Introduction 1 2 3 4 5 6 7 9 10 Materials and methods Substrates ® 11 12 Batch in vitro digestion model 7 13 2 4 2 4 α 2 2 Dynamic in vitro digestion model (TIM-1) 8 2 2 3 2 2 3  3 ,  Characterization 10 14 15 16 16 17 Wide-angle X-Ray powder diffraction (X-Ray) measurements were performed using a Scintag PADX Diffractometer (PANAlytical, Almelo, NL) operating at 45 mA and 40 kV at an angular range of 2θ from 1 to 40° with a step size of 0.03°. Counting time was 5 s on each step. Differential scanning calorimetry (DSC) measurements were performed using the Thermal Analyst system 2000 (DSC 2910, TA Instument Ltd, Newcastle, UK). Samples at a starch:water ratio of 1:4 were prepared and heated from 4°C to 200°C at 10°C/min, thereafter the samples were cooled at 20°C/min to 4°C and heated again at the same heating rate. All the DSC results were evaluated from the mean of three separate determinations for each sample, unless stated otherwise. Statistical analysis P Results Composition and digestibility of RS containing preparations 1 1 16 Table 1 Average total starch (TS) content of retrograded tapioca maltodextrins (RTmd) and high amylose maize starch (HAM) and their indigestible starch fractions obtained by dynamic digestion, batch digestion and the Megazyme resistant starch (RS) method (mean values ± SD) Sample TS (g/100 g dm) Digestion method Indigestible starch fraction (g/100 g TS dm) RTmd n Dynamic n a,b Batch n a Megazyme n b HAM n Dynamic n d Batch n c Megazyme n d P Composition of digestion residues 2 1 Table 2 Total starch (TS) and resistant starch (RS) contents of in vitro and in vivo digestion residues of retrograded tapioca maltodextrins (RTmd) and high amylose maize starch (HAM) (mean values ± SD) Sample TS (g/100 g dm) RS (g/100 g dm) RTmd dynamic n n RTmd batch n n RTmd in vivo n n HAM dynamic n n HAM batch n n 2 1 α 1 α 1 Fig. 1 a b  Characterization of digestion residues X-ray diffraction 2 Fig. 2 X-ray diffraction patterns of retrograded tapioca maltodextrins (RTmd) and their digestion residues  3 18 19 2 Fig. 3 X-ray diffraction patterns of high amylose maize starch (HAM) and their digestion residues  DSC 3 T p 20 21 3 3 20 Table 3 n T 0 T p T c Sample T 0 T p T c ΔH (J/g) First heating n 60.7 112.8 140.4 a RTmd dynamic 58.4 108.6 132.4 a RTmd batch 61.8 109.1 136.4 a RTmd in vivo 58.4 105.0 123.5 b HAM 61.8 96.9 110.0 c HAM dynamic 55.5 98.7 127.0 c n 59.9 91.5 110.5 c Second heating RTmd dynamic 79.7 87.9 144.0 *2.4 ± 0.4 HAM 60.0 88.2 109.6 d HAM dynamic 84.9 94.2 146.1 d HAM batch 59.8 83.2 101.9 e P Discussion Digestibility of RS preparations In vitro digestion models use a strictly standardized procedure based on an average human digestibility. Therefore standard deviations of in vitro experiments are expected to be low. In vitro obtained HAM fractions showed higher standard deviations compared to in vitro digested RTmd fractions. Possibly the molecular structures of HAM have been more sensitive to small differences in experimental conditions such as mechanical stress, temperature or pH between experiments. 1 10 8 7 16 22 1 2 Structural aspects of the RS preparations X-Ray diffraction 2 12 23 11 23 25 26 2 27 DSC 20 1 28 27 Conclusions The results of this study show that both, a simple and a more sophisticated in vitro digestion model and the ileostomy study led to similar amounts of starch escaping digestion when using a RS3 containing carbohydrate source as substrate. In contrast, when the two in vitro digestion models were compared with a RS2 containing preparation, different amounts of indigestible starch were found. It is not possible to predict which model simulated the in vivo RS2 fraction more precisely. The more sophisticated in vitro digestion model attacked starch probably more strongly compared to the batch in vitro digestion model using an RS2 and an RS3 containing substrate. This was confirmed by differences in crystalline fractions and structural rearrangements as measured by X-ray and DSC measurements. These structural differences, caused by different experimental parameters influenced starch digestibility of HAM but not of RTmd. The behavior of other RS preparations in the two in vitro models described here can not be deduced from the obtained results and has to be examined case by case.