The high levels of non-communicable diseases such as CVD and type 2 diabetes mellitus are linked to obesity and poor diet. This continuing emphasis on health in relation to food is proving a powerful driver for the development of cheap but palatable and more functional foods. However, the efficacy of such foods is often hard to prove in human subjects. Thus, a suite of tools has been developed including in silico and in vitro simulations and animal models. Although animal models offer physiologically relevant platforms for research, their use for experimentation is problematic for consumers. Thus, in vitro methods such as Infogest protocols have been developed to provide digestion endpoints or even an indication of the kinetics of digestion. These protocols have been validated for a range of food systems but they still miss the final absorption step. This review discusses the use of such in vitro models and what further steps need to be included to make the bioaccessibility determination more relevant to bioavailability and human health.

The aim of this study was to test the effective separation of shape indices of otoliths of three species belonging to the family Sciaenidae before and after in vitro digestion. We measured 328 sagittal otoliths and applied six shape indices. Before the experiment, the aspect ratio (otolith height/otolith length%), circularity, ellipticity and relative surface of the sulcus acusticus were suitable for differentiating the species of genera Paralonchurus and Stellifer. Among the species of Stellifer, the aspect ratio and rectangularity were suitable. Otoliths exposed to in vitro digestion showed no significant differences in their morphometry before and after the experiment. After in vitro digestion, the aspect ratio and circularity were effective in separating Paralonchurus and Stellifer. However, none of the indices used in the present study were efficient to separate otoliths of congeneric species after in vitro digestion.

We report the fatty acid profile of raw milk and of the corresponding digested milk from different sources (human milk, formula milk and donkey, bovine, ovine and caprine milk) to gain information on the nutritional quality of different milk sources in infant nutrition.

Short chain fatty acids (SC-FA) were higher in bovine and caprine milk, intermediate in ovine and donkey and lower in human and formula milk. Medium chain fatty acids (MC-FA) showed the highest values for bovine and caprine milk and the lowest for donkey and formula milk, whereas long chain fatty acids (LC-FA) were the highest in donkey and formula milk and intermediate in human milk.

The percentage distribution of fatty acids liberated after in vitro digestion did not reflect the patterns found in the corresponding milk sources. In particular, MC free fatty acids (MC-FFA) showed the highest and the lowest values in donkey and in formula milk, LC-FFA showed the highest value in human milk. The total FFA was highest in human milk, lowest in formula milk and intermediate in donkey, bovine, ovine, and caprine milk.

Se bioavailability in commercial pet foods has been shown to be highly variable. The aim of the present study was to identify dietary factors associated with in vitro accessibility of Se (Se Aiv) in pet foods. Se Aiv is defined as the percentage of Se from the diet that is potentially available for absorption after in vitro digestion. Sixty-two diets (dog, n 52; cat, n 10) were in vitro enzymatically digested: fifty-four of them were commercially available (kibble, n 20; pellet, n 8; canned, n 17; raw meat, n 6; steamed meat, n 3) and eight were unprocessed (kibble, n 4; canned, n 4) from the same batch as the corresponding processed diets. The present investigation examined if Se Aiv was affected by diet type, dietary protein, methionine, cysteine, lysine and Se content, DM, organic matter and crude protein (CP) digestibility. Se Aiv differed significantly among diet types (P< 0·001). Canned and steamed meat diets had a lower Se Aiv than pelleted and raw meat diets. Se Aiv correlated positively with CP digestibility in extruded diets (kibbles, n 19; r 0·540, P =0·017) and negatively in canned diets (n 16; r − 0·611, P =0·012). Moreover, the canning process (n 4) decreased Se Aiv (P =0·001), whereas extrusion (n 4) revealed no effect on Se Aiv (P =0·297). These differences in Se Aiv between diet types warrant quantification of diet type effects on in vivo Se bioavailability.

Active films were developed for food coating applications. Entrapped phenol susceptibility to digestion was studied. Sodium caseinate (Na-CN) coatings were formulated with 0, 10, 20% Casein (CN) incorporating selected phenols as model antioxidants. This study investigated phenol/CN/Na-CN interactions, in vitro bioaccessibility of phenols and CN role in phenols retention during in vitro gastric and pancreatic digestion. The antioxidant activity of catechin (CAT), rutin (RUT), chlorogenic acid (CHL), gallic acid (GAL), and tannic acid (TA) in coatings varied with the phenolic compound type and CN concentration and was related to phenol hydrophobic binding to CN. ABTS method gave activities ranged from 412 down to 213, and DPPH method gave values from 291·7 to 190·9. An inverse relationship was found with CN content due to CN/phenol interaction. During digestion, a part of phenols was degraded by alkaline pH of pancreatic fluid. Simultaneously, CN proteolysis led to release of phenols and the bioaccessibility index remained above 80% for all phenols. The results suggested the possibility of protecting phenols against oxidation and digestive alteration by entrapment in CN and Na-CN coating films. These positive results showed the ability to produce antioxidant-enriched edible coatings to increase food protection and phenol nutritional intake.

Potatoes are usually a high-glycaemic index (GI) food. Finding a low-GI potato and developing a screening method for finding low-GI cultivars are both health and agricultural priorities. The aims of the present study were to screen the commonly used and newly introduced cultivars of potatoes, in a bid to discover a low-GI potato, and to describe the relationship between in vitro starch digestibility of cooked potatoes and their in vivo glycaemic response. According to International Standard Organisation (ISO) guidelines, seven different potato cultivars were tested for their GI. In vitro enzymatic starch hydrolysis and chemical analyses, including amylose content analysis, were carried out for each potato cultivar, and correlations with the respective GI values were sought. The potato cultivars had a wide range of GI values (53–103). The Carisma cultivar was classified as low GI and the Nicola cultivar (GI = 69) as medium GI and the other five cultivars were classified as high GI according to ISO guidelines. The GI values were strongly and positively correlated with the percentage of in vitro enzymatic hydrolysis of starch in the cooked potatoes, particularly with the hydrolysis percentage at 120 min (r 0·91 and P <0·01). Amylose, dietary fibre and total starch content was not correlated with either in vitro starch digestibility or GI. The findings suggest that low-GI potato cultivars can be identified by screening using a high-throughput in vitro digestion procedure, while chemical composition, including amylose and fibre content, is not indicative.

Practical values to guide food choices for control of postprandial glycaemia need to refer to entire foods in amounts customarily consumed. We tested an in vitro method for determining the relative glycaemic impact (RGI) of customarily consumed portions of foods. Sugars released during in vitro pancreatic digestion of eighty-three foods were measured as glucose equivalents (GE) per gram of food, adjusted by the glycaemic indexes of the sugars to obtain glycaemic GE (GGE) per gram and multiplied by food portion weight to obtain the GGE contribution of the food portion, its RGI. The results were compared with clinical GGE values from subjects who consumed the same food amounts. In vitro and in vivo GGE values were significantly correlated, but the slope of the regression equation was significantly less than one, meaning in vitro GGE values overestimated in vivo GGE values. Bland–Altman method comparison showed the in vitro–in vivo disparity to increase as mean GGE increased, suggesting the need to allow for different rates of homeostatic blood glucose disposal (GD) due to different GGE doses in the customarily consumed food portions. After GD correction, Bland–Altman method comparison showed that the bias in predicting in vivo GGE values from in vitro GGE values was almost completely removed (y = 0·071x − 0·89; R2 0·01). We conclude that in vitro food values for use in managing the glycaemic impact of customarily consumed food quantities require correction for blood GD that is dependent on the GGE content of the food portions involved.

Glycaemic responses to foods reflect the balance between glucose loading into, and its clearance from, the blood. Current in vitro methods for glycaemic analysis do not take into account the key role of glucose disposal. The present study aimed to develop a food intake-sensitive method for measuring the glycaemic impact of food quantities usually consumed, as the difference between release of glucose equivalents (GGE) from food during in vitro digestion and a corresponding estimate of clearance of them from the blood. Five foods – white bread, fruit bread, muesli bar, mashed potato and chickpeas – were consumed on three occasions by twenty volunteers to provide blood glucose response (BGR) curves. GGE release during in vitro digestion of the foods was also plotted. Glucose disposal rates estimated from downward slopes of the BGR curves allowed GGE dose-dependent cumulative glucose disposal to be calculated. By subtracting cumulative glucose disposal from cumulative in vitro GGE release, accuracy in predicting the in vivo glycaemic effect from in vitro GGE values was greatly improved. GGEin vivo = 0·99GGEin vitro+0·75 (R2 0·88). Furthermore, the difference between the curves of cumulative GGE release and disposal closely mimicked in vivo incremental BGR curves. We conclude that valid measurement of the glycaemic impact of foods may be obtained in vitro, and expressed as grams of glucose equivalents per food quantity, by taking account not only of GGE release from food during in vitro digestion, but also of blood glucose clearance in response to the food quantity.

Lycopene β-cyclase (tlcy-b) tomatoes, obtained by modulating carotenogenesis via genetic engineering, contain a large amount of β-carotene, as clearly visible by their intense orange colour. In the present study we have subjected tlcy-b tomatoes to an in vitro simulated digestion and analysed the effects of digestate on cell proliferation. To this aim we used HT-29 human colon adenocarcinoma cells, grown in monolayers, as a model. Digested tomatoes were diluted (20 ml, 50 ml and 100 ml/l) in culture medium and added to the cells for different incubation times (24 h, 48 h and 72 h). Inhibition of cell growth by tomato digestate was dose-dependent and resulted from an arrest of cell cycle progression at the G0/G1 and G2/M phase and by apoptosis induction. A down-regulation of cyclin D1, Bcl-2 and Bcl-xl expression was observed. We also found that heat treatment of samples before digestion enhanced β-carotene release and therefore cell growth inhibition. To induce with purified β-carotene solubilised in tetrahydrofuran the same cell growth inhibition obtained with the tomato digestate, a higher amount of the carotenoid was necessary, suggesting that β-carotene micellarised during digestion is utilised more efficiently by the cells, but also that other tomato molecules, reasonably made available during digestion, may be present and cooperate with β-carotene in promoting cell growth arrest.

The food industry is playing an increasing role in the development and marketing of new products although little is known regarding the bioavailability of the phytochemicals they contain. Our aim was to assess the effect of the presence of absorption modifiers (milk and iron) on the in vitro bioaccessibility and the serum response in vivo of carotenoids and α-tocopherol from fruit juices. Thirty-two young women participated in a three-period (21 d each) supplementation study with a 2-week wash-out in between. Subjects consumed consecutively 2 × 250 ml/d vitamin C-fortified juices supplied as fruit juice, fruit juice containing milk and fruit juice containing milk and iron. Fasting blood samples were collected before and after each supplementation period. In vitro bioaccessibility of carotenoids and α-tocopherol was assessed by a static digestion model. Vitamin E and carotenoids from both studies were determined by HPLC. In vitro, xanthophyll ester hydrolysis and transference of free xanthophylls and α-tocopherol into the micellar phase were higher in the presence of absorption modifiers. In vivo, consumption of the fruit juices provoked significant increments (within-subject) of α-tocopherol and some carotenoids in serum. Dose-adjusted increments in serum of some carotenoids were higher when subjects consumed juices with milk and milk plus iron, although differences did not reach statistical significance. In conclusion, the presence of milk and milk plus iron do not influence the bioavailability of carotenoids and α-tocopherol from fruit juices in vivo. Our results support the use of in vitro models to assess food-related factors affecting bioavailability of carotenoids and tocopherols from foods.

In the present study, we utilised an in vitro digestion procedure to deliver molecules contained in tomatoes to cultured cells and to analyse potential mechanisms underlying the antitumoural effects of tomatoes reported in the literature. Ripe tomatoes underwent in vitro simulated digestion and the aqueous fraction obtained was delivered to HT-29 and HCT-116 colon adenocarcinoma cells. The amount of lycopene released during digestion and transferred to the aqueous fraction during digestion was 10-fold lower than that present in tomato homogenate before digestion. The carotenoid was accumulated by colon adenocarcinoma cells in a dose-dependent manner after the addition of tomato digestate (20–100 ml/l) for 24 h. Tomato digestate inhibited the growth of HT-29 and HCT-116 cells in a dose-dependent manner. Growth inhibition resulted from an arrest of cell cycle progression at the G0/G1 phase and by apoptosis induction. A down regulation of cyclin D1, Bcl-2 and Bcl-xL expression was also observed, without apparent changes in p53, p21, p27 and Bax. In conclusion, the present data demonstrate that the in vitro digestion procedure represents a useful approach to supply tomato to colon cultured cells. Moreover, we have shown that tomato digestate is able to inhibit the growth of colon cancer cells by modulating the expression of regulators of the cell cycle and apoptosis.

The effect of rate of substitution of processed, urea-treated whole-crop wheat (pWCW) for grass silage on intake, performance and whole-tract digestibility was evaluated using 44 dairy cows. Cows received 10.5 kg of concentrates per day and one of the following forage mixtures (dry matter (DM) basis): grass silage alone (W-0); 0.75 grass silage, 0.25 pWCW (W-25); 0.5 grass silage, 0.5 pWCW (W-50) or 0.25 grass silage, 0.75 pWCW (W-75). Forage DM intake increased linearly with inclusion rate of pWCW from 9.7 kg DM per day in cows fed W-0 to 14.6 kg DM per day in W-75. By contrast, milk and protein yield (kg/day) were higher (P < 0.05) in cows receiving W-25 compared with W-0, but there was no effect (P>0.05) of treatment on fat yield (kg/day). From week 11 of the experiment onwards, body condition score increased with rate of inclusion of pWCW (P < 0.05). Whole-tract apparent digestibility of organic matter (OM) and fibre (kg/kg), decreased linearly with rate of inclusion of pWCW. Assuming a constant digestibility of starch in the other diet components, the apparent digestibility of starch in pWCW was 0.95 kg/kg and was not affected by rate of inclusion (P>0.05). Four continuous culture vessels were used to determine the effect of rate of inclusion of pWCW on ruminal metabolism in four periods, each of 14 d duration with sampling conducted on days 9 to 14. Vessel ammonia concentration increased linearly (P < 0.05) with rate of inclusion of pWCW whilst mean pH tended (P = 0.06) to decrease. The ratio of acetate to propionate increased from 2.5 in vessels receiving W-0 to 3.2 in those receiving W-75 (P < 0.001). There was no effect (P>0.05) of treatment on digestibility (g/g) of OM, fibre or starch or microbial protein flow (g/day). It is concluded that forage DM intake increased linearly with rate of inclusion of pWCW, but there was no further improvement in milk yield from inclusion rates above 0.25 of the forage DM, with body condition score increasing instead. Increasing the inclusion rate of pWCW resulted in a more ketogenic volatile fatty acid profile but did not affect the efficiency of microbial protein synthesis when determined in vitro.

Seven mature Welsh-cross pony geldings provided the faecal inocula in a cross-over design experiment, consisting of two 14-day periods. In period 1, four ponies (group 1) were offered a low-starch fibre mix (LS), and three (group 2) were offered a conventional high-starch coarse mix (HS). Both groups were offered these mixes in a 50:50 ratio with mature grass hay, to give a total daily dry-matter intake of 17·5 g/kg live weight per day. Diets were then switched in period 2. At the end of each experimental period freshly voided faeces were collected from each animal and assessed for their ability to ferment grass hay (H), fibre mix (FMix) or starch-based coarse mix (SMix) using the gas production (GP) technique of Theodorou et al. (1994). Donor animal diet and donor animal had no effect on any end-point measurements. Lag times recorded for the SMix were significantly (P<0·001) greater in LS-inoculated bottles compared with the HS inocula (1·74 v. 2·25 h, respectively). Lag times for FMix and SMix varied significantly (P<0·001) between ponies (0·82 to 1·78 h in the FMix and 1·64 to 2·51 h in the SMix). The degradation rate of H also differed significantly (P<0·001) between ponies with the time taken to reach 50% of GP (T50) ranging from 12·70 to 17·30 h. Consequently, it would appear that the effect of feeding LS or HS on the in vitro fermentative capacity of equine faecal inocula is minimal; moreover, the GP technique appears to be valuable tool for evaluating such effects.

In this three-phase study we first compared the availability of fourteen Fe forms in a wheat-based ready-to-eat breakfast cereal using an in vitro digestion/human colonic adenocarcinoma (CaCo-2) cell model. We then investigated the effect of milk and/or coffee on those fortified cereals found in phase 1 to show promising increases in Fe availability. The Fe forms assessed in phase 1 were reduced (control), carbonyl, electrolytic, FePO4, FeSO4, FeCO3, Na2FeEDTA, Ferrochel® (Albion Laboratories, Clearfield, UT, USA; ferrous bis-glycinate), encapsulated ferrous fumarate, FeSO4, ferrous lactate and Biofer® (LipoTech, Britwell Salome, Oxfordshire; FeSO4), SQM® (Sea-Questra-Min Iron; Quali Tech, Chaska, MN, USA; polysaccharide-complexed FeSO4) and Sun Active® (Taiyo Kagaku, Yokkaichi, Japan). All these forms increased Fe uptake compared with the unfortified cereal. Relative to the control, the following increases in Fe availability were observed: electrolytic, 52 %; ferrous fumarate, 30–35 %; Sun Active, 78 %; Ferrochel, 125 %; Na2FeEDTA, 291 %. Recent human studies have shown similar data with regard to Ferrochel, FeSO4 and Na2FeEDTA, with the latter being more bioavailable. Our phase-2 studies indicated that the addition of milk to FeSO4-fortified cereal increased Fe availability, but this availability was markedly decreased by the addition of coffee to the digest. Conversely, a loss in availability from Na2FeEDTA was observed with the addition of milk; however, the addition of coffee did not markedly affect Fe availability from this form. In phase-3 studies we observed increased Fe availability upon the addition of milk to cereals containing Ferrochel, FeSO4, Sun Active, a mixture of reduced Fe and Na2FeEDTA or reduced Fe. For these forms we did not assess the behaviour after the addition of coffee. In conclusion, when considering possible fortificants for optimizing Fe bioavailability within a foodstuff, it is of paramount importance to consider the interaction between the fortified foodstuffs and other components of the meal (such as milk and coffee with a breakfast).