Broccoli leaf powder in gluten-free bread (GFB) -

Broccoli leaf powder in gluten-free bread (GFB)

Broccoli leaves are perceived as waste products, they are characterized by a high content of nutrients and bioactive compounds. The present study evaluated the nutritional value, technological quality, antioxidant properties, and inhibitory activity against the formation of advanced glycation end-products (AGEs) of GF enriched with broccoli leaf powder (BLP). The obtained results indicate that BLP can be successfully used as a component of gluten-free baked products. The newly developed GFB with improved technological and functional properties is an added-value bakery product that could provide health benefits to subjects on a gluten-free diet.

Materials and Methods

Undamaged leaves of mature broccoli (Brassica oleracea L. var. italica) were cleaned of soil residues, washed with water, then blanched shortly (1 min) in hot water to inactivate enzymes and decrease the microbial load. Afterward, petioles and main midribs were removed, and leaf blades were freeze-dried. Dry leaves were ground and sieved to obtain homogenous powder (particle size ≤ 0.60 mm). The obtained BLP was packed in a sealed plastic box and kept in a refrigerator for further analysis.

Preparation of Experimental Gluten-Free Bread

Corn starch, potato starch, sugar, fresh yeast, pectin, rapeseed oil “Kujawski”, salt, and water were the main ingredients of GFC. Previously characterised BLP was incorporated into the GFB by replacing 5% (w/w) of corn starch in the GFC formula. This level of substitution was based on a preliminary study that showed that 5% was the acceptable replacement level that did not affect the sensory properties of bread, whereas the GFB with 7% BLP had too intense cabbage flavor

To prepare GFs, all solid ingredients were mixed for 5 min at minimum speed using a KitchenAid Professional K45SS mixer in the stainless-steel bowl with a flat beater. Yeast, salt, and sugar were dissolved in the water and added to the dry mixture, together with oil. The batter was mixed for 12 min at speed 2. Then, a 240-g sample of the resulting batter was placed in a greased hexagon-shaped bread pan and proof for 40 min at 35 ◦C and 70% humidity. Experimental GFs were baked for 30 min at 220 ◦C in the laboratory oven. Nine loaves were baked from each formula. After baking, all bread loaves were cooled for at least 2 h at room temperature. Then, GFs were packed in clip-on plastic bags and kept in the dark at room temperature for further analysis. Products of two independent batches, fresh (2 h after baking) and/or stored (24 and 72 h after baking), were analysed.

Characteristics of Experimental Gluten-Free Breads

The basic chemical composition was determined in freeze-dried GFs according to the standard method: moisture content was analysed using the drying method (AOAC 925.10), proteins content was determined with the Kjeldahl method and fat content using Soxhlet extraction with hexane (AOAC 923.03); total ash was determined using the gravimetric method by burning in a muffle furnace at 550 ◦C for 10 h (AOAC 923.03). The total carbohydrate content was calculated by subtracting the values of the moisture, protein, fat, and ash content from 100. The energy values (kJ) were calculated by multiplying the amount of macronutrients by the corresponding conversion factors (17 kJ/g for protein, 37 kJ/g for fat, and 17 kJ/g for carbohydrates).

The crust and crumb colour of GFs was evaluated using a HunterLab ColorFlex. Crust colour was determined at the middle point of the top of the loaf crust, while crumb colour was analysed at the middle point of the central 2-cm slice. The measurements were performed through a 3-cm diameter diaphragm containing an optical glass. The colour was expressed in accordance with the CIELab system.

The texture profile (TPA test) of fresh (2 h) and stored (for 24 and 72 h after baking) crumbs of GFs were analysed using a TA.HD Plus Texture Analyser equipped with a 30-kg load cell. The middle bread slices of 25-mm thickness underwent a double compression cycle up to 40% deformation of its original height with a 35-mm flat-end aluminium compression disc (probe P/35).

Evaluation of the Antioxidant Capacity of BLP and GFs

The total phenolic content (TPC) was determined with the use of the Folin–Ciocalteu reagent method.The Trolox Equivalent Antioxidant Capacity (TEAC) by the 2,20 -azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) assay was performed. Trolox Equivalent Antioxidant Capacity by DPPH Assay, The TEAC by 2-diphenyl-picryl-hydrazyl (DPPH) radical scavenging assay. Photochemiluminescence Assay A photochemiluminescence (PCL) assay was performed to measure the antioxidant capacity of BLP. The inhibiting activity against advanced glycation end-products (AGEs) was assessed using two in vitro model systems: bovine serum albumine (BSA)-glucose and BSA-methylglyoxal (MGO).

Results 

Fresh GFC and GFB were similarly soft (13.21 and 13.80 N, respectively); however, fresh GFB was significantly springier and more cohesive than GFC. Besides, the chewiness of the GFB was over 50% higher compared with the GFC. The chewiness informs about the time required to mastication a piece of food before it is swallowed. The incorporation of BLP into the gluten-free formulation prolonged the chewing time for the GFB crumb.

The application of BLP in the gluten-free formulation caused a significant reduction of crumb springiness; thus, the GFB became very crumbly. However, in comparison with fresh GFB, the chewiness of stored crumb did not change meaningfully, contrary to the GFC stored for 72 h. The broccoli bread was harder compared to the control wheat bread both on the day of baking and during storage. However, the deterioration in texture attributes was more pronounced in the oil-free wheat bread.

The GFC was characterised by a relatively low antioxidant activity evaluated using all assays. Contrary, the BLP was found as a good source of TFC, consequently exerting a high antioxidant capacity. Freeze-drying, which was used to prepare BLP, is a well-known method that allows preserving the nutritional value of the starting material, including bioactive compounds. Therefore, as expected, the fortification of GF with BLP significantly increased the antioxidant potential of experimental GFB. Among broccoli parts, leaf tissue had the highest TFC and antioxidant activity (DPPH), compared with florets and stems. ABTS, DPPH, and PCL-ACW assays are associated with the activity of hydrophilic compounds like polyphenols, which have confirmation in TFC. On the other hand, the PCL-ACL assay informs about the activity of         lipophilic compounds, like fat-soluble vitamins and carotenoids. The result showed that broccoli bread contained the β-carotene and lutein that are characterised by a strong antioxidant activity. BLP was characterised by very high PCL-ACL activity, and consequently, this assay was the highest among all analysed in GF, suggesting that BLP can be a good source of lipophilic compounds. In this study TFC and antioxidant capacity increased after in vitro digestion, suggesting that the health-promoting potential of products fortified with broccoli by-products is even higher. This study also confirmed that underestimated by-products of broccoli processing can be a valuable additive to GF improving its nutritional and functional quality.

The presence of phenolic compounds, besides the improvement of antioxidant potential, can contribute also to other bioactive activities. The advanced glycation end-products (AGEs) are formed continuously in the human body, the intensity of AGEs formation is increased by hyperglycemia and oxidative stress status. Hence, the challenge is to evaluate food products with natural inhibitors of the AGEs formation. The AGEs inhibitory activity was monitored in two model systems of BSA-MGO and BSA-glucose. The extracts of BLP had high activity against the AGE formations (83.53%) in the BSA-MGO study, almost the same as the reference material of aminoguanidine (84.03%). Additionally, a high effectiveness against AGE formation was noted in GFs after the addition of BLP (77.60%) in comparison to the control (67.47%). Therefore, the incorporation of BLP resulted in 1.15 times higher anti-AGE activity of the designed gluten-free product.  The results obtained in this study are in agreement with other studies utilisin g bypr oducts in bread formulation to improve the anti-AGE activity.

Conclusions

Based on the results obtained, it can be noticed that BLP can be successfully used as an additive in gluten-free bakery particular, the specific volume and the bake loss of GFB have been significantly improved, compared to GFB, human intervention studies are needed.

Reference:

Krupa-Kozak, U., Drabińska, N., Bączek, N., Šimková, K., Starowicz, M. and Jeliński, T., 2021. Application of broccoli leaf powder in gluten-free bread: An innovative approach to improve its bioactive potential and technological quality. Foods, 10(4), p.819.