2.4.1. Pre-Hydration of By-Products

By-product powders were soaked in distilled water overnight at room temperature. The amount of soaking in water required for this differed depending on the results of the water-holding capacity for each sample (Table 2). According to Chen and Rubenthaler [42], pre-hydration of the by-product reduces the absorption of water necessary to form the dough and reduces the water competition between the dough ingredients and the dry by-product.


**Table 2.** Formulations of wheat and gluten-free bread.

CTRL: control; DOP: dried orange pomace; DAP: dried apple pomace; DTP: dried tomato peel; DPP: dried pepper peel; DPPP: dried prickly pear peel; DPPSP: dried prickly pear seed peel; B-P: by-product; X: water-holding capacity (WHC).

## 2.4.2. Preparation of Bread

Bread with 100 g of wheat flour was prepared as control of wheat bread (Wheat CTRL 1). Gluten-free bread made from a mixture of corn flour and chickpea flour at a ratio of 2/1 (*w*/*w*) and with 0% *w*/*w* addition of by-products represented the gluten-free control bread (GF CTRL 2). Gluten-free breads were prepared and enriched with pomace and peel powders.

Gluten-free bread recipes were prepared by replacing the base flour with 0, 2. 5, 5, or 7.5% (*w*/*w*) by-products (DOP, DAP, DPP, DTP, DPPP, or DPPSP). All by-product powders were used after their pre-hydration. The flour, yeast, by-products, and water were mixed for 1 min at medium speed. Then salt, previously dissolved in water, was gradually added to the mixture. The dough was mixed for an additional 6 min; then, the obtained dough was divided, molded, placed in an aluminum foil pan (9 × 7 × 3 cm), and fermented at 37 ◦C. The fermentation time varied depending on the T1—the time at which dough attains the maximum height (Hm) in rheofermentometer measurements. The baking test was carried out in an aluminum foil pan in an electric oven at 230 ◦C for 30 min. After baking, the bread was cooled at room temperature. The wheat control bread was prepared without any additives, as was the gluten-free control bread. The formulations for each type of bread are presented in Table 2.

#### *2.5. Bread Quality Evaluation*

The analysis of wheat and gluten-free bread was performed after 1 h of baking and included: weight loss, specific volume, moisture content, pH of bread, color, and crumb cell analysis.

*Weight loss (WL)* is the difference between the weight of the dough and the weight of the bread just after baking dividing by the weight of dough [43].

#### 2.5.1. Specific Volume

The loaf volume was calculated by the rapeseed displacement method (10.05) [44], and the specific volume (cm3/g) was calculated by dividing the volume by the weight.

#### 2.5.2. Moisture Content

Moisture content in bread was determined according to the ICC 110/1 method [45].

#### 2.5.3. Measurement of pH Values

The bread was ground into a fine powder; 10 g of this powder was mixed with 100 mL of distilled water which had an initial pH of 5.68 ± 0.16. The mixture was then placed on an orbital shaker (Heidolph polymax 1040, Schwabach, Germany) and stirred for 30 min at room temperature. The pH was measured in the supernatant solution after a resting time of 10 min [46,47].

#### 2.5.4. Color Analysis

The color of the crust and the crumb of the bread were determined according to the method of He and Pei [48] using the Color Grab color-extracting application (version 3.6.1, 2017, Loomatix Ltd., Munchen, Germany). To ensure that the color capture was not affected by ambient light, a closed polystyrene box (39 × 17 × 28 cm) was used, integrated with a 1.2 W 5 V white LED to obtain an evenly scattered light on top of the sample. The *CIE-L\*a\*b\** color space mode was chosen. This is a mathematical color model based on the sensitivity of the human visual spectrum [49], where *<sup>L</sup>\** represents the lightness, *<sup>a</sup>\** represents green − /red+, and *<sup>b</sup>\** represents blue − /yellow+. Color measurements were taken at 5 different locations on the same crust or crumb.

#### 2.5.5. Crumb Cell Analysis

Image processing was performed using the Image J software (1.43u; Java1.7.0-2132 bit, Wayne Rasband, National Institute of Mental Health (NIH), Bethesda, MD, USA). The bread was cut on a horizontally into 2 slices. The slice images were captured and saved in TIFF format. Each image was cropped to keep only the bread crumb and then converted to 8-bit grayscale to obtain black (crumb pores) and white (crumb) thresholds. The measured parameters were as follows: cell number/mm2, average pore size, pore area fraction, perimeter, and solidity.

#### *2.6. Statistical Analysis*

Statistical analysis was performed using the statistical JMP Trial 15(SAS Institute Inc., Cary, NC, USA). The significance of the difference between the mean values was estimated using one-way analysis of variance (ANOVA), and the means were compared using the Tukey's test (*p* ≤ 0.05). Principal component analysis (PCA) and hierarchical cluster analysis (HCA) were carried out to establish the correlation between dough properties and bread quality. The HCA was based on the Ward method using Euclidean distances to show the similarities and differences between the bread samples. The by-product addition percentage parameter was excluded from the HCA. The results of PCA and HCA are presented in graphical form.
