1. RelEASE OF FLAVONOLS IN SIMULATED IN VITRO GASTROINTESTINAL DIGESTION AND ITS RELATIONSHIP WITH VISCOSITY IN ONION AND APPLE PRODUCTS AND COMMERCIAL QUERCETIN SUPPLEMENT
M.D., Alvarez*, B., Herranz, I., Fernández-Jalao, C., Sánchez-Moreno, B., De Ancos
Department of Characterization, Quality and Safety, Institute of Food Science and Nutrition (ICTAN-CSIC),
José Antonio Novais 10, Madrid, Spain.
The aim of this work was to study the release of flavonols in onion and apple products and in a commercial quercetin supplement throughout in vitro gastrointestinal digestion (GID) using a Dynamic Gastrointestinal Digester (DGD) and its relationship with the viscosity changes in the different digestion phases at a physiological shear rate (10 s–1). The physical stability of the digests was also studied throughout the changes in backscattered light monitored in time with a Turbiscan®
Simulated GID procedure
A multi-compartmental DGD with a computer-controlled system, developed at AINIA Technological Center (Valencia, Spain) was used:
Freeze-dried powder Onion (Allium cepa L. 'Recas', Carabaña, Madrid, Spain).
Freeze-dried powder Apple (Malus Domestica, 'Golden Delicious', Aragón, Spain).
Quercetin supplement powder (70.5 mg; Solaray, Nutraceutical Corp. USA).
Experimental analyses
All digests of the different phases (ND, OP, GD and ID) for the three samples (onion, apple and commercial quercetin supplement) were performed in triplicate.
1. Physical stability of digest
Total flavonol content
-Methodology by Bonoli et al. (2004)
-UV-vis-Spectrophotometer (GE Amersham Biosciences Pharmacia, Sweden).;
-quercetin as external standard calibration curve (1 to 50 µg/mL.)
-Total flavonol content = mg quercetin equivalents (QE) per 100 g dried weight (dw) of sample. -
Rheological measurements (Steady shear tests)
-Kinexus Pro Rotational Rheometer (Malvern Instruments Ltd., Worcestershire, UK) ;
-a cone and plate geometry (4° cone angle, 40 mm diameter);
-shear rates (100 to 0.1 s–1) at 37 °C. + a pre-shearing at 100 s–1 during 5 min;
-apparent viscosity at 10 s–1 (ηa,10) values from apparent viscosity vs.
shear rate curves and compared to K (consistency) values from power law fits.
Principle of TURBISCAN measurement
27 g
-Turbiscan MA classic 2000 (Iesmat S.A., Alcobendas, Spain)
-emulsion sample (7 mL);
-a light beam emitted in near infrared (λ=880 nm);
-each minute for a period of 60 min.;
- Backscattering profiles (BS).
37.5 g
Experimental
Digests of GID phases for onion, apple and commercial quercetin supplement:
- ND (non-digested)
- OP (oral phase)
- GD (gastric digest)
- ID (intestinal digest)
Bioaccessibility (%) = (BCdigested/BCnon-digested) x 100
BCdigested = Concentration of bioactive compounds (BC) in the soluble fraction of digested sample and BCnon- digested = Concentration of BC in the non-digested sample.
In vitro Dynamic Gastrointestinal Digester (DGD) and Outline of the in vitro digestion process. a
-The backscattering (BS) profiles of the digests in the different GID phases (ND, OP, GD and ID) for the three samples studied are shown in Figs. 1-4.
-All three samples presented a stable BS pattern in ND phase (Figs.1a-c). In OP and GD phases, the BS profiles of onion (Figs. 2a,3a) and apple (Figs. 2b,3b) show an aggregation process by an increase of the BS in the middle of the sample (the particle size increase), and at the same time, a notable creaming phenomenon (migration of particles) by the presence of cream layer (increase of BS) at the top part of the sample (right part of the graph). The cream layer increases with the time and it was more marked in onion (thicker layer; Figs. 2a,3a) than in apple (Figs. 2b,3b).
-However, in commercial quercetin supplement OP and GD phases (Figs. 2c,3c, respectively) were very stable. In turn, in ID phase (Fig. 4), it was produced a remarkable change in the BS profiles for the three samples related with the release of BC in the intestine.
-Therefore, the mild alkaline conditions of the intestinal phase produced a change in the physical stability of the digests facilitating the release of the bioactive compounds. a a a b b b b c c c c
A progressive decrease in total flavonols (Table 1) was also observed in onion and quercetin supplement from ND to ID phases, meanwhile a significant increase in total flavonols release was observed in GD and ID phases for apple. Therefore, onion flavonols and quercetin in the commercial supplement appear to have been more resistant in the acidic conditions of the stomach than in the alkaline ones of the small intestine. However, in apple total flavonol content was similar in GD and ID phases.
Results and Discussion
Figure 1. Backscattering (BS) profiles of the non-digested (ND) products.
Figure 2. Backscattering (BS) profiles of the digests in oral phase (OP).
Figure 3. Backscattering (BS) profiles of the digests in gastric phase (GD).
Figure 4. Backscattering (BS) profiles of the digests in intestinal phase (ID).
Table 2. Effects of different in vitro GID phases (ND, OP, GD and ID) and food matrix on steady shear rheological properties for different food products
Table 1. Effects of different in vitro GID phases on total flavonols
GID phases
Onion product
Non-digested (ND)
382±19Ab
Oral-phase (OP)
335±5.8Bb
Gastric digest (GD)
338±12Ba
Intestinal digest (ID)
292±3.5Ca
Bioaccessibility (%)
16.6±2.3b
Apple product
27.1±0.71B,Cc
26.2±2.7Cc
32.3±2.1A,Bc
37.3±3.9Ac
59.7±7.4a
Quercetin supplement
840±19Aa
543±2.8Ba
168±4.5Cb
88.0±7.0Db
4.12±0.27c
Onion
K (mPa sn)
ηa,10 (mPa s) ND
237±18Ab
48.0±11Ab OP
111±18Bb
32.8±7.3Bb GD
22.3±2.5Cb
4.77±1.1Cb ID
14.2±0.50Cb
2.00±0.047Cb
Apple
376±13Aa
72.2±17.6Aa
207±15Ba
45.6±14.5Ba
178±21Ba
23.7±8.60 B,Ca
23.8±1.8 Ca
3.41±0.22Ca
Quercetin
12.2±0.18 A,Bc
1.60±0.038Bc
10.5±0.47Cc
1.27±0.087Cc
12.8±0.37Ac
1.78±0.076Ac
11.7±0.30Bc
1.52±0.065Bc
A–Cfor each rheological property and the same matrix, mean values with different letter between different phases are significantly different (P < 0.05).
a-c for each rheological property and the same GID phase, mean values with different letter between different products are significantly different (P < 0.05).
Values are expressed as mg QE/100 g dw.
A–Dfor the same food matrix, mean values with different letter between different phases are significantly different (P < 0.05).
a-cfor the same digestion phase among different products, mean values with different letter are significantly different (P < 0.05).
Flow curves of each GID phase for the three samples showed non-Newtonian shear-thinning behavior (Fig. 5) due to rearrangement in the conformation of the molecules in the dispersion as a result of shearing. As can be seen in Table 2, a decrease in K and ηa,10 values was observed for onion and apple products from ND to ID.
Figure 5. Apparent viscosity changes versus shear rate at the different in vitro GID phases for onion and apple products and commercial quercetin supplement.
The behavior detected in apple is due to the different physico-chemical composition of the food matrix (higher pectin content, total fiber, soluble solids and acidity than onion) which made apple powder more resistant to the intestinal phase. Apple presented the higher bioaccessibility (59.72%) followed by onion (16.55%) and quercetin supplement (4.12%), reflecting the importance of the food matrix effect in the release and bioaccessibility of the flavonols.
However, in commercial quercetin, K and ηa,10 values were similar between different GID phases and much lower than those in onion and apple. Therefore, it seems that a diluting effect is produced in onion and apple products as the samples passed throughout GID. K and ηa,10 values correlated well with the bioaccesibility; apple had the highest K and ηa,10 values in ID, followed by onion and quercetin supplement, in that order.
Concluding Remarks
The release of bioactive compounds in the intestine corresponded with a change in the physical stability of the digest. There is a diluting effect throughout gastrointestinal digestion as it was shown by the K and ηa,10 values of the different GID phases of apple, onion and quercetin supplement. This diluting effect is related to the release of flavonols. Food matrices (apple and onion products) showed higher bioaccessibility than commercial quercetin supplement. Therefore, the evolution of the apparent viscosity and BS profiles throughout in vitro simulated GID can be related with the release of flavonols and their bioaccessibility.