Effect of High Pressure on the Digestibility of Unabsorbed Proteins Surrounding Soybean Oleosomes

Particle Size

Crude oleosomes

Purified oleosomes

Crude-HPP oleosomes

Purified-HPP oleosomes

Size Exclusion Chromatography

SDS Page

Confocal Microscopy

In vitro Digestion

bD. Maurer, aE. McAllister c,eM. Lamballerie, d,eE. David-Briand, d,eM. Anton and aS. Jung

aDepartment of Food Science and Human Nutrition, Iowa State University, Ames, IA 50011, bDepartment of Agricultural and Biosystems Engineering , Iowa State University, cONIRIS, Nantes, France, dINRA-BIA, Nantes, France, eSFR 4204, IBSM Joint Federation, Nantes, France

Results Soybean oleosomes (oil bodies) are a natural

occurring emulsion with high physical stability that are extracted from full-fat soy flour.

High pressure processing (HPP), because of its pasteurization effect, can potentially increase shelf-life of soybean oleosomes, while also modifying protein structure and therefore their digestibility and physical stability.

Level of purification during extraction impacts surrounding environment, mostly presence of unabsorbed proteins.

The objective of the study was to determine the effect of HPP on in vitro digestibility and characteristics of soybean oleosomes depending on the level of unabsorbed proteins around the oleosomes (crude vs purified).

Potential outcome(s): soybean oleosomes, despite their unique properties, are not used by the food industry. By having a better understanding on how the level of purification might impact these emulsions, the food industry might find appropriate applications. HPP could modify these oleosomes and give them properties that can be of interest.

8th International Conference on High Pressure Bioscience and BiotechnologyJuly 15th-18th 2014, Nantes, France

Figure 3: SEC-HPLC throughout a static in vitro digestion.

*HPP starting fluid temperature was 10°C, dwell time was 5 min. G: Gastric in vitro. I: Intestinal in vitro.

A: Crude oleosomes

Sa

mp

le (

%)

B: Crude oleosomes treated at 600 MPa*

G

G

G

G

GG

GG

G

G

G

G

II

I

I

I

I

II

I

I

I

I

Figure 2: : Effect of purification level and treatment on mean diameter throughout in vitro digestion.

Gastric End/Intestinal Start

Crude Oleosomes

Purified Oleosomes

By-products:Insoluble and

Protein-rich fractions

Stir at 57 °C for 15 hours

Grind

Full Fat Soybean

Flour

Centrifuge

Separate

3x

HPP HPP-Crude Oleosomes

Deionized Water Wash

Centrifuge

3x

HPP HPP-Purified Oleosomes

A B

Oil body matrix Cytosol

C

A: Crude oleosome = oleosome with secondary layer surrounded by water-soluble compounds. B: Purified oleosome with secondary layer. C: Purified oleosome. : Phospholipids. : Oleosins. : Soluble proteins. : Soluble carbohydrates. : Triacylglycerides.

Oil body matrix

Cytosol

Increasing degree of purification

Oil body matrix

Flow Chart of Oleosomes Extraction and Processing

Schematic Representation of Oleosome in Function of Purification Stage

Figure 1: Effect of purification level and treatment on particle size distribution and mean diameter.

Oleosomes Gastrica

1h, pH 2.5, 37oCIntestinalb

20h, pH 6.5, 37oCDigested Solution

120090030012006030 0

IntestinalGastricStd.(kDa)

Crude oleosomes

1200900

30012006045300

IntestinalGastricStd. (kDa)

Crude-HPP oleosomes

1320

 480300 120 60 30 0 60 45 30 15 0

IntestinalGastricPureCrude Std. (kDa)

Purified oleosomes

1440132048030012060 30 0604530150

IntestinalGastricHPP pure

PureStd. (kDa)

Purified-HPP oleosomes

Summary HPP destabilized crude oleosomes in gastric phase

illustrated by an increase in particle size.

Although HPP did not change the peptide profile of the crude oleosomes, the treatment significantly modified the rate of digestion of the major soy proteins as illustrated by the SDS-PAGE profile.

Treatment D[3,2]

Crude 8.5±1.5

Crude-HPP 11.0±1.3

Purified 5.3±1.8

Purified-HPP 7.8±2.5Vo

lum

e (

%)

aIn gastric fluid, incubated in the dark, stirred at 400 rpm.

bBile extract and pancreatic juice added, stirred at 400 rpm.

Purification impacts particle size distribution (bimodal to unimodal) and decreased average D[3,2] values.

For crude oleosomes, HPP increased particle size while variability in D[3,2] in purified pressurized oleosomes was high.

Level of purification had higher impact than HPP on particle size during the gastric and intestinal phases.

Purified oleosomes were more stable during the gastric phase compared to crude. During the intestinal phase purified oleosomes had significant particle size reduction.

HPP does not impact the % of particles with a size lower than 17,000 Da during the gastric and intestinal phases.

For particles >17,000 Da, time and HPP seem to influence % of particle size during the gastric and intestinal digestion phases.

Confocal microscopy confirmed removal of larger oil droplets (red) during the purification process (crude vs purified)

Less proteins (green) are also observed in purified sample but still present.

Proteins from crude oleosomes are rather stable during both the gastric and intestinal phases. After HPP, in vitro digestion during intestinal phase is accelerated.

During the gastric phase, HPP seems to inhibit in vitro digestion of the proteins from purified oleosomes.

Introduction

Methods

This project was funded by