LWT - Food Science and Technology 186 (2023) 115245 Contents lists available at ScienceDirect LWT journal homepage: www.elsevier.com/locate/lwt Enriched nutraceuticals in gluten-free whole grain rice cookies with alternative sweeteners Hameeda Banu Itagi a,1, Kristel June D. Sartagoda b,1, Nitesh Gupta a, Vipin Pratap a, Priyabrata Roy a, Rhowell N. Tiozon Jr. b, Ahmed Regina a, Nese Sreenivasulu a,b,* a Centre of Excellence in Rice Value Addition (CERVA), International Rice Research Institute (IRRI)-South Asia Regional Centre (ISARC), Varanasi, Uttar Pradesh, India b International Rice Research Institute, Los Baños, Philippines A R T I C L E I N F O A B S T R A C T Keywords: Cookies are a popular snack worldwide, but the presence of gluten in most wheat-based cookies poses problems Gluten-free cookies for people with gluten intolerance. Furthermore, gluten-free products are often deficient in nutraceuticals. This Rice study investigated the potential of two traditional Indian rice landraces, Kalanamak and Chak-hao, as alternative Whole grain cereals for producing whole grain gluten-free cookies with enriched bioactive compounds. The study also Antioxidants Sensory evaluation evaluated the influence of whole grain rice flours (WGRFs) and different sweeteners on the physical and Nutraceuticals biochemical properties of the cookies. The substitution of refined wheat flour with WGRFs significantly affected the physical and chemical properties of the cookies. WGRF cookies were generally crispier and had a lower spread ratio resulting in higher sensory evaluation scores. The added health benefits of WGRF derived cookies are likely due to the inherently higher levels of bioactive compounds such as quercetin equivalents with higher hydrogen peroxide scavenging (HPS) capacity and antioxidant activity derived from 2,2-diphenyl-1-picrylhydra- zyl (DPPH) in Chak-hao rice and jaggery. This work shows that WGRFs from Kalanamak and Chak-hao could be viable alternatives to refined wheat flour for producing gluten-free cookies with enhanced nutraceutical benefits. 1. Introduction taste (Kowsalya et al., 2022). Both of these landraces are widely culti- vated in geographical indicator regions. Our previous research has Rice (Oryza sativa L.) has been a fundamental part of the human diet shown that popped rice made from these landraces retain high levels of for thousands of years and has played a significant role in the green phytochemicals and antioxidants, making them not just flavorful, but revolution to achieve food security in Asia. However, the milling process also nutritious (Itagi et al., 2023). Due to changes in lifestyle and so- removes the outer bran and embryo, which results in white rice grain cioeconomic conditions and increased awareness of their nutritional with a high glycemic index and reduced nutrient content (Anacleto benefits, pigmented rice, as a stand-alone food product or as an ingre- et al., 2019). In contrast, brown rice is an unpolished whole grain that dient in food products, has attracted increased attention in recent years contains more dietary fiber, amino acids, phytosterols, phenolics, and (Itagi et al., 2023; Kasote et al., 2021). Therefore, the deployment of bioactive compounds compared to white rice (Brotman et al., 2021; geographical indicator (GI)-tagged rice landraces can help in the Tiozon et al., 2021). Additionally, pigmented rice varieties, such as red development of additional rice food products with unique and desirable rice, black rice, and purple rice, have been found to be even more traits to diversify the consumer demands. nutrient-dense than brown rice due to their enriched antioxidant prop- A considerable proportion of the world population exhibits intoler- erties (Itagi et al., 2023; Mbanjo et al., 2020). Kalanamak, which gets its ance and sensitivity to gluten, which is an integral component of grains name from the black husk (kala) and salt (namak), is a prominent belonging to the Triticeae tribe such as wheat, rye, barley, and oats. The landrace from Uttar Pradesh. Chak-hao, a black rice accession, is a development of a gluten-free (GF) diet to cater for people with gluten fragrant variety of sticky rice, which derives its name from its delicious intolerance and celiac disease has been undertaken by studying * Corresponding author. Centre of Excellence in Rice Value Addition (CERVA), International Rice Research Institute (IRRI)-South Asia Regional Centre (ISARC), Varanasi, Uttar Pradesh, India. E-mail address: n.sreenivasulu@irri.org (N. Sreenivasulu). 1 Contributed equally. https://doi.org/10.1016/j.lwt.2023.115245 Received 22 July 2023; Received in revised form 27 August 2023; Accepted 30 August 2023 Available online 6 September 2023 0023-6438/© 2023 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/). H.B. Itagi et al. L W T 186 (2023) 115245 alternative starch sources from grain outside of the tribe Triticeae (Vici in a laboratory mill (UDY Corporation, Cyclone Sample Mill, Screen- et al., 2016). Due to the increased prevalence of gluten intolerance and 0.25 mm, Fort Collins, Colorado, USA) to make rice flour and sieved to a celiac disease, the market for GF foods are projected to reach almost $24 particle size of about 1 mm. The rice flour samples were stored in billion by 2027 (Aguiar et al., 2023). It is well known that rice is airtight containers in a refrigerator (4 ◦C), before being subjected to extensively used to make GF foods, but most of these prior studies used product formulations. refined white sugar and commercial rice flour normally made from polished rice, which is rich in starch and lacks nutrients (Paz et al., 2.4. Reagents, solvents, and standards 2020). Because of this, GF diets usually lack adequate levels of essential nutrients, including micronutrients and bioactives, which can have a Analytical grade reagents were utilized to measure nutritional number of negative health implications. To ensure that people receive components. The National Institute of Standards and Technology the best dietary intervention, it is crucial to develop GF food products (NIST®) (Gaithersburg, MD, USA) provided the standard reference derived from whole grain of nutritious rice varieties or landraces and material (rice flour, 1568 b). Methanol and hexane of HPLC grade were assess the nutritional value of foods that fall within this diet group (Vici procured from Fisher Scientific Co. in India. Takadiastase, DPPH (1,1- et al., 2016). diphenyl-2-picrylhydrazyl radical), and Folin-Ciocalteu (FC) reagent Functional foods and nutraceuticals, that offer benefits beyond were provided by Parke, Davis and Co., Ltd. in the United States and SRL meeting caloric requirements, are becoming more and more familiar to (Sisco Research Laboratories Pvt. Ltd., India), respectively. Merck, India, consumers. Cookies are a widely consumed snack that is crunchy and provided the sodium hydroxide, other reagents, solvents, and standards sweet, which are either baked or fried. In general, cookies are handy and for fatty acids, vitamins, and minerals. All aqueous solutions were pre- have a long shelf life and these are created from a blend of ingredients pared using ultrapure water (Milli-Q water purification system, LAB Q including flour, sugar, eggs, and fats (Giuberti et al., 2017). Jaggery is a Ultra, India). natural sweetener that is popular in India due to its perceived health advantages, wherein the majority of the vitamins and minerals present 2.5. Cookie formulation and preparation in sugarcane are retained. Hence substituting white sugar with sweet- eners like brown sugar and jaggery in cookies offer extra functionality Cookies were prepared following the Approved Method 10-50D and health advantages (Iqbal et al., 2017). (AACC, 2000) with slight modifications on ingredients: RWF (control) Despite vast research on the role of rice in a GF diet, traditional and WGRF (from Kalanamak or Black rice) (100 g), salted butter (65 g), whole-grain Indian rice landraces have not been fully investigated to test water (12–14 mL), sweetener (white sugar, brown sugar or jaggery) (50 its potential to develop GF-free functional foods and cookies. Addi- g/100 g) and chopped cashew nuts as toppings (2 g/100 g). The tionally, little information is known about the physicochemical, func- creaming process of the salted butter and powdered sweetener was tional, and nutritional qualities of landraces when they are added to carried out in a planetary mixer (Berjaya laboratory in Kuala Lumpur, food matrices. The goal of the current work was to create formulations of Malaysia). Subsequently, flour and water were added to the mixture. novel, functional, nutraceutical-rich, gluten-free whole grain rice flour The cookie batter was portioned using a standard tablespoon and for making cookies using two well-known, GI-tagged, aromatic Indian manually shaped into circular forms. The baking process was conducted rice landraces (Kalanamak from Uttar Pradesh and Chak-hao from in a commercial baking oven (Arise Equipments, New Delhi, India) that Manipur) and raw sugarcane products (jaggery and brown sugar). had been preheated to a temperature of 180 ◦C, for 18–20 min. After Cookies made as a result were examined for their physical and chemical cooling for 30 min at ambient temperature, the cookies were subse- properties, nutritional content, nutraceuticals, and sensory qualities. quently transferred to an airtight container made of clear polyethylene The findings of this study will offer useful knowledge for manufacturing terephthalate (PET). A comparative analysis was conducted to assess the GF high-quality convenience snacks from Indian rice landraces that are physical, nutritional, nutraceutical, and sensory attributes of cookies nutrient-dense and palatable. prepared using Kalanamak and Chak-hao WGRF in contrast to those made with RWF. 2. Material and methods 2.6. Water activity (aw) and moisture content (MC) 2.1. Raw materials The MC of flour and cookie samples (2 g) was determined according Popular GI-tagged landraces Chak-hao (black aromatic waxy rice) to the AOAC (1990) approved method. The aw of samples was measured and Kalanamak aromatic rice (non-pigmented) were cultivated under using a Model Series 4 TE water activity analyzer (Aqua Lab, Meter well-maintained irrigated conditions during the monsoon season of Group Inc., Pullman, WA, USA). The experiments were conducted in 2021 at the International Rice Research Institute South Asia Regional triplicates (Itagi et al., 2023). Centre (ISARC) experimental farm in Varanasi, Uttar Pradesh. Paddies were dehusked, milled and prepared rice flour according to Itagi et al. 2.7. Physical characteristics and texture of cookies (2023). The refined wheat flour (RWF), salted butter, white sugar, brown sugar, and jaggery available in local standard brands were pur- Using an analytical balance (GR-202, A&D Co., Japan), cookie chased from the local market (Varanasi, India). weight was calculated. A digital Vernier caliper with 0.001mm accuracy was used to measure the cookies diameter and thickness. The diameter/ 2.2. Shelling/dehulling of paddy for preparation of whole grain rice thickness formula was used to calculate the spread ratio (SR) of cookies after baking (Naseer et al., 2021). A colorimeter (Chroma Meter CR-410, The Paddy De-Husker (Model No. 67004, Osaw Industrial Products, Konica Minolta, Inc., Osaka, Japan) was used to assess the color of the Pvt. Ltd., Indosaw, Ambala, Haryana, India) was used to de-hull around cookies. The measuring head was placed in the center of each sample 1 kg of paddy. Rubber rollers had to be adjusted in order to produce after the instrument’s calibration was completed using a reference dehulled whole grain rice. standard that was white in color. Color values using the CIE L* a* b* scales were recorded using five samples for each cookie formulation. 2.3. Preparation of whole grain rice flour (WGRF) Following that, the mean values were documented as L* = lightness (100 = white, 0 = black), a* (-a* = greenness, +a* = redness), and b* Grain that had been dehulled was washed and dried. The dehulled (-b* = -blueness, +b* = yellowness) (Selvakumaran et al., 2019). The rice grains from the Kalanamak and Chak-hao varieties were processed browning index was calculated following (Klunklin & Savage, 2018). 2 H.B. Itagi et al. L W T 186 (2023) 115245 The texture analysis of cookies was conducted to determine their 2.8.6. Estimation of total phenolic content and antioxidant potential breaking strength, utilizing a TA. XT plus texture analyzer (Stable Micro Systems Ltd, Surrey, UK) equipped with a 50 kg load cell. The peak force 2.8.6.1. Extraction of phenolic content. A 1g sample was subjected to required to break a single whole cookie was recorded and the average extraction using 10 mL of petroleum ether in an ultrasonicator (PCI value of ten replicates was reported (Pal et al., 2019). Analytics, India) for 15 min. After centrifugation (5 min at 2520×g), the supernatant was decanted and collected. The polyphenols were extrac- 2.8. Nutritional characteristics and bioactive potential of flours and ted from defatted samples following Itagi et al. (2023). cookies 2.8.6.2. Total phenolic content (TPC). FC reagent (800 μl) and Na2CO3 2.8.1. Nutrient composition (7.5 g/100 mL) (2 mL) were added to 200 μl of sample. Then, the sample The Kjeldahl method was employed to determine the protein content volume was increased to 7 mL with deionized water and then left to (Kjeltec™ 8200, Kjeltec, Foss, Sweden). The fat was extracted with pe- stand in a lightprotected environment for 30 min. The absorbance was troleum ether (40–60 ◦C) through a Foss ST243 Soxtec Extraction Unit taken at 725 nm. TPC was reported according to Itagi et al. (2023). and quantified through gravimetric analysis (Itagi et al., 2023). The determination of ash content was carried out through the incineration of 2.8.6.3. Total flavonoid content (TFC). The extract (1 mL) was diluted to the samples at a temperature of 550 ◦C (AOAC, 2000). The estimation of 5 mL with ultrapure water, followed by the addition of NaNO2 (5 g/100 total carbohydrate in the samples was carried out following FAO (2003, mL) (300 μl). The mixture was incubated for 5 min. Next, AlCl3 (10 g/ p. 77) and the Gross Energy (Calories/100 g dry matter) was calculated 100 mL) (600 μl) was added, and the mixture was incubated for an based on the methods outlined by Ganogpichayagrai and Suksaard additional 6 min. A 2 mL NaOH (1 mol/L) was added, and the volume (2020). was adjusted to 10 mL. The quantification of TFC was performed by measuring the absorbance at a wavelength of 510 nm, and the resulting 2.8.2. Micronutrient analysis values were expressed as milligrams of catechin equivalents (CE) per All the trace mineral compounds and metals were quantified using an 100 g of the sample, as previously reported by Itagi et al. (2023). ICP-MS (Agilent 7800 ICP-MS) by following the prior protocol outlined in Itagi et al. (2023). For the vitamin analyses, finely ground samples (5 2.8.6.4. Total anthocyanin content (TAC). A mixture comprising of 2 mL g) were weighed in a 100 mL volumetric flask, added with HCl (30 mL, of potassium chloride buffer (0.03 mol/L, pH 1.0) and 2 mL of sodium 0.1 mol/L), and incubated at 120 ◦C for 30 min. Then, the pH was acetate buffer (0.4 mol/L, pH 4.5) was introduced to 20 μl of extract. adjusted to 7 with NaOH (0.1 N). Takadiastase (5 mL, 1 mol/L) was Following a 15min incubation period, the absorbance was quantified at added, followed by overnight incubation at 35 ◦C. The sample was 550 nm and 700 nm relative to a blank sample consisting of ultrapure diluted to 100 mL with distilled deionized water and filtered (0.22 μm, water. TAC was recorded in terms of milligrams of cyanidin-3-glucoside PVDF Whatman filter paper). A 10 μL of the filtrate and standards were (C-3-G) equivalents per 100 g of the sample (Itagi et al., 2023). analyzed using LC-MS/MS that had an autosampler and MS detector (Agilent Technologies, 6470 Triple Quad LC-MS/MS). This system was 2.8.6.5. Total antioxidant capacity. The extract (500 μL) was combined equipped with a 1.8-μm Agilent ZORBAX RRHD Eclipse Plus C-18 sta- with phosphomolybdenum reagent (0.6 mol/L sulfuric acid, 28 mmol/L tionary phase in 3.0 mm × 100 mm formats. The mobile phase of sodium phosphate, and 4 mmol/L ammonium molybdate) (1.23 mL) and gradient delivery composed of a mixture of solvent A (water: formic incubated at 90 ◦C for 90 min. The total antioxidant capacity was re- acid, 100:0.3, v/v) and B (methanol: formic acid, 100:0.3, v/v) and had ported as quercetin equivalents (QE) per 100 g of the sample at an a flow rate of 0.50 mL/min (0–0.4 min, 1% B; 0.4–6 min, 1%–45% B; absorbance of 695 nm (Itagi et al., 2023). 6–7.5 min, 45–90% B; 7.5–9min, 90%–1% B). The standard vitamins B1, B2, B5, and B6 appear at 0.96, 7.2, 4.8, and 2.2 min retention times (Rezaei et al., 2022). 2.8.6.6. Hydrogen peroxide scavenging capacity (HAS). The extract (0.4 mL) was combined with 40 mmol/L H2O2 (0.6 mL). The mixture was 2.8.3. Dietary fiber (DF) content diluted to 2 mL with 50 mmol/L sodium phosphate buffer (pH 7.4) and The Megazyme K-TDFR kit (Megazyme Wicklow, Ireland) was uti- then incubated for 40 min at 30 ◦C. The findings were reported as mg lized to evaluate the levels of total dietary fiber (TDF), insoluble (IDF), quercetin equivalents (QE) per 100 g of sample (Itagi et al., 2023). and soluble (SDF). The computation of IDF and SDF was performed utilizing the Megazyme Mega-Calculation method (Itagi et al., 2023). 2.8.6.7. DPPH radical scavenging activity. The DPPH assay was con- ducted on 500 μL of extract according to Itagi et al. (2023). The results 2.8.4. Total sugar, total starch, and amylose content were represented in terms of % DPPH radical scavenging activity. The total starch was conducted utilizing a Megazyme assay kit spe- cifically designed for total starch (Megazyme K-TSTA, Wicklow, Ireland) 2.8.6.8. Ferric reducing antioxidant power (FRAP). A 200 μL of volume (Itagi et al., 2023). The determination of amylose content was conducted of freshly made FRAP reagent (300 mmol/L acetate buffer (pH 3.6):10 in accordance with the methodology outlined by Cuevas et al. (2018), mmol/L 2,4,6-tri (2- pyridyl)-1,3,5-triazine solution:20 mmol/L FeCl3 and the categorization of samples was based on the contents as described solution, (10:1:1, v:v:v)) was added to 20 μL of extract. At 620 nm, the by Graham (2002). The quantification of the overall quantity of soluble mixture’s absorbance was measured after 5 min at 37 ◦C incubation. sugar present in the cookies was carried out using the anthrone method Results were expressed as mmol/L Trolox Equivalents per gram of ma- as described in Roy et al. (2021). terial (Tomasina et al., 2012). 2.8.5. Extraction and estimation of oryzanol 2.8.6.9. Targeted bioactive profiling. The defatted samples were used to Oryzanol extraction and estimation were done following the method extract the phenolic compounds. One milliliter of 80% aqueous meth- described in Itagi et al. (2023). The findings were reported in mg per anol (acidified with 1% HCl) was added to a 50 mg sample. After soni- 100 g, and all subsequent measurements were taken with the same cation for 10 min at 120 W, 800 μL of the supernatant was collected and spectrophotometer. centrifuged (28000×g, 10 min). Extraction was done twice, and the pooled supernatant was filtered through a 0.22 PVDF membrane filter, then subjected to liquid chromatography-electrospray ionization-tan- dem mass spectrometry (LC-MS/MS) to profile and identify phenolic 3 H.B. Itagi et al. L W T 186 (2023) 115245 compounds. The HPLC system used in this study was an Agilent 6470 0.05) using R statistical package, version 4.2.1 (R Foundation for Sta- Triple quad LC/MS System (Agilent Technologies, Santa Clara, CA, USA) tistical Computing, Vienna, Austria). To reveal a distinct clustering be- equipped with column C18, 2.1 × 100 mm, 1.8 μm which was used for tween data sets, the principal component analysis (PCA) scores plot was phenolic compound separation. Mobile phases A and B were composed used. The correlation between physical properties and texture profile, of solvent A (water: formic acid, 100:0.3, v/v) and B (methanol: formic nutritional composition, bioactives, and antioxidant activity for each acid, 100:0.3, v/v), respectively. The flow rate was set at 0.40 mL/min rice genotype was analyzed by Pearson’s correlation. All data visuali- (0–0.5 min, 10% B; 0.5–5 min, 10%–40% B; 5–12 min, 40–80% B; 12–15 zations were performed using the R statistical package. min, 80%–2% B) and the column was maintained at 40 ◦C. An aliquot of each sample solution (10 μL) was injected into the system equipped with 3. Results and discussion an ESI source and a triple-quadrupole mass spectrometer (MS/MS). The ESI source and the MS/MS were operated in the negative ion and posi- Kalanamak from Uttar Pradesh and Chak-hao from Manipur are tive ion multiple reaction monitoring (MRM) modes, respectively. All popular GI-tagged Indian rice landraces that are renowned for their measurements were conducted in duplicate. Calibrations with R2 = 0.99 aroma and superior grain quality (Kowsalya et al., 2022). The bran and were used. broken rice (byproducts of the rice milling process) of these two land- races are likewise known to possess higher nutraceutical properties. 2.8.7. Fatty acid (FA) profiling Rather than being used as a low-value commodity for fodder, these FA content in samples was analyzed following a method by Jarukas byproducts could be instead leveraged as raw material in food appli- et al. (2020), with slight modifications. Powdered samples were cations with enriched functional properties for different value-added extracted with hexane (1:7, w/v) in a shaking water bath (65 ◦C, 30 product development purposes. In this study, whole grain rice flour min), then centrifuged at 7000×g for 15 min. Total lipid fraction was derived from the well-known aromatic rices, Kalanamak (greenish recovered after solvent removal in a stream of nitrogen. The samples brown) and Chak-hao (black rice), and raw sugarcane products (jaggery were then derivatized using 2 mL of 7% (v/v) BF3 in methanol and 1 mL and brown sugar) were used to explore novel formulations of of toluene and placed in a warm bath at 80 ◦C for 45 min. After the nutraceutical-rich gluten-free cookies. The physicochemical, nutri- addition of 5 mL of distilled water, the trans-methylated FAs were tional, and sensory properties of these cookies were compared with that extracted with 1 mL hexane. The aliquot of the hexane phase was of refined wheat-based cookies and discussed. analyzed by gas chromatography. An Agilent 7890B gas chromatograph (Agilent Technologies, USA) with a Flame-Ionization Detector was used 3.1. Physicochemical properties of flour and cookie samples to separate and quantify FAs. One microliter aliquot of the hexane phase was injected in split-mode onto a DB-Wax column (30 m × 0.25 mm ID, Along with texture and flavor, the surface color of a baked product is 0.25 μm DB-Wax (J&W 122–7032)). The injector temperature was set at a crucial factor in the initial acceptability of baked products among 250 ◦C, detector at 280 ◦C, oven at 50 ◦C initially, then 50 ◦C, 1 min, consumers. The flour and sweetener had remarkable and complex effects 25 ◦C/min to 200 ◦C, 3 ◦C/min to 230 ◦C, 18 min. The carrier gas was on the color formulation of the resulting cookies, as shown in Fig. 1. A Hydrogen. Detector gasses were Hydrogen: 40 mL/min; Air: 450 higher L* value, which indicates lightness, is observed in refined wheat mL/min; Helium make-up gas: 30 mL/min. An electronic pressure flour (RWF) and Kalanamak whole grain rice flour (KWGRF)based control in the constant flow mode was used. The FAME calibration cookies than in black whole grain rice flour (BWGRF)based cookies. standards were used for the quantification of FAs in the various lipid RWF cookies made with white sugar were noted with the highest L* extracts. values, which could be attributed to the typical light color of well-milled wheat kernels. The lower L* values of all BWGRF cookies, on the other 2.9. Sensory evaluation hand, can be attributed to the presence of pigmented bran in the present study. These findings align with previous studies by Joo and Choi (2012) A hedonic sensory evaluation was conducted on cookies made from and Jang et al. (2010) that observed a decreasing L* value with whole grain rice flour. The evaluators were 27 volunteer staff members increasing rice bran substitution in a cookie formulation. Moreover, the from ISARC (Varanasi) (untrained), ranging in age from 23 to 58 years L* value of Kalanamak whole grain rice flour white sugar (KWGRFWS) and including both male and female participants. Each of the panelists cookie was not substantially different from refined wheat flour white has provided their informed consent to partake in the study. The sensory sugar (RWFWS) cookie and was similar to that reported by Joo and Choi evaluation was conducted within the confines of the sensory and prod- (2012) for cookies made from commercially available rice flour. Dif- uct development laboratory at CERVA. The cookies were prepared in ferences in rice flour color were attributed to their polyphenols, which advance of the sensory evaluation and were subsequently stored at relate to the purple color of the rice grain (Buenafe et al., 2022; Klunklin ambient temperature. In the context of sensory evaluation, the samples & Savage, 2018). The L* values generally decreased with the addition of were presented in their entirety on white plastic dishes that were labeled jaggery as it contains reducing sugars that could participate in the with codes. The panelists were then served the samples in a randomized Maillard reaction. Chand et al. (2011) reported that reducing sugars in sequence. The panelists were furnished with distilled water and unsalted jaggery typically increases with storage regardless of the storage con- crackers to rinse their palates in between tastings. The cookies under- ditions. The Maillard reaction, which occurs during the baking process, went an evaluation process that assessed their surface color, surface is another factor that affects the final color of baked food items (Giuberti cracking pattern, crumb color, texture, mouth feel, flavor, aroma, and et al., 2017). This reaction could result in reddish-brown hues from the overall acceptability. The evaluation was conducted using a nine-point interaction of reducing sugars with proteins and explain the lower L* hedonic scale which ranged from “like extremely” to “dislike value of 60.49 for refined wheat flour brown sugar (RWFBS) cookie extremely,” corresponding to the highest and lowest scores of “9” and compared to a higher value of 67.00 for Kalanamak whole grain rice “1” respectively (Naseer et al., 2021). flour brown sugar (KWGRFBS) cookie (Ryan & Brewer, 2006). All WGRF cookies had significantly lower redness (a*) and yellowness (b*) 2.10. Statistical analysis compared to RWF cookies, with BWGRF showing the lowest values for both parameters. The differences in a* and b* values were presumably The experiments were performed in triplicate (n = 3), unless stated correlated with the differences in the amino acid profile and content of otherwise, and the results were reported as the mean values along with reducing sugars responsible for the intensity of the Maillard reaction their standard deviations. Data from the results were analyzed statisti- (Torbica et al., 2012; Zucco et al., 2011). cally using one-way ANOVA, followed by a Tukey’s post hoc test (P < The physical and chemical characteristics of RWF and WGRF from 4 H.B. Itagi et al. L W T 186 (2023) 115245 Fig. 1. Color measurements of cookies from RWF and WGRFs from Kalanamak and Chak-hao rice with different sweeteners. Abbreviations: RWF–Refined wheat flour; WGRF–Whole grain rice flour. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.) Kalanamak and Chak-hao rice, and their corresponding cookies were flour samples (BWGRF and KWGRF) have a satisfactory shelf life. Due to shown in Table 1. The WGRF samples exhibited lower moisture content water evaporation during baking, which results in the distinctive crusty (9.93% and 10.13% for KWGRF and BWGRF, respectively) compared to features of cookies, all cookie formulations exhibited considerably lower RWF (13.71%). These findings are consistent with previous studies that moisture content in PC1 than flour samples (Fig. 2b). Regardless of flour have also reported lower moisture content in rice flour relative to wheat type, the moisture content values of cookies prepared with jaggery were flour (Islam et al., 2012; Rai et al., 2014; Torbica et al., 2012). Moisture up to 51% higher than those prepared with refined white sugar. This content is a crucial consideration for flour storage since flour with a distinction may be ascribed to the hygroscopic character of inverted moisture content exceeding 13% is more prone to microbial deteriora- sugar and the presence of mineral ions in jaggery (Lamdande et al., tion (Oppong et al., 2021). Accordingly, the moisture content of WGRFs 2018; Rao & Singh, 2022). Apart from moisture content, the water ac- in the present study was below the threshold level, indicating that both tivity (aw) also exhibited lowest in cookies, in comparison to flour (PC1, Table 1 Physicochemical properties of refined wheat flour and whole grain rice flour samples and resulting cookies with different sweeteners. Samples/ Refined Refined Wheat Flour Cashew Kalanamak Kalanamak Whole Grain Rice Black Black Whole Grain Rice Cashew Parameter Wheat Cookies Whole grain Cashew Cookies Whole Cookies Flour Rice Flour grain Rice White Brown Jaggery White Brown Jaggery Flour White Brown Jaggery Sugar Sugar Sugar Sugar Sugar Sugar Moisture 13.71 ± 2.44 ± 4.10 ± 3.74 ± 9.93 ± 0.12b 2.02 ± 2.70 ± 3.42 ± 10.13 ± 2.53 ± 3.18 ± 3.78 ± Content 0.04a 0.09g 0.04c 0.03d,e 0.15h 0.16g 0.10e,f 0.04b 0.09g 0.05f 0.12c,d (%) Water 0.71 ± 0.28 ± 0.35 ± 0.31 ± 0.40 ± 0.00b 0.22 ± 0.26 ± 0.29 ± 0.43 ± 0.29 ± 0.33 ± 0.33 ± Activity 0.01a 0.02e,f 0.02c 0.01d,e 0.01g 0.01f,g 0.01d,e,f 0.01b 0.002d,e, 0.01c,d 0.01c,d (aw) f Total starch 75.13 ± 56.29 ± 56.87 ± 54.86 ± 80.0 ± 0.32a 55.10 56.48 ± 61.62 ± 72.85 ± 56.14 ± 57.36 ± 57.57 ± (g/100g) 0.04b 0.12d 0.65d 1.66d ± 0.07d 0.72d 0.20c 0.72b 0.96d 0.53c,d 1.60c,d Total 21.80 ± 16.41 ± 16.26 ± 17.03 ± 16.11 ± 0.4c 13.87 14.29 ± 13.96 ± 11.17 ± 5.16 ± 4.75 ± 3.79 ± amylose 0.00a 0.01b,c 0.1c 0.1b ± 0.00d 0.2d 0.00d 0.04e 0.1f 0.01f 0.1g (g/100g) Total sugar – 23.97 ± 23.27 ± 20.33 ± – 30.77 26.97 ± 20.77 ± – 30.40 ± 27.80 ± 22.20 ± (g/100g) 1.40b,c, 3.71c,d 1.85d ± 2.04a 1.24a,b, 2.07d 1.99a 2.54a,b 1.33d d c Diameter (D, – 40.44 ± 39.00 ± 39.44 ± – 37.33 37.00 ± 36.00 ± – 39.89 ± 39.67 ± 39.00 ± mm) 0.87a 0.72a,b 0.42a ± 0.27c 0.27c 0.16a 0.54a 0.27a,b 0.27b,c Thickness – 12.00 ± 12.78 ± 12.78 ± – 12.67 12.78 ± 12.67 ± – 12.67 ± 13.22 ± 13.33 ± (T, mm) 0.27b 0.16a,b 0.16a,b ± 0.58a, 0.16a,b 0.27a,b 0.27a,b 0.16a 0.27a b Spread ratio – 3.37 ± 3.05 ± 3.09 ± – 2.95 ± 2.90 ± 2.84 ± – 3.15 ± 3.00 ± 2.93 ± (D/T) 0.15a 0.06b,c, 0.05b,c 0.04b,c, 0.04c,d 0.05d 0.06a,b 0.02b,c, 0.07b,c,d d d d Weight (g) – 8.11 ± 7.78 ± 8.40 ± – 8.11 ± 7.96 ± 8.02 ± – 8.69 ± 8.76 ± 8.44 ± 0.17b,c, 0.11d 0.05a,b,c 0.03b,c, 0.19d 0.06c,d 0.06a 0.11a 0.13a,b d d Hardness – 3.97 ± 5.01 ± 4.31 ± – 2.33 ± 2.49 ± 2.25 ± – 3.98 ± 4.60 ± 5.40 ± (N) 0.54c 0.62a,b 0.44b,c 0.37d 0.20d 0.23 d 0.40c 0.64b,c 0.69a Values are expressed as the mean of three (3) replicates for moisture content, aw, total amylose, and total sugar, and ten (10) for thickness, diameter, spread ratio, weight and, hardness parameters ± standard deviation; “- “–Not applicable parameters; Different lowercase and uppercase letters denote a significant difference (P < 0.05). 5 H.B. Itagi et al. L W T 186 (2023) 115245 Fig. 2. Principal component analysis of RWF and WGRF and resulting cookies with different sweeteners. Abbreviations: RWF–Refined wheat flour; WGRF–Whole grain rice flour; RWFWS (Refined wheat flour white sugar); RWFBS (Refined wheat flour brown sugar); RWFJ (Refined wheat flour jaggery); KWGRF (Kalanamak whole grain rice flour); KWGRFWS (Kalanamak whole grain rice flour white sugar); KWGRFBS (Kalanamak whole grain rice flour brown sugar); KWGRFJ (Kalanamak whole grain rice flour jaggery); BWGRF (Black whole grain rice flour); BWGRFWS (Black whole grain rice flour white sugar); BWGRFBS (Black whole grain rice flour brown sugar); BWGRFJ (Black whole grain rice flour jaggery). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.) Fig. 2b). All cookie samples had values in the range of 0.28–0.35, while WGRF resulted in a slight reduction in the spread ratio of cookies. the wheat flour exhibits less than 0.85. Interestingly, the KWGRFWS However, the spread ratio values of BWGRF cookies were almost cookies have the lowest aw value (0.22) (Table 1). Lower MC and aw equivalent to those of the control, indicating that they are commercially values are crucial for extending the shelf life of the final product and viable. Additionally, BWGRF cookies were heavier than RWF and reducing the risk of foodborne illnesses. In addition, appropriate pack- KWGRF cookies. Ryan and Brewer (2006) observed that cookies made aging materials are also necessary for quality preservation and shelf-life with low-protein flour tended to be smaller in diameter and have lower extension (Itagi et al., 2023; Yildiz & Gocmen, 2021). spread ratios, indicating denser properties. The term “hardness”, which describes the amount of force used to distort a sample, is measured among various cookies. In general, the hardness values of RWF (3.97–5.01N) and BWGRF (3.97–5.40N) cookies 3.2. Nutritional properties of flour and cookie samples were comparable. Interestingly, all KWGRF cookies had a low breaking point (2.25–2.49 N) (Table 1). The contribution to PC1 (Fig. 2b) was 3.2.1. Macronutrient content significantly influenced by the hardness value and spread ratio, which Although RWF had the highest protein levels (13.24 g/100g), fol- accounts for the characteristic features of the cookie samples. These lowed by BWGRF (12.41 g/100g), and KWGRF (10.55 g/100g), the results are consistent with those of Yildiz and Gocmen (2021) and Paz cookies derived from the flour has substantially lowered the protein et al. (2020), who reported a reduction in hardness values when content (Table 2). RWF cookies contained protein ranging between 7.90 substituting rice flour for wheat flour in a cookie formulation. In addi- and 8.50%. Compared to KWGRF cookies (7.09–7.80 g/100g), BWGRF tion, these studies attribute the high hardness values to the higher cookies retained higher protein levels (7.17–8.50 g/100g). Prior studies protein content of RWF compared to WGRF. Proteins on the surface of reported cookies made from rice and wheat flour of regular varieties starch granules function as adhesives, which strengthen the starch–- (Islam et al., 2012; Klunklin & Savage, 2018; Torbica et al., 2012). Also protein bond in cookie dough and increase the overall hardness of the observed was a significant difference in the total carbohydrates of the cookie (Ryan & Brewer, 2006). In addition, a higher DF content may also flour samples. KWGRF was found to have the highest carbohydrate level enhance the texture of cookies. This was the case with GF cookies made (74.15 g/100g), which was 3% and 4% higher than BWGRF and RWF, with almond flour (Yildiz & Gocmen, 2021). Likewise, BWGRF had a DF respectively. Variations in flour carbohydrate content could be attrib- that was up to 49% greater than the other two flour types, which may uted to differences in protein and lipid content (Oppong et al., 2021). explain the higher hardness values of BWGRF cookies. Yildiz and Goc- However, in this study, only protein showed a strong positive correlation men (2021) propose that DF can reduce the amount of free water in the with total carbohydrates (Fig. 4b). All cookie samples exhibited a sig- dough, thereby decreasing the resulting spread ratio of cookies. nificant drop in carbohydrates, which may be ascribed to amylose and Differences in starch, DF, and protein proportions in the flour could amylopectin leaching from the starch granules when it swells during the account for the dimensional property variations (Mudgil et al., 2017). thermal process (Itagi et al., 2023). Diameter, thickness, and spread ratio are typical parameters used to The fat content of flour samples ranged from 0.87 in RWF to 3.34 g/ determine cookie quality. The cookies in the study varied in diameter 100g in KWGRF (Table 2). These differences in lipid content are pri- (37–40.44 mm), with significant differences observed primarily among marily attributed to the presence of bran components in both WGRFs, KWGRF cookies (Table 1). In addition, the WGRF substitution led to an which are virtually absent from RWF due to the nature of processing increase in the overall thickness of the cookies. These changes are re- (Ciccoritti et al., 2017; Oppong et al., 2021). The addition of extra fat flected in spread ratio values. In general, cookies with a higher spread resulted in an increase of total fat in the range 25.83–27.76 g/100g ratio are regarded as the most commercially appealing (Giuberti et al., (Table 2). The fat content of resulting cookies remains higher, regardless 2017; Mudgil et al., 2017). Table 2 suggests that replacing RWF with of the sweetener used (Klunklin & Savage, 2018). The fatty acid profile of BWGRF and KWGRF cookies generally 6 H.B. Itagi et al. L W T 186 (2023) 115245 Table 2 Nutritional composition of refined wheat flour and whole grain rice flour samples and resulting cookies with different sweeteners. Samples/ Refined Refined Wheat Flour Cashew Kalanamak Kalanamak Whole Grain Rice Black Black Whole Grain Rice Cashew Parameter Wheat Cookies Whole grain Cashew Cookies Whole Cookies Flour Rice Flour grain Rice White Brown Jaggery White Brown Jaggery Flour White Brown Jaggery Sugar Sugar Sugar Sugar Sugar Sugar Total 71.43 ± 62.27 60.77 59.94 ± 74.15 ± 0.44a 62.27 ± 61.60 ± 58.62 ± 72.29 ± 62.01 61.08 ± 58.70 ± Carbohydrate 0.13b ± 0.38c ± 0.30e 0.49c 0.23c,d 0.62f 0.14b ± 0.29c 0.30c,d, 0.18f (g/100g) 0.18d,e e Protein (g/100g) 13.24 ± 7.90 ± 7.90 ± 8.50 ± 10.55 ± 0.39b 7.09 ± 7.80 ± 7.27 ± 12.41 ± 7.17 ± 7.27 ± 8.50 ± 0.11a 0.22c,d 0.20c,d 0.30c 0.09d 0.21c,d 0.45d 0.16a 0.23d 0.14d 0.07c Fat (g/100g) 0.87 ± 25.87 25.83 26.01 ± 3.34 ± 0.02g 27.76 ± 26.67 ± 27.67 ± 3.02 ± 26.35 27.15 ± 26.98 ± 0.13h ± 0.20f ± 0.09f 0.28e,f 0.19a 0.18c,d 0.44a 0.26g ± 0.15b 0.16b,c 0.15d,e Ash (g/100g) 0.64 ± 1.19 ± 1.19 ± 1.80 ± 1.85 ± 0.10c 1.41 ± 1.46 ± 2.20 ± 1.90 ± 1.40 ± 1.44 ± 2.07 ± 0.07g 0.06f 0.06e,f 0.07c 0.04d,e 0.05d 0.04a 0.05b,c 0.04d, 0.08d 0.01a,b e,f Energy (Calories/ 347.5 ± 516.6 509.0 508.1 ± 370.6 ± 0.62d 522.3 ± 518.3 ± 517.5 ± 368.3 ± 518.8 516.7 ± 511.4 ± 100g) 0.18e ± ± 1.24c 0.90c 2.60a 2.94a 0.61a 0.24d ± 2.41a 1.57a,b 0.89b,c 1.80a,b Insoluble Dietary 1.88 ± 1.31 ± 1.33 ± 1.67 ± 3.10 ± 0.02a,b 2.51 ± 2.18 ± 2.23 ± 4.12 ± 2.62 ± 2.72 ± 3.24 ± Fiber (g/100g) 0.21c,d 0.00d 0.40d 0.09c,d 0.12b,c 0.18b,c, 0.06b,c, 0.32a 0.09b,c 0.17b,c 0.26a,b d d Soluble Dietary 1.28 ± 1.34 ± 0.71 ± 0.75 ± 0.77 ± 0.03a,b 0.67 ± 0.44 ± 0.13 ± 1.03 ± 0.17 ± 0.56 ± 0.89 ± Fiber (g/100g) 0.05a 0.13a 0.16a,b 0.04a,b 0.41a,b 0.04a,b 0.03b 0.27a,b 0.01b 0.20a,b 0.12a,b Total Dietary 3.16 ± 2.65 ± 2.05 ± 2.42 ± 3.13 ± 0.01b, 3.19 ± 2.62 ± 2.37 ± 5.16 ± 2.79 ± 3.28 ± 4.13 ± Fiber (g/100g) 0.26b,c,d 0.14c,d 0.24d 0.14c,d c,d 0.30b,c 0.22c,d 0.10c,d 0.05a 0.08c,d 0.38b,c 0.14a,b Minerals Magnesium (mg/ 428.1 ± 286.7 323.0 520.5 ± 1692 ± 5.82a 1010 ± 1069 ± 1497 ± 1491 ± 899.4 966.6 ± 1466 ± kg) 4.11g ± 2.74h ± 3.26f 15.99c,d 0.46c 20.92b 20.11b ± 2.27e 1.81d 16.55b 11.09h Potassium (mg/ 1806 ± 1082 ± 1406 ± 2287 ± 3056 ± 23.42d 1840 ± 1997 ± 4124 ± 3770 ± 2234 ± 2458 ± 4759 ± kg) 20.31g 13.79i 48.97h 16.98e,f 30.72g 24.90g 62.56b 28.97c 5.82f 6.75e 70.18a Manganese (mg/ 5.89 ± 3.90 ± 4.41 ± 5.84 ± 24.24 ± 0.07a 17.43 ± 17.48 ± 19.33 ± 21.71 ± 9.69 ± 10.29 ± 13.25 ± kg) 0.00g 0.05h 0.15h 0.02g 0.36d 0.01d 0.22c 0.12b 0.05f 0.08f 0.14e Iron (mg/kg) 15.25 ± 15.70 29.21 25.12 ± 18.33 ± 0.50c 19.76 ± 22.27 ± 34.25 ± 20.60 ± 21.99 19.65 ± 25.65 ± 0.40c ± 0.66c ± 5.32a, 0.14a,b,c 0.73b,c 1.69b,c 2.61a 0.63b,c ± 1.33b,c 0.32a,b,c b 0.78b,c Copper (mg/kg) 1.80 ± 1.31 ± 1.33 ± 1.36 ± 4.96 ± 0.10a 1.67 ± 1.90 ± 1.82 ± 3.19 ± 2.23 ± 2.01 ± 2.18 ± 0.01d,e 0.07f 0.11f 0.02f 0.01e,f 0.17c,d, 0.00d,e 0.00b 0.00c 0.05c,d, 0.03c,d e e Zinc (mg/kg) 7.42 ± 4.68 ± 4.93 ± 5.39 ± 17.45 ± 0.05b 8.72 ± 9.24 ± 9.88 ± 18.95 ± 9.42 ± 9.64 ± 11.23 ± 0.14f 0.13h 0.22g,h 0.05g 0.13e 0.18d,e 0.09d 0.09a 0.13d 0.01d 0.01c Selenium (mg/ 0.09 ± 0.07 ± 0.06 ± 0.06 ± 0.20 ± 0.01a 0.11 ± 0.10 ± 0.14 ± 0.08 ± 0.11 ± 0.09 ± 0.11 ± kg) 0.01c 0.02c 0.00c 0.01c 0.01b,c 0.00b,c 0.00a,b 0.00c 0.01b,c 0.02b,c 0.01b,c Vitamins Thiamine 0.40 ± 0.23 ± 0.17 ± 0.35 ± 0.32 ± 0.02b,c 0.71 ± 0.56 ± 0.41 ± 0.29 ± 0. 0.71 ± 0.56 ± 0.41 ± (Vitamin B1) 0.06b,c 0.03c 0.06c 0.04b,c 0.05a 0.10a,b 0.02b,c 02b,c 0.05a 0.1a,b 0.02b,c (ug/g) Riboflavin 0.25 ± 0.20 ± 0.26 ± 0.31 ± 0.22 ± 0.07a,b 0.25 ± 0.25 ± 0.29 ± 0.41 ± 0.41 ± 0.39 ± 0.43 ± (Vitamin B2) 0.02a,b 0.00b 0.01a,b 0.02a,b 0.02a,b 0.02a,b 0.02a,b 0.11a,b 0.01a,b 0.02a,b 0.04a (ug/g) Pantothenic acid 2.05 ± 0.61 ± 0.54 ± 1.38 ± 2.98 ± 0.04b 1.15 ± 1.12 ± 1.45 ± 4.40 ± 1.49 ± 1.28 ± 1.66 ± (Vitamin B5) 0.08c 0.00h 0.04h 0.06d,e, 0.01f,g 0.02g 0.07f,g 0.12a 0.01d,e 0.01e,f,g 0.08d (ug/g) f,g Pyridoxine 0.95 ± 0.70 ± 0.82 ± 1.22 ± 0.81 ± 0.02e 0.96 ± 0.95 ± 1.29 ± 1.01 ± 1.55 ± 1.60 ± 1.73 ± (Vitamin B6) 0.04d,e 0.01e 0.00e 0.03c,d 0.02d,e 0.02d,e 0.04b,c 0.08c,d,e 0.04a,b 0.09a,b 0.15a (ug/g) Values are expressed as the means of (3) replicates for total carbohydrate, protein, ash, energy, fat, and minerals two (2) replicates for fiber and vitamins ± standard deviation; Different lowercase letters between rows denote a significant difference (P < 0.05). contain higher levels of unsaturated and polyunsaturated FAs such as (PC2, Fig. 2b). Cookies made with jaggery had more ash content oleic, myristic, linoleic, linolenic, and gadoleic acid (shown in Fig. 5a), (1.80–2.20 g/100g) than those made with brown sugar (1.19–1.44 g/ compared to RWF and RWFbased cookies (Supplementary Table 1). 100g) and white sugar (1.19–1.44 g/100g). Similar results were Unsaturated and polyunsaturated FAs are crucial in reducing cholesterol observed by Lamdande et al. (2018) when substituting jaggery for sugar levels which provides various health benefits (Ciccoritti et al., 2017; Joo in a muffin formulation. Previously, it was reported that white sugar has & Choi, 2012; Kasote et al., 2021; Ruan et al., 2015). Among the fatty an ash value of approximately 0.015% (McKee et al., 2015), while acids profiled across samples, oleic acid is particularly abundant in brown sugar and jaggery have values of approximately 0.20% and WGRF and its cookies (Fig. 5d). This is particularly the case in KWGRF. 1.56%, respectively (Lamdande et al., 2018). The ash content reflects Several studies suggest that oleic acid is the most prevalent fatty acid in the mineral, fiber, and inorganics remaining in the sample after it has rice bran (Joo & Choi, 2012; Ruan et al., 2015). been heated to a very high temperature, eradicating moisture, volatiles, Ash content across flour samples ranged from 0.64 to 1.90 g/100g. and organics compounds (Altındağ et al., 2015; Islam et al., 2012). Similar trends were observed among the cookies. The ash content con- tributes to the distinction of BWGRFJ cookies from other cookie samples 7 H.B. Itagi et al. L W T 186 (2023) 115245 3.2.2. Micronutrient content B1 level (0.71 μg/g), among cookie samples examined in the present ICP-MS data of flour samples revealed varying mineral concentra- study. These results are predominantly attributed to the presence of bran tions between samples (Table 2). KWGRF flour samples had the highest components in both WGRFs, as these components are a known abundant Mg content (1692 mg/kg), followed by BWGRF (1491 mg/kg) and RWF source of B-vitamins, which provide a variety of health benefits (Kasote (428.1 mg/kg). The K, Fe, and Zn values in BWGRF were substantially et al., 2021; Tiozon et al., 2021). higher than those in the other two varieties of flour. Interestingly in the baked cookies of BWGRF made with white and brown sugar, the Mg and 3.3. Antioxidant-based nutraceutical properties of flour and cookie K levels were reduced, and substituting with jaggery maintained the Mg samples and K levels to the same basal level as in the flour. The mineral content of jaggery-sweetened cookies was found to be generally higher than that Table 3 presents the levels of γ-oryzanol, phenolics, and total anti- of white and brown sugar. For instance, KWGRF cookies with jaggery oxidant capacities in flour and cookie samples. In general, our findings offer the highest % daily value for Mn per 32g serving (26.99%, KWGRF concur with previous research indicating that whole grain rice is an with jaggery). In PC1, Fe distinguishes KWGRF jaggery-formulated abundant source of bioactive compounds. Furthermore, pigmented cookies from the other formulations (Fig. 2a). KWGRF jaggery- whole grain rice is far superior to non-pigmented rice with enriched formulated cookies could provide a 6.26% daily value of Fe per 32g nutraceutical properties (Goufo & Trindade, 2014; Itagi et al., 2023; serving. The enhanced mineral content in KWGRF and BWGRF cookies Kasote et al., 2021; Tiozon et al., 2021). BWGRF (37.56mg/100 g rice could therefore be attributed to the application of mineral rich WGRFs flour) had 4-folds and 1.7-folds more γ-oryzanol than KWGRF and RWF, and the inclusion of jaggery in the formulation. During dough prepa- respectively. γ-oryzanol distinguishes jaggery-sweetened BWGRF ration and baking, jaggery crystals dissolve as a result of their interac- cookies (48.82 mg/100 g rice flour) from the other formulations (PC2, tion with water molecules. This may have caused micronutrients from Fig. 2b). These BWGRF cookies had 35% and 105% γ-oryzanol than jaggery crystals to migrate throughout the cookie matrix resulting in KWGRFJ and RWFJ cookies. Previous reports have shown that γ-ory- mineral-rich cookies (Verma et al., 2019). The presence of vitamin and zanol concentrates on the rice bran (Goufo & Trindade, 2014; Kumari mineral-rich rice bran in pigmented varieties as compared to et al., 2015). The presence of bran in both WGRFs may account for the non-pigmented varieties also likely accounted for these results (Ciccor- high oryzanol content of the resulting cookies. Recent research evidence itti et al., 2017; Oppong et al., 2021). Lamdande et al. (2018) made suggests that γ-oryzanol may alleviate obesity and cognitive impair- comparable findings on muffins formulated with jaggery. In many re- ment, highlighting its importance to bioactive rice research (Mastinu gions of Asia and Africa, jaggery has a well-established reputation as a et al., 2019; Masuzaki et al., 2019). nutraceutical due to its abundance of essential amino acids, antioxi- There was a general decline in the levels of total phenolic compounds dants, phenolics, minerals such as Mg, K, Fe, Zn, and Cu, and vitamins. (TPC) upon baking. Be that as it may, in comparison to RWC cookies This nutrient-dense profile made jaggery a suitable substitute for white with added refined white sugar, the cookies made from KWGRWS have and brown sugar (Lamdande et al., 2018; Rao & Singh, 2022). 2-folds higher phenolics and BWGRWS cookies have 5-folds higher The WGRF cookies, regardless of the sweetener used, generally phenolics content (Table 3). Interestingly, the KWGR and BWGR retained the highest levels of B-vitamin (B1, B2, B5, and B6) than RWF- formulated with jaggery not only retained the highest levels of pheno- based cookies (Table 2). The vitamin B5 content of cookie samples lics, but also flavonoids, and anthocyanins (Table 3; Fig. 3b). The TPC ranged from 1.66 to 0.54 μg/g, with the BWGRFJ cookies containing the value of BWGRFJ cookies (178.1 mg GAE/100g) was 1.5 times greater most vitamin B5 and RWFBS cookies containing the least. BWGRFJ than in KWGRFJ cookies, 6 times greater than in RWFJ, and 8-times cookies also retained the highest vitamin B2 (1.43 μg/g) and vitamin B6 greater than that of RWFWS cookies, which is the formulation with (1.73 μg/g), while BWGRFWS and KWGRFWS had the highest vitamin the lowest TPC observation. The total flavonoid content (TFC) of the Table 3 Nutraceutical properties of refined wheat flour, wholegrain rice flour samples, and resulting cookies with different sweeteners. Samples/ Refined Refined Wheat Flour Cashew Kalanamak Kalanamak Whole Grain Rice Black Black Whole Grain Rice Cashew Parameter Wheat Cookies Whole grain Cashew Cookies Whole Cookies Flour Rice Flour grain Rice White Brown Jaggery White Brown Jaggery Flour White Brown Jaggery Sugar Sugar Sugar Sugar Sugar Sugar Total oryzanol 10.14 ± 11.65 13.26 15.27 ± 21.49 ± 0.43g 32.80 31.62 40.81 ± 34.31 ± 37.56 37.78 48.82 ± (mg/100g flour) 0.37k ± 0.29j ± 0.15i 0.20h ± 0.31e ± 0.23f 0.36b 0.28d ± 0.15c ± 0.26c 0.79a Total phenolics 18.76 ± 22.57 27.07 31.80 ± 411.0 ± 8.47a 44.61 47.28 119.0 ± 632.6 ± 103.3 166.4 178.1 ± (mg GAE/100g 1.22d ± 0.57d ± 0.47d 0.37d ± 0.49c ± 0.71c 1.53 16.20a ± 0.84b ± 1.27a 1.84a flour) Total flavonoids 26.17 ± 11.80 9.75 ± 41.31 ± 166.3 ± 2.74c 88.52 118.0 169.3 ± 271.7 ± 116.5 98.01 222.7 ± (mg CE/100g 1.66g ± 1.81h 1.92g ± 1.66f ± 1.31d 5.14c 10.90a ± 2.83d ± 4.28e 3.10b flour) 1.92g,h Total anthocyanin ND ND ND 33.40 ± 239.6 ± 116.9 139.2 144.7 ± 634.6 ± 356.2 339.5 434.2 ± (mg C-3-GE/ 1.67f 25.51d ± ± 9.64e 25.51e 33.40a ± 9.64c ± 16.70b 100g flour) 16.70e 34.76c Total Antioxidant 18.76 ± 5.80 ± 4.96 ± 32.11 ± 164.4 ± 3.39b 44.61 47.28 54.60 ± 289.4 ± 34.62 29.13 77.40 ± Capacity (mg 1.22f,g 0.57g 0.75g 1.31e,f ± 0.49d ± 0.71d 0.47d 16.20a ± ± 1.27f 5.64c QE/100g flour) 0.84d,e HAS (mg QE/100g 0.5 ± 0.1f ND ND 25.83 ± 201.0 ± 1.43b 14.29 14.75 21.91 ± 275.1 ± 24.73 23.17 32.85 ± flour) 0.73d ± 0.25f ± 0.19f 0.19e 1.64a ± 0.42d ± 0.42d 0.62c DPPH radical 16.96 ± 14.78 30.80 26.30 ± 43.62 ± 1.61d 35.87 17.61 40.36 ± 79.06 ± 45.36 50.58 59.57 ± scavenging 1.16g ± 1.55g ± 0.45f 0.35f ± 0.47e ± 3.12g 0.51d,e 0.37a ± 0.80d ± 1.21c 1.34b activity (%) FRAP (mmol/L 0.61 ± 0.28 ± 0.13 ± 0.49 ± 1.75 ± 0.02d 0.80 ± 0.83 ± 1.05 ± 8.56 ± 2.87 ± 3.10 ± 6.17 ± TE/g flour) 0.01f 0.02g 0.01g 0.07f,g 0.01f 0.01e,f 0.05e 0.23a 0.04c 0.02c 0.11b Abbreviations: C-3-GE–Cyanidin-3-Glucoside Equivalents; CE–Catechin Equivalents; GAE–Gallic Acid Equivalents; HAS–Hydrogen Peroxide Scavenging Activity; QE–Quercetin Equivalents TE– Trolox Equivalents; ND–Not detected; QE–Quercetin Equivalents; Values are mean ± standard deviation of three independent de- terminations (n = 3); Different lowercase letters between rows denote a significant difference (P < 0.05). 8 H.B. Itagi et al. L W T 186 (2023) 115245 Fig. 3. Heatmaps of the physical attributes and macromolecules (a) and antioxidant components and capacity (b) of RWF and WGRF and resulting cookies with different sweeteners. Abbreviations: RWF–Refined wheat flour; WGRF–Whole grain rice flour; RWFWS (Refined wheat flour white sugar); RWFBS (Refined wheat flour brown sugar); RWFJ (Refined wheat flour jaggery); KWGRF (Kalanamak whole grain rice flour); KWGRFWS (Kalanamak whole grain rice flour white sugar); KWGRFBS (Kalanamak whole grain rice flour brown sugar); KWGRFJ (Kalanamak whole grain rice flour jaggery); BWGRF (Black whole grain rice flour); BWGRFWS (Black whole grain rice flour white sugar); BWGRFBS (Black whole grain rice flour brown sugar); BWGRFJ (Black whole grain rice flour jaggery). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.) Fig. 4. Correlation plots for antioxidant components and capacity (a) and physical attributes and macromolecules (b) of RWF and WGRF and resulting cookies with different sweeteners. The p-values were signified as follows: ** = P < 0.05, ***P < 0.001. Abbreviations: RWF–Refined wheat flour; WGRF–Whole grain rice flour; RWFWS (Refined wheat flour white sugar); RWFBS (Refined wheat flour brown sugar); RWFJ (Refined wheat flour jaggery); KWGRF (Kalanamak whole grain rice flour); KWGRFWS (Kalanamak whole grain rice flour white sugar); KWGRFBS (Kalanamak whole grain rice flour brown sugar); KWGRFJ (Kalanamak whole grain rice flour jaggery); BWGRF (Black whole grain rice flour); BWGRFWS (Black whole grain rice flour white sugar); BWGRFBS (Black whole grain rice flour brown sugar); BWGRFJ (Black whole grain rice flour jaggery). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.) cookie samples ranged from 9.75 to 222.7 mg CE/100g, with BWGRFJ scavenging activity (HAS), DPPH radical scavenging activity, and cookies having the highest values and RWFBS cookies having the lowest. FRAP), phytochemical rich BWGRF consistently demonstrated the BWGRF-based cookies had a higher total anthocyanin content (TAC) highest values (289.4 mg QE/100g, total antioxidant capacity; 275.1 mg than cookie samples derived from other flours. In addition, the TAC QE/100g, HAS; 79.06%, DPPH; 8.56 mM TE/g, FRAP). A study con- values of BWGRFJ cookies (434.2 mg C-3-GE/100g) were 13-folds ducted by Iqbal et al. (2017) on the antioxidant content and capacity of greater than those of RWFJ cookies, which had the lowest TAC results. raw and processed sugars showed that jaggery had 118 and 138 times Antioxidants derived from pigmented rice protect vital lipids, pro- more phenolic compounds than brown and white sugar, respectively. teins, and DNA from oxidative stress. Therefore, the potential of the The DPPH radical scavenging and reducing power of the samples fol- antioxidants to combat free radicals was assessed through in vitro tech- lowed a similar pattern. Retaining higher levels of TPC, TFC and TAC niques (Goufo & Trindade, 2014; Tiozon et al., 2021). Across antioxi- correlated strongly with antioxidant properties (Fig. 4a). This is further dant capacity assays (total antioxidant capacity, hydrogen peroxide supported by the strong positive correlation between bioactive 9 H.B. Itagi et al. L W T 186 (2023) 115245 Fig. 5. Fatty acid profile (a), heatmap (b), correlation plot (c), and level of oleic acid in RWF and WGRF and resulting cookies with different sweeteners. Abbreviations: RWF–Refined wheat flour; WGRF–Whole grain rice flour; RWFWS (Refined wheat flour white sugar); RWFBS (Refined wheat flour brown sugar); RWFJ (Refined wheat flour jaggery); KWGRF (Kalanamak whole grain rice flour); KWGRFWS (Kalanamak whole grain rice flour white sugar); KWGRFBS (Kalanamak whole grain rice flour brown sugar); KWGRFJ (Kalanamak whole grain rice flour jaggery); BWGRF (Black whole grain rice flour); BWGRFWS (Black whole grain rice flour white sugar); BWGRFBS (Black whole grain rice flour brown sugar); BWGRFJ (Black whole grain rice flour jaggery). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.) compounds and antioxidant capacity, especially hesperidin, rutin, ella- gluten-containing products. However, our results suggest that KWGRF gic acid, and quercetin with FRAP and DPPH (Fig. 4a). Heating starch in and BWGRF cookies, particularly those made with white sugar and water causes the granules to rupture. Phenolic and bioactive compounds jaggery, could compete with RWF-based cookies and thus may be more in the matrix could complex with starch molecules either through in- preferred by consumers. clusion (where the compounds are captured within the starch helices), or non-inclusion (where the compounds are trapped between the heli- 4. Conclusions ces) (Sudlapa & Suwannaporn, 2023). This complexation process occurred during baking. These results highlight the potential of formu- The whole grain cookies made from GI-tagged rice landraces, Kala- lating nutritionally superior GF-products from whole grain rice and namak and Chak-hao, offer distinct sensory qualities and are nutritious nutrient-dense sweetener such as jaggery. that are rich in polyunsaturated fatty acids, minerals, fiber, and bioac- tive compounds. The Chak-haobased cookies retained the highest levels 3.4. Sensory analysis of cookie samples of phytochemicals with greater antioxidant activities and adding jaggery as sugar alternative exhibited higher levels of Fe and helped to retain The sensory properties of cookies produced using WGRF and RWF higher antioxidant compounds upon baking. These cookies demonstrate with three different sweeteners (white sugar, brown sugar, and jaggery) good shelf-life stability with aw levels under 0.85. Although gluten-free were evaluated, and the results are presented in Fig. 6. KWGRF cookies formulations spread less than the wheat control, sensory evaluation sweetened with white sugar were rated the highest in all sensory suggests that acceptability of KWGRF and BWGRF cookies are compa- properties (8.00) except for crumb color (Fig. 6c). The panelists rable to RWFbased cookies. The rising demand for nutritious foods preferred the crumb color of BWGRF cookies with jaggery (8.00), fol- provides manufacturers an opportunity to diversify their products to lowed by RWF with white sugar (7.85). In terms of surface color (8.00) cater to specific markets. Future research can explore the development and cracking patterns (8.00), BWGRF cookies formulated with jaggery of more gluten-free functional foods using Kalanamak and Chak-hao were a close second to KWGRF cookies with white sugar. For aroma, the rice, catering to both local and global demands. Investigating pack- panelists showed the highest preference for KWGRF cookies with aging and storage options is also essential for maintaining shelf stability jaggery (7.89). The preference for mouthfeel in KWGRF and RWF and nutritional quality. cookies (both versions of sweetened with white sugar), could be due to a clean mouthfeel without any residue formation, attributed to lower TDF Informed consent declaration compared to BWGRF cookies. Similar findings were reported by Baumgartner et al. (2018) on cookies made with dephytinized oat flour. All panelists granted informed consent before taking part in the Yildiz and Gocmen (2021) argued that gluten-free bakery products have study. The research protocol was explained to the panelists, detailing the weaker sensory properties and may not meet consumer expectations due cookies and their ingredients. Panelists could opt out of evaluation to their harder structure, darker color, unpleasant appearance, and sessions without needing to explain their choice. Evaluations were dry-sandy feeling in the mouth compared to conventional conducted at the Center of Excellence in Rice Value Addition, Product 10 H.B. Itagi et al. L W T 186 (2023) 115245 Fig. 6. Sensory profile (a), dot plots (b), spider plots (c), and normalized scores in RWF and WGRF and resulting cookies with different sweeteners. Abbreviations: RWF–Refined wheat flour; WGRF–Whole grain rice flour; RWFWS (Refined wheat flour white sugar); RWFBS (Refined wheat flour brown sugar); RWFJ (Refined wheat flour jaggery); KWGRF (Kalanamak whole grain rice flour); KWGRFWS (Kalanamak whole grain rice flour white sugar); KWGRFBS (Kalanamak whole grain rice flour brown sugar); KWGRFJ (Kalanamak whole grain rice flour jaggery); BWGRF (Black whole grain rice flour); BWGRFWS (Black whole grain rice flour white sugar); BWGRFBS (Black whole grain rice flour brown sugar); BWGRFJ (Black whole grain rice flour jaggery). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.) Development, and Sensory Laboratory, located at the IRRI South Asia Acknowledgments Regional Center in Varanasi, Uttar Pradesh, India. Cookies suitable for consumption were included in the study. The samples of rice were provided by Vivek Kumar Singh and Ajeet Kumar, while Hari Narain Singh and Santhosh Kumar helped with the Funding experiments. The authors are appreciative of their help. The Department of Agriculture and Farmers Welfare (DA&FW) Appendix A. Supplementary data Government of India, the CGIAR Research Initiative on Transforming Agrifood Systems in South Asia (TAFSSA), and the Agricultural and Supplementary data to this article can be found online at https://doi. Processed Food Products Export Development Authority (APEDA) pro- org/10.1016/j.lwt.2023.115245. vided funding for this study. References CRediT authorship contribution statement Aguiar, E. V., Santos, F. G., Krupa-Kozak, U., & Capriles, V. D. (2023). Nutritional facts Hameeda Banu Itagi: Conceptualization, Investigation, Resources, regarding commercially available gluten-free bread worldwide: Recent advances and future challenges. Critical Reviews in Food Science and Nutrition, 63(5), 693–705. 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