**10. Results and discussion**

### **10.1. Characterisation of fibers**

The dietary fibers used in this study have different water solubility characteristics: inulin is a soluble fiber, bamboo and wheat are insoluble fibers, apple is partially insoluble fiber, and psyllium forms a viscous dispersion at concentrations below 1% and a clear gelatinous mass at 2%. Chitosan is a fiber of a different origin, i.e. from animal source and is soluble in an acidic medium and flocculates in an alkaline medium. We used these fibers because they present different physicochemical behaviors that have been described in literature [144,4]. The commercial fiber compositions used in this study, regarding total, soluble and insoluble fractions, are shown in Table 4. Analysis for dietary fiber using the AOAC method 991.43 showed that wheat and bamboo have high amounts of insoluble fraction.


nd: no detectable

470 The Complex World of Polysaccharides

standard curve was developed [138].

**9.6. Statistical analysis** 

**10. Results and discussion** 

**10.1. Characterisation of fibers** 

digestive simulation solution (10 mL) was mixed with 1.0 mL of reagent, tubes were incubated for 5 min in water bath at 37°C and developed colour were read in spectrophotometer (Spectronic 20 Genesys TM, Spectronic Instrument, USA) at 505 nm. Final reaction colour is stable for 30 min. Glucose calibration curve was carried out. The amounts of glucose used in

To determine calcium concentration a spectrophotometric method was used. Calcium reacts with 3.7 mmol/L cresolphtalein complexone (Cpx) at pH 11 (buffer 0.2 mol/L aminomethylpropanol (AMP) solution in 35%v/v methanol) (Wiener Lab Ca-color Kit, Argentina). Assays were carried directly in spectrophotometer test tubes: 50 lL Cpx were mixed with a plastic rod and absorbance was read in spectrophotometer (Spectronic 20 Genesys TM, Spectronic Instrument, USA) at 570 nm (internal blank), then 20 mL of each digestive mimicking sample were added, immediately mixed and read after 10 min. A

To determine iron concentration in the dialysates a spectrophotometric method was used, 500 μL of dialyzates was reduced with 2 mL of mercaptoacetic acid (succinic acid buffer, pH 3.7). Then, iron reacted with one drop of pyridyl bis-phenil triazine sulfonate (PBTS) producing a pink color due to the complex formed (Wiener Lab Fe-colour Kit, Rosario, Argentina). Absorbance was read on a spectrophotometer (Spectronic 20 Genesys Thermo Electron Scientific Instruments Corp., Madison, WI, USA) at 560 nm (internal blank). All glassware used in sample preparation and analysis was rinsed with 10% (v/v) concentrated HCl (37%) and deionised water before using, to avoid mineral contamination. A regression equation (y = 2.5333x + 0.0042, R2 = 0.995) derived from data generated from standards of FeSO4 was used to calculate iron concentrations in the samples. Iron retention percentages for each studied fibers were calculated as a percentage of the amount of iron measured in

Experiments were performed at least five times for each dietary fiber using freshly prepared yogurt. For total iron concentration in dialyzates, each individual sample was run in duplicate. Averages and standard deviations were calculated and expressed in each case as the mean ± SD for n replicates. Normality of the data was checked with the Lilliefors test. The influence of different dietary fibers on the retention percentages of glucose, calcium and iron were statistically analyzed by a one-way analysis of variance (ANOVA) (p < 0.05) to

The dietary fibers used in this study have different water solubility characteristics: inulin is a soluble fiber, bamboo and wheat are insoluble fibers, apple is partially insoluble fiber, and psyllium forms a viscous dispersion at concentrations below 1% and a clear gelatinous mass

the dialysed medium obtained with the control yoghurt without fibers [139].

find significant differences and Tukey's test to compare means.

this study correspond to available carbohydrates in the human mixed diet.

**Table 4.** Total, soluble and insoluble fiber content (g/100g) of employed fibers

Inulin presents only soluble fraction as expected. Psyllium and apple have both soluble and insoluble fractions. Apple fiber is characterized by a well balanced proportion between soluble and insoluble fraction [145]. The total dietary fiber content is 45.2% for psyllium, which is an acceptable value, taking into account that the supplier declared a 49.15% content for Plantago ovata seed husk in Metamucil preparation. Van Craeyveld et al. [146] reported 3.4% (dm) ash and 7.1% (dm) protein contents for Plantago ovata seed husks. The total dietary fiber content is 58.1% for apple, which is about 10–14% higher than the values reported by Sudha et al. [147]; however, this value was in accordance with suppliers. The chitosan used in this study has 98% of insoluble fraction and no detectable soluble fraction. Furthermore the characteristics of this biopolymer are a deacetylation degree of 89%, a viscosity of 120 mPa.s, 6.7 g% moisture and 0.67 g% ash content.

Plant fiber characterisations were completed with the study of Acid Detergent Fiber (ADF) and Neutral Detergent Fiber (NDF), lignin, cellulose and hemicellulose contents (Table 5). Apple presents the highest lignin content. Wheat fiber mainly has cellulose. Bamboo has proportional amounts of cellulose and hemicellulose, but compared with other fibers, has the highest hemicellulose content. These results are in accordance with their plant fiber origins and previous works [145-149]. Frutafit-Inulin was not analysed because its composition was ≥85.5% (w/w) of inulin, ≤9.5% of mono and disaccharides, ≤0.1% of ash with polymerisation degree ≥9 according to suppliers. Chitosan was not analysed either, because of its animal origin.


**Table 5.** Acid Detergent Fiber (ADF), Neutral Detergent Fiber (NDF), lignin, cellulose and hemicellulose (%) of fibers

Scientists who deal with animal nutrition usually use Van Soest's method to analyse feed. Scientists working on human nutrition use methods of the AOAC, because of their interest in soluble fiber. It is known that soluble fiber plays an important role in human health and the food industry. However, it could be useful in human nutrition to know the composition of insoluble fiber, as it is possible that insoluble fibers do not all have the same effect on human health. The NDF and insoluble fiber methods were applied to the same samples. Insoluble fiber includes hemicellulose, cellulose, lignin, cutin, suberin, chitin, chitosan, waxes and resistant starch. NDF includes hemicellulose, cellulose and lignin. Escarnot et al. [149] studied three wheat varieties and four spelt genotypes. They analysed three milling fractions from those grains for insoluble and soluble fiber contents, lignin, hemicellulose and cellulose. They found a very high correlation (r2 = 0.99) between the two methods, showing that NDF and insoluble fiber methods cover the same types of fiber. For insoluble fiber analysis, the NDF method is faster and more thorough.
