INTRODUCTIONBiscuit is a popular, versatile bakery item consumedby all levels of society worldwide due to its taste,affordability, convenience, and an extended shelf life [1].Biscuits, or cookies, are usually low in fiber, vitaminsor minerals, and are highly calorific [2]. Thus, theycannot be added to the group of healthy foods. However,consumers are seriously concerned about their healthissues. For this reason, bakery products with a high fibercontent may be a choice for health-conscious people.Biscuits have good consumer acceptance, just as snacksdo, and a long shelf life. Therefore, there is scope fornutritional development and fortification [3].Okara is a pulpy, fiber-rich by-product of tofuand soy milk processing. Soy okara is a rich source offiber [4]. It is composed of cellulose, hemicellulose,and lignin, as well as protein, lipids, vitamins,phytochemicals, and phytosterols [5, 6]. According toGrizotto et al., one ton of processed soybeans couldproduce about two tons of okara following soymilkproduction [7]. As a result, every year enormousquantities of okara create disposal problems. To solvethem, okara may be used as a dietary additive. It canfurther be processed to convenient and useful formssuch as powders or extrudates [8]. It can be used directlyin soups or salads. In addition, we can now find manyonline recipes where freshly produced okara is used as araw ingredient.However, it is hard to find an industrial product ofsoy okara because of its high moisture content (around85 g) and poor textural quality that may cause rapiddeterioration [5]. Some possible applications of okara might include baked goods, beef patties, and coconutcookies due to its high amount of fiber and protein [8, 9].Dietary fiber is a significant component in bakeryproducts, confectionery, meat, beverage, and dairyitems [8]. However, supplementing baked products withdietary fiber may change the flavor, texture, and taste offinal products [3].A number of studies have shown the applicationof okara flour in tortillas, cookies, roti and parata, andeven in breakfast cereals [8, 10–12]. The use of okara asa gluten free all-purpose flour may add further value tothis agro waste and bring significant nutritional benefits.Our study aimed to identify the effect of okara flouron the properties and nutritional composition of biscuits.STUDY OBJECTS AND METHODSSoybeans and refined wheat flour were collectedfrom K-R Market (Mymensingh, Bangladesh). Sodiumbi-carbonate (food grade) was supplied by MitaliScientific Co. Ltd., Bangladesh.Whole soybeans were soaked in a 0.5% NaHCO3solution (1:2) at 60°C for four hours in a water bath(Schufzart, Membart GmBH+ Co., Bϋchenbach,Germany). The water was discarded and the soakedbeans were dehulled before grinding to removeunwanted substances using a dehuller. The hydratedsoybeans were blanched at 90 ± 2°C for 10 min with theaddition of 0.5% NaHCO3 (w/v), and the solution wasdrained well. The beans were washed with potable waterfor three times [13]. The blanched beans were groundwith the addition of hot water (100°C) [bean to waterratio = 1:4] using a super mass collider (Masuko SangyoCo. Ltd., Kawaguchi, Japan).Soy okara was collected after soy milk extraction byfiltering through double layers of cheese cloth. Soy okarawas dried in a cabinet drier (Dayton Electric MFG. Co.Ltd., USA) at 60°C for 24 h and ground using a grinder.The ground okara was sieved (420-micron mesh size)and kept in a desiccator to reduce the moisture contentby up to 5% [11]. The powder was finally placed in asealed polyethylene-laminated aluminum foil bag andkept at –20°C before analysis and further processing.Moisture, protein, ash, and fat contents of soy okaraflour, refined wheat flour, and biscuits were determinedaccording to the AOAC method [14]. Genistein wasdetermined by the HPLC method as modified by [15].Particle size was measured according to [16].Average particle sizes (d3,2 – surface-weighted meandiameter, Sauter mean diameter and d4,3 – volumeweightedmean diameter, De Brouckere mean diameter)of refined wheat flour, as well as 2, 4, 6 and 8% soyokara flour were determined using a particle sizeanalyzer (Malvern Zetasizer Nano ZS, UK) with theattachment of dry feed.Refined wheat flour (RWF), okara flour, and otheringredients were weighed according to Table 1 andmixed together. Fat was mixed with the ingredients andwater was added to the mixer to form dough. The doughwas kneaded, rolled to uniform thickness (5 mm) andcut in round shape biscuits of 4 cm in diameter. Theywere baked at 220°C for 10 min and cooled at roomtemperature. The biscuits were packed in HDPE andkept in desiccators for further analysis.The spread ratio, an essential quality parameter ofbiscuits, was determined as follows:Spread ratio = D / Twhere D is the average diameter and T is the averagethickness of biscuits after baking, cm.Weight loss (WL) of biscuits during baking wascalculated by the following formula [17]:WL = (Wdough – Wbiscuit) / Wdough × 100where Wdough is the weight before baking and Wbiscuit isthe weight after baking five samples, g.The doughs made from different amounts of soyokara flour (2, 4, 6, and 8%) and only refined wheat flour(control) were tested for firmness by a penetration test.The dough was placed in a concentric cylinder (30 mmin diameter) under a cylindrical probe (5 mm) (StableMicro Systems, UK). The test conditions included2 mm/s pretest speed, 3 mm/s test speed, 10 mm/s posttestspeed, 50 kg load, and 60% strain. When the probepenetrated 60% of the dough, it was found to gain itsoriginal position. The absolute peak force of the forcetimecurve was taken as dough firmness [18]. Eachdough was tested three times.Dough strength, adhesion, and stickiness tests werecarried out using an SMS/Chen-Hosney Stickiness Celland Prespex cylinder probe (25 mm) (Stable MicroSystems, UK). The test conditions included 2 mm/spretest speed, 2 mm/s test speed, 10 mm/s post-testspeed, 40 g trigger force, 3 mm return distance, and 10 scontact time [19–21]. The positive peak constraint fromthe curve was considered as stickiness force. The areafalling under this force-distance curve indicates thework of adhesion. The distance of sample extensionTable 1 Basic formulation for preparation of biscuits (on 100 gflour basis)Ingredients, g SamplesControl(wheat flour)Experimental(with okara flour)Wheat flour 100 98 96 94 92Okara flour – 2 4 6 8Sugar 50 50 50 50 50Oil 40 40 40 40 40Baking powder 1.5 1.5 1.5 1.5 1.5Milk powder 5 5 5 5 5Salt 0.5 0.5 0.5 0.5 0.5Egg 45 45 45 45 45Ammonium bicarbonate 0.5 0.5 0.5 0.5 0.5424Momin M.A. et al. Foods and Raw Materials, 2020, vol. 8, no. 2, pp. Х–Хduring prove return was considered as dough strength orcohesiveness [18].The three-point bending test was carried outemploying a 3-point bending rig (Stable Micro Systems,UK) connected to a texture analyzer. The test conditionsincluded 10 mm/s pretest speed, 1 mm/s test speed,10 mm/s post-test speed, 10 mm distance, and 50 kgload cell; descending development was continued tillthe biscuits broke. The most extreme constraint wasrecorded as the “hardness” of the biscuits [22].The cutting strength of biscuits was measured usingan HDP/BS blade-type texture analyzer (Stable MicroSystems, UK). The biscuits were set on the platform,and the blade was connected to the crosshead of theinstruments. The test conditions included 2 mm/s pretestspeed, 2 mm/s test speed, 10 mm/s post-test speed, and5 mm distance. The outright peak force of the curve wasrecognized as the cutting strength of the biscuits [18,21]. Textural properties of the dough and biscuits weredetermined by a TA-XT plus texture analyzer (StableMicro Systems, UK) with Texture ExpertTM software.The color of biscuits was analyzed by a colorimeter(Chroma Meter CR400, Konica Minolta, Japan) underilluminant: *C, D65 and space: LAB. It was determinedin L*, a* and b* system, where L* is lightness (100:white, 0: black), a* is redness (+)/greenness (−), and b*indicates yellowness (+)/blueness (−). All analyses wereperformed in triplicate.The sensory evaluation of the control andexperimental samples included color, texture, flavor,and overall acceptability by ten semi-trained panelistson a 9-point hedonic scale (9 = like extremely,8 = like very much, 7= like moderately, 6 = like slightly,5 = neither like nor dislike, 4 = dislike slightly,3 = d islike m oderately, 2 = d islike v ery m uch a nd1 = dislike extremely). The results were evaluated byanalyses of variance (ANOVA) and Duncan’s newmultiple range test (DMRT) of the Statistical AnalysisSystem (SAS).The physicochemical, nutritional, and texturalproperties were determined in replicate and statisticallyanalyzed by a two-way ANOVA using the MicrosoftExcell-2010.RESULTS AND DISCUSSIONTable 2 shows the nutritional composition ofokara and refined wheat flour (RWF). Okara flour hadsignificantly higher contents of protein, fat, ash, andcrude fiber compared to wheat flour. However, whenmaking biscuits, it is important to add wheat flour asit contains gluten, which makes the dough adhesiveand cohesive. Yet, gluten is also responsible for celiacdiseases [23]. In our study, we substituted 2, 4, 6, and8% of wheat flour with the same quantities of gluten-freeokara flour.Figure 1 illustrates the distribution of particle sizesin the mixed flour (2, 4, 6, and 8% okara flour mixedwith wheat flour) and control flour (RWF). We identifiedtwo distinct peaks for all mixed flour samples, whereasthe first peak in the control was not as distinct, showingthat the particle size distribution of all flours wasbimodal. The results were in agreement with [16] and[24]. We also found the maximum particle size to be ~100 μm, indicating a higher volume. The particle sizesfor both the experimental and control samples rangedfrom 0.1 to 100 μm, but the volume of particles was thehighest at ~ 145 μm and 91 μm, respectively. The resultsindicated that refined wheat flour had lower proteinand fiber contents [25]. Hard dough prepared with finerparticle-size flour has higher density, resulting in lessdeveloped biscuits during baking [26]. Thus, coarser,mixed or composite, flour is most desirable for preparinghard dough biscuits.The mean volume diameter and the surface meandiameter of refined wheat flour (70.05 and 27.5 μm,respectively) were significantly (P < 0.05) smaller thanthose of all mixed flour samples (83.7 and 32.8 μm,respectively). There was no significant difference among2, 4, 6, and 8% okara flour samples (P > 0.05). The flourparticle size often affects the biscuits’ water absorptioncapacity, density, and spread ratio. When fine particle-Table 2 Chemical composition of okara and refined wheatflour (RWF)Components Okara flour Refined wheat flourMoisture, % 13.75a ± 1.25 12.65a ± 1.50Protein, % 35.14a ± 2.00 13.00b ± 1.90Fat, % 10.78a ± 1.40 1.80b ± 1.00Ash, % 3.95a ± 0.50 1.47b ± 0.25Crude fiber, % 30.01a ± 2.25 3.23b ± 0.50Genistein, mg/100 gof okara flour5.05 ± 0.50 ndMean ± SD represents the average of three replicates for each analysisDifferent letters in the same row show significant differences(P < 0.05)nd = not determinedFigure 1 Particle size of refined wheat flour and wheat flourmixed with okara flourwheat flour with 2% okara flourwheat flour with 4% okara flourwheat flour with 6% okara flourwheat flour with 8% okara flourwheat flour only425Momin M.A. et al. Foods and Raw Materials, 2020, vol. 8, no. 2, pp. Х–Хsize flour is used for hard dough biscuits, it usuallyresults in higher density and less effective bakingproperties [26]. Therefore, coarser composite flour ismore desirable.The weight of okara flour biscuits was higher thanthat of biscuits prepared with only refined wheat flour(control). The biscuits prepared with coarser 2, 4, 6,and 8% okara flour mixes had a significantly (P < 0.05)higher thickness and a smaller diameter than the control(RWF) biscuits (Table 3) due to higher developmentduring baking [26]. As a result, they had a reducedspread ratio compared to the control. The spread ratiocorrelates with texture, grain fineness, bite, and overallmouth feel of the biscuits [27]. Additionally, using flourhigh in protein and fiber in place of wheat flour providesa reduced spread ratio [28].Among the experimental biscuits, 8% okarasamples had a smaller diameter and a lower spread ratiocompared to the others (Table 3). This might be due tocomparatively higher protein and fiber contents in 8%okara biscuits. Further, mixed flour biscuits showed asignificant (P < 0.05) reduction in weight loss comparedto biscuits from wheat flour (Table 3). However, therewas no significant difference (P > 0.05) in weight lossamong 2, 4, 6, and 8% okara flour biscuits. The lowerweight loss in the experimental biscuits might be dueto better water absorption by the flour components dueto protein hydration, with less water evaporated duringbaking [17].Moisture, ash, protein, and fat contents in the biscuitsprepared with okara flour were higher than those in thebiscuits from control flour (RWF) (Fig. 2). Among mixedflours, 8% okara flour provided significantly (P ≤ 0.0.5)higher nutrient contents compared to the others. Thebiscuits made with 8% okara flour had a higher moisturecontent than the control or the other mixed-flour samplesdue to a greater volume of water required at the time ofdough making for its high fiber and protein contents.Table 3 Effect of okara flour on physical properties of biscuitsBiscuits Weight, g Diameter (D), cm Thickness (T), cm Spread ratio (D/T) Weight loss, %Control (refined wheat flour) 4.95b ± 0.40 4.62a ± 0.32 0.62e ± 0.06 7.45a ± 0.80 11.46a ± 0.352% okara flour 5.94a ± 0.50 4.58b ± 0.50 0.63d ± 0.05 7.27b ± 0.70 10.01b ± 0.554% okara flour 5.95a ± 0.35 4.56c ± 0.42 0.64c ± 0.02 7.13c ± 0.50 10.03b ± 0.506% okara flour 5.94a ± 0.70 4.55c ± 0.45 0.65b ± 0.01 7.00d ± 0.90 10.01b ± 0.658% okara flour 5.93a ± 0.55 4.53d ± 0.60 0.66a ± 0.01 6.86e ± 0.80 10.09b ± 0.75Mean ± SD represents the average of five replicates for each analysisDifferent letters in the same column indicate significant difference (P < 0.05)Figure 2 Nutritional components of biscuits from wheat flourand okara flour-enriched biscuits.(wheat flour)Table 4 Textural properties of dough and biscuits from refined wheat flour and okara flourSamples Dough BiscuitsFirmness, N Stickiness, N Work of adhesion(N-S)×10-3Doughstrength, mmCuttingstrength, NHardness, NControl (refined wheat flour) 4.45a ± 0.05 0.28a ± 0.05 5.20a ± 0.5 0.65a ± 0.10 64.94a ± 2.00 15.20c ± 19.052% okara flour 3.90b ± 0.09 0.21b ± 0.07 4.75b ± 0.3 0.54b ± 0.09 55.75b ± 1.75 17.55b ± 1.554% okara flour 3.79c ± 0.07 0.18b ± 0.05 4.50c ± 0.4 0.50b ± 0.01 52.55b ± 1.55 18.10b ± 1.256% okara flour 3.66cd ± 0.05 0.16b ± 0.03 4.25c ± 0.1 0.40c ± 0.03 43.90c ± 2.50 18.76a ± 2.008% okara flour 3.45d ± 0.04 0.14b ± 0.05 3.50d ± 0.7 0.35c ± 0.05 40.35cd ± 2.75 19.05a ± 2.05Mean ± SD represents the average of five replicates for each analysisDifferent letters in the same column indicate significant difference (P < 0.05)Figure 3 Color parameters of biscuits from wheat flourand okara flour-enriched biscuitsL* a* b*(wheat flour)426Momin M.A. et al. Foods and Raw Materials, 2020, vol. 8, no. 2, pp. Х–ХFlours rich in protein require much water to makemachinable dough as protein is not sufficiently hydratedto form a network [29].Dough firmness, strength (cohesiveness), andstickiness were significantly lower in 2, 4, 6, and 8%okara flour samples compared to the control (wheatflour only) (Table 4). Decreased dough firmness isusually related to a high fat content in formulations.This disrupts the development of a gluten network bylubricating the complete matrix and making it hydrated[17, 30]. Okara flour is gluten-free, which alsoaccounted for lower firmness in mixed flour doughcompared to dough made from wheat flour only.Stickiness is a significant parameter of dough qualityas it affects the handling convenience and may damagethe apparatus [31]. Okara flour dough showed lowerstickiness compared to the control, owing to a higherwater absorption ability. Lower stickiness and adhesionof okara dough correlate with a greater water absorptioncapacity, a comparatively low gluten content, and ahigher fat content [16].The three-point bending test showed significantlylower (P ≤ 0.05) hardness for 2, 4, 6, and 8% okaraflour biscuits compared to wheat biscuits (Table 4). Thiswas due to a high protein content and a better waterabsorption capacity in the mixed flour. 8% okara flourbiscuits seemed harder than those with 2, 4, and 6%okara flour due to a lower gluten content compared to thecontrol and the other mixed flours.There was a significant (P < 0.05) difference for allfive types of biscuits in terms of color (Fig. 3). The L*value of all composite flour biscuits was lower than thatin the refined wheat flour biscuits due to the presenceof natural anti-browning substance such as genisteinin okara flour [32]. In particular, its genistein contentwas 5.05 ± 0.5 mg/100 g of okara flour (Table 2). 8%okara flour had the highest L* value, indicating lessbrown pigment formation. The result suggested thatokara flour could reduce brown pigments. We observedhigh positive a* values (redness) for 2, 4, 6 and 8%okara flour biscuits. Positive b* values (yellowness)were significantly (P < 0.05) higher in mixed flourbiscuits compared to the control due to the presence ofphytochemicals and crude fiber in okara flour [11].The results of sensory evaluation of biscuits enrichedwith okara flour and control biscuits are shown inTable 5. The sample with 4% okara flour showed thefinest sensory characteristics in terms of color, texture,flavor, and overall acceptability. However, the othersamples were also found acceptable. A DMRT analysisrevealed that 4% okara biscuits were significantlybetter in color, texture, flavor, and overall acceptabilitythan other biscuits containing 2, 6, and 8% okara flour.However, increasing the amount of okara flour decreasedthe level of overall acceptability.CONCLUSIONBiscuits prepared from mixed okara (2, 4, 6, and 8%)and refined wheat flour were found to outperform refinedwheat flour biscuits in physicochemical, nutritional,textural, and sensory attributes. Okara flour biscuitshad an inferior spread ratio, but higher fiber and proteincontents. We also found them to have poor cuttingstrength and greater hardness. Okara flour biscuits hadbetter color due to the presence of genistein. 4% okarabiscuits had higher consumer acceptability on a 9-pointhedonic scale. Decreased dough hardness due to okaraflour diminished the cutting strength and increasedhardness in the corresponding biscuits. Thus, we canconclude that biscuits prepared from okara flour can beconsidered as a healthy snack option.CONTRIBUTIONSMd. A. R. Mazumder and Thottiam V. Ranganathanconceptualized and supervised the work. AnjumanA. Begum and Md. F. Jubayer were involved inmanuscript writing and data analysis. Md. A. Mominand Asmaul H. Nupur performed laboratory experimentsand data collection.CONFLICT OF INTERESTThe authors declare that there is no conflictof interest.