquid and dry powder form, provided that BBI is required to be removed from said processing stream.

When making soymilk, trypsin inhibitors (TI) need to be inactivated. But the process of inactivating TI takes long time. Bowman-Birk inhibitor (BBI), as one of TI, accounts for 20% to 40% activity in soy milk. It is the main bottleneck for fast making of soymilk. Described herein are novel processes for separating BBI from the soymilk processing stream, which consequently shortens the cooking time. This method can be applied in soymilk maker, tofu making machine and soy food processor. For example, the processes of the present disclosure comprise a dialysis selected and designed to provide the remove of BBI from the soymilk processing stream.

Dialysis is a protocol for separating small, unwanted compounds from macro molecules in solution by selective and passive diffusion through a semi-permeable membrane. A sample and a buffer solution are placed on opposite sides of the membrane. Due to the gradient between the sample and the buffer, the molecules in the sample attempt to flow through the membrane but only those smaller than the pore size can actually leave the sample and passing the membrane, reducing the concentration of those molecules in the sample (Fig 1).

In one aspect, the present disclosure relates to a method of making soymilk comprising the steps of:

(a) preparing a raw soymilk processing stream, (b) subjecting the soymilk processing stream to a dialysis separation so as to separate

Bowman-Birk inhibitor (BBI) from the soymilk processing stream, and

(c) cooking the soymilk processing stream.

In one embodiment, step (b) and step (c) can be carried out at the same time. In one embodiment, step (b) can be carried out prior to step (c). In another embodiment, step (b) can be carried out subsequent to step (c).

In one embodiment, step (a) comprises the steps of: washing beans, optionally subjecting the beans to other treatments, and adding water to the beans and grinding the beans to produce the raw soymilk processing stream.

In one embodiment, the method of making soymilk further comprises step (d) filtering the soymilk processing stream. In a further embodiment, step (b) and step (c) can be carried out at the same time prior to step (d). In another further embodiment, step (b) and step

(d) can be carried out at the same time prior to step (c). In another further embodiment, step (b) and step (c) can be carried out at the same time subsequent to step (d).

Three embodiments of soymilk making process are exemplified in Figure 2.

Dialysis process is carried after grinding and before serving. Beans are washed, optionally subjected to other treatments; then water is added into beans and the beans inside the water are grinded to produce the raw soymilk processing stream. Then, raw soymilk is subjected to dialysis step, cooking step and filtering step. In Embodiment 1 in Figure 2, dialysis step and cooking step can be carried out at the same time prior to filtering step to obtain milk ready for serve. Meanwhile, heating can accelerate dialysis efficiency. In Embodiment 2 in Figure 2, dialysis step and filtering step can be carried out at the same time prior to cooking step. In Embodiment 3 in Figure 2, dialysis step and filtering step can be carried out at the same time subsequent to cooking step.

In one embodiment, during the method of making soymilk as mentioned above, the pore size of a dialysis membrane used in the dialysis is 8,000 Da-14,000 Da. In a further embodiment, the pore size of a dialysis membrane is selected from the range of 8,000 Da- 13,000 Da, 8,000 Da-12,000 Da, 8,000 Da-11,000 Da, 8,000 Da-10,000 Da, 8,000 Da-9,000 Da, 9,000 Da-14,000 Da, 10,000 Da-14,000 Da, 11,000 Da-14,000 Da, 12,000 Da-14,000 Da and 13,000 Da-14,000 Da. In one preferred embodiment, the pore size of a dialysis membrane used in the dialysis is smaller than heat stable material such as cellulous.

In one aspect, the present disclosure relates to an equipment for separating Bowman-Birk inhibitor (BBI) from a soymilk processing stream comprising a water tank and a dialysis device.

In some embodiments, the dialysis device is selected from dialysis bag, dialysis tube and other membrane based separation devices.

In one embodiment, the equipment further comprises a device for one or more chromatographic separation techniques. Preferably, the chromatographic separation techniques are selected from ion exchange chromatography, affinity chromatography and hydrophobic interaction chromatography.

In one embodiment, the pore size of a dialysis membrane used in the dialysis device is 8,000 Da-14,000 Da. In a further embodiment, the pore size of a dialysis membrane is selected from the range of 8,000 Da-13,000 Da, 8,000 Da-12,000 Da, 8,000 Da-11,000 Da, 8,000 Da-10,000 Da, 8,000 Da-9,000 Da, 9,000 Da-14,000 Da, 10,000 Da-14,000 Da, 11,000 Da-14,000 Da, 12,000 Da-14,000 Da and 13,000 Da-14,000 Da. In one preferred

embodiment, the pore size of a dialysis membrane used in the dialysis is smaller than heat stable material such as cellulous.

One embodiment of the equipment for separating BBI from a soymilk processing stream is exemplified in Fig. 3, wherein the accelerator for dialysis is a stirring system, in particular, a stir bar.

In one aspect, the present disclosure provides a food processor comprising the equipment for separating Bowman-Birk inhibitor (BBI) from a soymilk processing stream as mentioned above.

In one aspect, this disclosure provides a food processor comprising:

a machine head connected with a grinding blade;

a grinding container;

a solution switching control;

a cooking container comprising the equipment for separating Bowman-Birk inhibitor (BBI) from a soymilk processing stream as mentioned above; a heat plate;

an electric connector;

In one embodiment, a food processor, in particular, a soymilk maker exemplified in Fig. 4 includes:

Machine head 1 connected with grinding blade;

Grinding Container 2;

Solution switching control 3;

Cooking container with dialysis function 4;

Heat plate 5;

Electric connector 6;

Solution switching control 3 and electric connector 6 are fixed; machine head 1 can plug in 6; 2 and 4 are detachable;

Parts 1, 3, 5 and 6 are connected, to enable the machine function as designed;

Cooked Soymilk is in dialysis tube in part 4 and then transfer to the cup.

In another embodiment, as shown in Fig. 5, the grinding container 4' is configured to be inside the cooking container 4. Machine head 1 connected with grinding blade, Heat plate 5 and Electric connector 6 are connected.

In one aspect, the present disclosure provides a method for separating

chromatographic separation techniques which are performed prior to and/or after the dialysis separation. In one embodiment, the pore size of a dialysis membrane used in the dialysis separation is 8,000 Da-14,000 Da. In a further embodiment, the pore size of a dialysis membrane is selected from the range of 8,000 Da-13,000 Da, 8,000 Da-12,000 Da, 8,000 Da- 11,000 Da, 8,000 Da-10,000 Da, 8,000 Da-9,000 Da, 9,000 Da-14,000 Da, 10,000 Da-14,000 Da, 1 1,000 Da-14,000 Da, 12,000 Da-14,000 Da and 13,000 Da-14,000 Da. In one preferred embodiment, the pore size of a dialysis membrane used in the dialysis is smaller than heat stable material such as cellulous.

In some embodiments, one or more chromatographic separation techniques used in the method for separating Bowman-Birk inhibitor (BBI) from a soy processing stream are selected from ion exchange chromatography, affinity chromatography and hydrophobic interaction chromatography.

Other features and uses of the invention and their associated advantages will be evident to a person skilled in the art upon reading the description and the examples.

It is to be understood that this invention is not limited to the particular embodiments shown here. The following examples are provided for illustrative purposes and are not intended to limit the scope of the invention since the scope of the present invention is limited only by the appended claims and equivalents thereof.

EXAMPLES

Example 1 : The preparation of soymilk by dialysis with pore size of 8,000 Da

Raw soymilk was prepared by grinding 75 g beans and 1 lOOmL water for 20s and repeat 11 times with 5s interval. 50g of raw soymilk was put in refrigerator as a reference. Another 50g raw soymilk was sealed in a dialysis bag with pore size range of 8,000 Da - 14,000 Da. The dialysis bag with soymilk was put in a tank with 1L water, and left in refrigerator. Those two samples were store at 4°C overnight.

Example 2: TI activity and chymotrypsin inhibitor activity test of soymilk

TI activity (TIA) and chymotrypsin inhibitor activity (CIA) were tested by spectrometric method. For TIA measurement, the spectrometric method is based on catalytic reaction of a synthetic substrate BAPA (benzoyl-DL-arginine-p-nitroanalide hydrochloride). BAP A can be hydro lysed into p-nitroaniline and Na-Benzoly-L-Arginine with the presence of trypsin. The hydro lyzed product, p-nitroaniline, can be detected by UV-Vis spectrometer as it has the absorbance at 410nm. If TI has its activity and exists in the system, it will bind the trypsin, and few BAPA will be hydrolyzed. If no TI exists in the testing system, BAPA will be hydrolyzed. So more trypsin inhibitors in sample will result in less absorbance at 410nm. The method of CIA measurement is similar as TIA measurement, but using substrate, N-benzoyl-L-tyrosine p-nitroanilide (BTpNA) and chymotrypsin solution instead of BAP A and trypsin.

Figure 6 and Figure 7 show that TI activity and chymotrypsin inhibitor activity of soymilk with different treatments, respectively. TI activity and chymotrypsin inhibitor activity in soymilk with dialysis treatment is 21.7% and 30.8% respectively lower than that in untreated soymilk. Since BBI possesses both TIA and CIA in soymilk and KTI is the large molecule which can't flow out by dialysis, the reduction of TIA as shown in Examples 2-5 is mainly due to BBI reduction. These results indicates that dialysis in soymilk making can separate undesired protein such as BBI. In addition, dissolved protein content was detected by Kjeldahl method. As shown in Figure 8, dissolved protein content didn't change too much after dialysis treatment which indicates the minim influence of nutrition loss caused by this method.

One skilled in the art would readily appreciate that the methods, devices and equipments described herein are representative of exemplary embodiments, and not intended as limitations on the scope of the invention. It will be readily apparent to one skilled in the art that varying substitutions and modifications may be made to the present disclosure disclosed herein without departing from the scope and spirit of the invention.

All publications mentioned in the specification are indicative of the levels of those skilled in the art to which the present disclosure pertains. All publications are herein incorporated by reference to the same extent as if each individual publication was specifically and individually indicated as incorporated by reference.

Claims

CLAIMS:

1. A method of making soymilk comprising the steps of:

(a) preparing a raw soymilk processing stream,

(b) subjecting the soymilk processing stream to a dialysis separation so as to separate Bowman-Birk inhibitor (BBI) from the soymilk processing stream, and

(c) cooking the soymilk processing stream;

2. The method of claim 1, wherein the step (b) further comprises subjecting the soymilk processing stream to one or more chromatographic separation techniques which are performed prior to and/or subsequent to the dialysis separation.

3. The method of claim 2, wherein said chromatographic separation techniques are selected from ion exchange chromatography, affinity chromatography and hydrophobic interaction chromatography.

4. The method of any one of claims 1-3, wherein the pore size of a dialysis membrane used in the dialysis is 8,000 Da-14,000 Da.

5. The method of claim 1, wherein the step (b) is carried out via an equipment comprising a water tank and a dialysis device.

6. The method of claim 5, wherein the equipment further comprises a device for one or more chromatographic separation techniques.

7. The method of claim 6, wherein said chromatographic separation techniques are selected from ion exchange chromatography, affinity chromatography and hydrophobic interaction chromatography.

8. The method of any one of claims 5-7, wherein the pore size of a dialysis membrane used in the dialysis device is 8,000 Da-14,000 Da.

9. The method of any one of claim 5-7, wherein the dialysis device further comprises an accelerator for dialysis.

10. The method of claim 9, wherein the accelerator is selected from stirring system, ultrasonic system, heating system, water refreshing system and vibration system.

11. food processor comprising

a machine head connected with a grinding blade;

a grinding container;

a solution switching control;

a cooking container comprising the equipment of claims 5-10;

a heat plate;

an electric connector;

wherein the solution switching control and the electric connector are fixed; the machine head can plug in the electric connector; the grinding container and the cooking container comprising the equipment of claims 5-10 are detachable; the machine head, the solution switching control, the heat plate and the electric connector are connected, and wherein a food processing stream is cooked in the equipment of claims 5-10 inside the cooking container and then transferred to a cup.

12. The food processor of claim 11, wherein the grinding container is configured to be inside the cooking container.

13. The food processor of any one of claims 11-12, wherein the food processor is selected from a soymilk maker, a tofu making machine and a soy food processor.

14. A method for separating Bowman-Birk inhibitor (BBI) from a soy processing stream comprising the step of subjecting the soy processing stream to a dialysis separation, and optionally, one or more chromatographic separation techniques which are performed prior to and/or subsequent to the dialysis separation, wherein the pore size of a dialysis membrane used in the dialysis separation is 8,000 Da-14,000 Da.