*2.1. Effect on Dough Properties*

The effect of the α-, β-, and γ-CDs on flour water absorption was investigated by consistograph analysis, and the results can be seen in Figure 1. The water absorption increased when up to 4% CD was added after which water absorption was approximately constant or decreased again. The water absorption of the control dough (52.7%) increased the most by addition of 4% α-CD (55.4%), while it increased less for 4% γ-CD (54.1%) and 4% β-CD (53.4%). The decrease in water absorption at 8% CD compared to 4% CD was only significant for β-CD. In general, increased water absorption was expected with increasing CD concentration, as the CDs were added to a constant amount of flour, and

the total mass (dry matter) did, therefore, increase. However, the water absorption did not increase proportionally to the amount of CD added, and it stagnated or decreased when going from 4% to 8%, dependent on the type of CD.

**Figure 1.** Water absorption for different types and concentrations of CD determined by the consistograph method. The water absorption is the hydration needed to obtain a dough with a maximum pressure of 2200 mbar. The error bars indicate the standard deviation. Different letters indicate the significant difference between the treatments (*p* < 0.05).

Similar tendencies have been found by other authors. Up to 3.4% increase in water absorption dependent on the CD concentration was observed for wheat doughs supplemented with up to 1.6% β-CD by Kim and Hill [11]. Likewise, Zhou et al. [13] observed up to 6.4% increase in water absorption with increasing CD concentration of up to 3.0% α- or γ-CD for a durum wheat flour dough. Of particular interest, Duedahl-Olesen et al. [15] found that an increase in water absorption by 7.3% and 8.0% for the addition of 3% αand γ-CD, respectively, whereas a much lower increase in water absorption, was recorded when supplying glucose and maltooligosaccharides at the same level (wt%).

The effects of the different concentrations of α-, β-, and γ-CD on the biaxial extensional properties of the dough were tested by alveograph analysis. The results can be seen in Figure 2. The *P* value, which represents the tenacity of the dough, was approximately constant up to 2% CD (no significant difference from the control), after which it increased with increasing CD concentration for all three types of CD. A concentration of 8% α-, β-, and γ-CD caused an increase in *P* of 63%, 51%, and 43%, respectively. The biaxial extensibility of the dough measured by the *L* values decreased with increasing concentration of the three types of CDs. 8% addition of α-, β-, and γ-CD caused a decrease in *L* of 49%, 52%, and 37%, respectively. The deformation energy measured by the parameter *W* seemed to decrease for low concentrations of CDs, after which it increased again for higher CD concentrations. However, the decrease in *W* was only significant for β-CD, while the *W* values for α- and γ-CD supplemented doughs were not significantly different from the control for any concentration. The *W* value for all concentrations of β-CD was significantly below the value of the control. The *Ie* value, which is called the elasticity index, changed differently dependent on whether α- and γ-CD or β-CD were applied. The *Ie* value seemed to increase for a concentration of up to 4% α- and γ-CD. However, only the *Ie* value for 4% α-CD was significantly different from the control. At 8% α- and γ-CD, the *Ie* value decreased significantly. The *Ie* value decreased with increasing concentration of β-CD. The alveograph results reveal that addition of CDs entails a stiffer and less extensible dough, as *P* increases, while *L* decreases. Addition of α- and γ-CD did not change the strength of the dough significantly, as indicated by *W*, while addition of β-CD resulted in a weaker dough. This was further substantiated by the *Ie* values obtained for the β-CD series of doughs, as according to Kitissou [40], *Ie* is related to the gluten network quality of the

dough. However, the addition of 8% α- and γ-CD also resulted in a significant decrease of the *Ie* value.

**Figure 2.** Effect of the different types and concentrations of CD on the dough extensional properties as determined by the alveograph method. The results for the alveograph parameters *P* (**a**), *L* (**b**), *W* (**c**), and *Ie* (**d**) are shown. The error bars indicate the standard deviation. Different letters indicate the significant difference between the treatments (*p* < 0.05).

While a few studies have investigated the effect of CDs on the mixing properties of the dough, the effect on the extensional properties of the dough has only been studied to a limited extent. Zhou et al. [12] investigated the effect of β-CD on the dough using the extensograph. They found that 0.5–1.5% β-CD increased the maximum resistance to deformation compared to the control, while the maximum resistance to deformation decreased for 2.0–3.0% β-CD. The extensibility increased slightly up to 1.0% β-CD, after which it decreased slightly up to 3.0% β-CD. The results from the alveograph method and the extensograph method cannot be directly compared due to differences in the sample and analysis conditions. However, the study by Zhou et al. [12] supports that at least elevated amounts of β-CD resulted in a weaker dough, probably through a weakening of the gluten network.

The addition of CD to wheat dough has multiple effects, as the CDs can affect both water distribution and the other flour constituents. The water in wheat dough interacts with the different constituents of dough, but the water availability is in general limited [41]. CDs contain multiple hydroxyl groups, which are able to form hydrogen bonds with the water. The addition of CDs might, therefore, limit the availability of water and thereby affect the gluten network development, which would be observed as changes in the mixing and extensional properties of the dough, including the HydHA value and the alveograph parameters. However, our results and other studies indicate that the effects of CDs are also caused by their direct influence on other components of the dough matrix and not just a shift in the distribution of water. Duedahl-Olesen et al. [15] found that α- and γ-

CD resulted in higher water absorption during mixing compared to an equal amount of glucose or non-cyclic maltooligosaccharides. If the higher water absorption should only be attributed to the water-binding capacity of the hydrophilic CDs, similar effects should be expected using their non-linear counterparts, as the water binding capacity is considered to be comparable in the dough matrix with its limited water availability. Furthermore, in the alveograph analysis, doughs with similar consistencies according to the HydHA values were analyzed, which was considered to reduce the effect of the varying water absorption on the results. This suggests that the large changes that were observed in the resultant parameters cannot solely be explained by differences in varying water absorption.

One characteristic that distinguishes CDs from smaller carbohydrates and, to some extent starches, is their general ability to form inclusion complexes by exchanging water in the cavity with a hydrophobic molecule or part of a molecule. In this process, complex formation is mainly driven by the release of "enthalpy rich" cavity-bound water and hydrophobic interaction (removal of ordered low entropy, high enthalpy water around the hydrophobic guest) [42]. Both driving forces would be expected to be favorable in an environment with low water activity. The CDs are (relatively rigid) cyclic oligosaccharides, and they are, therefore, capable of forming rather stable inclusion complexes with a range of primarily lipophilic molecules [1,2,5]. The CDs might interact with lipophilic molecules (e.g., lipids) and lipophilic parts of molecules, e.g., lipophilic parts of gluten proteins, but the strength and selectivity will be dependent on the cavity size of the specific CD. In essence, α-CD is most suitable for complex formation with linear aliphatic molecules (such as lipids), β-CD is suitable for complex formation with aromatic molecules, and γ-CD is suitable for larger aromatic molecules [1,2,5]. Although this leads to a considerable degree of selectivity, the complex-forming ability of the CDs is somewhat general, as typically all three CDs will be able to form a complex with a given (preferably lipophilic) molecule, but with different association constants.

The starch in the dough might also be affected by the addition of CDs. It has previously been suggested that β-CD might disrupt the amylose-lipid complex formation as well as it might form amylose-β-CD and amylose-lipid-β-CD complexes [14,43–45]. This might change the crystallinity of the starches and thereby cause an indirect change in the distribution of water, which has been suggested to affect the mixing properties [15]. However, the disruption of amylose-lipid complex formation and formation of complexes with CDs have primarily been observed for starch, which was at least partly gelatinized. In the dough, most of the starch is organized in starch granules, and the accessibility of the starch is, therefore, limited [46]. The effects of interactions between CDs and starch are, therefore, assumed to be smaller for the dough compared to the bread where the starches have been subjected to extensive gelatinization.

The CDs might also interact with the gluten proteins in the dough due to their ability to form weak inclusion complexes with proteins, which might affect the development and the properties of the gluten network. This was identified by Zhou et al. [12], who found that the addition of β-CD to wheat dough changed the secondary structure of the gluten proteins by increasing the proportion of α-helixes and decreasing the proportion of β-sheets. α-, β-, and γ-CD have been shown to be able to influence the behavior of proteins [47–50], but β-CD causes by far the largest effects, which have been explained by a relatively large affinity towards solvent exposed aromatic amino acids [47,51]. The interaction between β-CD and aromatic amino acids reduces the formation of proteinprotein interactions by hydrophobic interaction in aqueous solutions [47,48]. All CDs had a significant influence on the viscoelastic properties of the dough, but especially β-CD was revealed to have large effect on the value of the alveograph parameters *W* and *Ie*, which are among other things dependent on the gluten network quality. These effects might be caused by interaction between the gluten proteins and β-CD, leading to changes in the strength of potential protein-protein interactions by non-covalent interactions, including hydrophobic interactions. The results indicate that the addition of a large amount of β-CD leads to a lower gluten network quality as assessed from the rheological properties. However, Zhou

et al. [12] suggested that the addition of up to 1.5% β-CD positively affected the gluten network, as the maximum dough tensile resistance in extension increased. In addition to the effects of β-CD, the addition of CDs dilutes the protein content in the dough, which might also decrease the strength of the gluten network. This might be a contributing cause to why the addition of 8% of any of the CDs results in a lower than expected increase in the water absorption as well as the low *Ie* values in the alveograph analysis.
