Devin Bates, Rossy Bueno & Tasha Perkins 11/18/2013

Effect of using hazelnut flour as a replacer of all-purpose flour on texture, appearance, and palatability of gluten-free cupcakes NUTR 45300 Research Project Devin Bates, Rossy Bueno & Tasha Perkins 11/18/2013

Hypothesis and Objectives Null hypothesis: The use of hazelnut flour to replace all-purpose flour will not have an effect on palatability, texture, and appearance of white cupcakes. The objectives of this experiment are to use hazelnut flour in place of all-purpose flour in the baking of white cupcakes, making the cupcakes safe for those with gluten intolerance without compromising quality or flavor, and add nutritional value to the original recipe of cupcakes with the incorporation of hazelnut flour, which is a good source of dietary fiber. Abstract We live in a world where more and more people are being affected by gluten. Gluten is a protein that is found in foods such as bread and pasta. Celiac disease presents an array of problems such as gastrointestinal disorders, acne and fatigue. These conditions can be cumbersome for millions of people in the American population to endure. This sets the foundation for this experiment in which hazelnut flour was tested to potentially replace all-purpose flour and obtain cupcakes for gluten-sensitive consumers. Two variations of hazelnut flour were made: Variation 1 combined 50% hazelnut flour with 50% all-purpose flour and Variation 2 contained 100% hazelnut flour. These variations were compared to a control group (composed of all-purpose flour) in terms of palatability, texture, and appearance. Objective evaluation of the cupcakes was conducted using the Texture Analyzer, Water Activity System, and Seed Volume Apparatus. It was apparent that the 100% hazelnut flour cupcakes were not conducive to baking. The subjective data results showed that there was no acceptability for the 100% hazelnut cupcakes. However a good number of panelists found the 50% hazelnut flour cupcakes acceptable based on taste, texture, and appearance. It was concluded that hazelnut flour has an effect on palatability, texture, and appearance of cupcakes. This experiment might serve as a starting point to find the right proportion of replacement of all-purpose flour to make cupcakes for gluten-sensitive individuals without sacrificing taste.

Introduction There is a growing concern for gluten sensitivity and an increasing popularity for gluten-free foods. Gluten is found in grains such as wheat, barley and rye. About 1% of American people have been diagnosed with celiac disease and 6% of Americans have non-celiac gluten sensitivity. Celiac disease is an autoimmune disease that causes damage to the villa of the small intestine when gluten is consumed. This can cause an interference with the absorption of nutrients, and therefore, may lead to malnourishment. The only treatment for celiac disease is a diet completely free of gluten. Those with non-celiac gluten sensitivity may be able to consume small quantities of gluten, but should decrease their gluten consumption significantly (National Foundation for Celiac Awareness, 2013). Going gluten-free may alleviate gastrointestinal discomfort or even eliminate discomfort if diagnosed with celiac disease or gluten-sensitivity. (Hafstrom et al, 2001). Studies have shown that the damage caused to the intestinal tract and the intestinal villa in patients with celiac disease may be reversed with a diet free of gluten. Once the intestinal tract heals, the problems with malabsorption will also disappear (Satumi et al, 2010). The purpose of this research experiment was to create a cupcake that decreased the amount of gluten and maintained a desirable texture, appearance, and palatability. For this experiment cupcakes made with all-purpose flour were replaced with hazelnut flour. Three variations of the cupcake recipe were created. The first variation was the original recipe made with all-purpose flour only. The second variation was made with 50% all-purpose flour and 50% hazelnut flour. The third variation was made with 100% hazelnut flour. For this research study the independent variable was the amount of hazelnut flour. The dependent variables were the texture, appearance and palatability of the cupcakes. The control was the original recipe made only with all-purpose flour.

Methods Overall design This experiment consisted on the preparation of cupcakes that involve a control group and two variations. The experiment was conducted in triplicate. Each trial was composed of three variables: a control group with 0% hazelnut flour, Variable 1 with 50% hazelnut flour, and Variable 2 with 100% hazelnut flour. The control group was prepared with all-purpose flour and contained no hazelnut flour as indicated by the percentage. Variable 1 was made with 50% hazelnut flour and 50% all-purpose flour. Variable 2 consisted solely of hazelnut flour. All variables were prepared using a recipe as reference that was modified accordingly based on flour amounts. The recipe and work plan used throughout is provided in the Specific Procedures section. The objective methods used were the Texture Analyzer, Seed Volume Apparatus, and Water Activity System to obtain direct texture measurement, water activity, and volume correspondingly. The cupcakes were evaluated subjectively by a taste panelist consisting of 8 students that filled out a sensory evaluation scorecard based on palatability, texture, and appearance. Specific Procedures The recipe used was obtained from http://allrecipes.com/recipe/simple-white-cake/detail.aspx. Recipe 1 (Control group: ID # 623) Original recipe for 12 cupcakes Ingredients 1 cup (200 g) granulated sugar ½ cup (113.5 g) butter 2 eggs 2 teaspoons (8.66 g) vanilla extract 1 ½ cups (187.5 g) all-purpose flour 1 ¾ teaspoons (8.050 g) baking powder ½ cup (123 g) milk 2% milk-fat Recipe 2 (Variable 1: 50% hazelnut flour, 50% all-purpose flour: ID # 982) Recipe for 12 cupcakes Ingredients 1 cup (200 g) granulated sugar ½ cup (113.5 g) butter 2 eggs 2 teaspoons (8.66 g) vanilla extract ¾ cups (93.75 g) hazelnut flour ¾ cups (93.75) g) all-purpose flour 1 ¾ teaspoons (8.050 g) baking powder ½ cup (123 g) milk 2% milk-fat

Recipe 3 (Variable 2: 100% hazelnut flour: ID # 456) Recipe for 12 cupcakes Ingredients 1 cup (200 g) granulated sugar ½ cup (113.5 g) butter 2 eggs 2 teaspoons (8.66 g) vanilla extract 1 ½ cups (187.5 g) hazelnut flour 1 ¾ teaspoons (8.050 g) baking powder ½ cup (123 g) milk 2% milk-fat The directions for all three recipes were the same and are provided in the list below. Directions 1. Preheat oven to 350 F (176.7 C). Add paper liners to muffin/cupcake pan. 2. In a medium bowl, combine sugar and butter until cream consistency is achieved. Beat in the eggs, one at a time, and stir the vanilla. 3. Combine flour and baking powder, add to creamed mixture and mix well. Add milk and stir until batter is smooth. 4. Pour or spoon batter into muffin pan. 5. Bake for 20 to 25 minutes. Check cupcakes by inserting toothpick in the center and ensuring it comes out clean before removing pan from oven. Equipment specifications In order to control the variables other than the ones that are being tested during the experiment it was necessary to be consistent and follow all the directions of the recipe carefully. Being familiar with the equipment that was used to obtain results based on texture and appearance of cupcakes was a key step for successful completion of the research experiment. Equipment specifications of each instrument (texture analyzer, seed volume apparatus, and water activity meter) were kept the same for the purpose of obtaining valid results during all three trials. The Stable MicroSystems Texture Analyzer with cone probe settings were the following: standard for muffin (load settings), parameters pre-test speed: 2.0 mm/s, test speed: 3.0 mm/s, post-test speed: 5.0 mm/s, rupture test distance: 1.00 mm, distance: 5.0 mm, force: 2000 g, time: 5.00 sec, count: 5. The CX-2 Aqualab Water Activity System was used to measure water activity by obtaining a quarter size amount of the cupcake sample and placing it in sample cup. The sample cup is then placed in sample drawer and the knob is turned from open/load to read. The stepby step procedure provided in The Food Chemistry Laboratory Manual- A Manual for Experimental Foods, Dietetics, and Food Scientists was followed to use the Seed Volume Apparatus (Weaver & Daniel, 2003). This manual was also consulted for full proper use of the other two instruments as well.

Work plan Set up utensils and ingredients. Preheat the oven to 350 F (176.7 C). Review the recipe and ensure that we are familiar with steps. Begin mixing the ingredients for the two variables and one control. Pour batter into pan and place in the oven. Set timer for 20 minutes. Monitor the baking process so that the cupcakes will not burn. Clean up our work area while cupcakes are baking. Check on the cupcakes. If done, take them out to cool. Prepare for subjective and objective method testing. Perform objective and subjective tests and record objective test results. Replications, randomization, and sampling As previously mentioned, the experiment was conducted in triplicate to obtain accurate results and eliminate chances of experimental error. Objective data was obtained for all variations during each trial. Subjective evaluation of samples was completed by eight panelists. In terms of randomization, the cupcakes were coded with a specific three code ID number for purpose of identification and to avoid any type of bias. The control group was referred as Sample 623, Variation 1 (50% hazelnut flour) was labeled as Sample 982, and Variation 2 (100% hazelnut flour) was named Sample 456. Each cupcake was cut into 4 pieces to provide samples for all panelists. The panelists filled out the sensory evaluation scorecard which consisted of a 9-point hedonic scale on palatability and two questions in terms of texture and appearance of the cupcakes. The two questions were based on a 3 point rating system in which 1 referred to best of the quality being measured and 3 constituted worst of the quality being measured. In other words, 1 was best texture and 3 was worst texture; 1 was best appearance and 3 worst appearance, corresponding to each question. The objective data was analyzed statistically using averages of results of all three trials and GraphPad software, Instat 3 through one way analysis of variance (ANOVA) and Tukey-Kramer Multiple Comparison Test to find significance between the variables tested at a P<0.05.

Sensory Evaluation Scorecard Overall, how much do you like or dislike the taste of the cupcake sample? Sample 982 Sample 456 Sample 623 Like Extremely Like Extremely Like Extremely Like Very Much Like Very Much Like Very Much Like Moderately Like Moderately Like Moderately Like Slightly Like Slightly Like Slightly Neither Like nor Dislike Neither Like nor Dislike Neither Like nor Dislike Dislike Slightly Dislike Slightly Dislike Slightly Dislike Moderately Dislike Moderately Dislike Moderately Dislike Very Much Dislike Very Much Dislike Very Much Dislike Extremely Dislike Extremely Dislike Extremely From 1 3 please rate the texture of the three samples, with 1 being best texture to 3 being worse texture. Sample 456 Sample 623 Sample 982 From 1 3 please rate the appearance of the three samples, with 1 being best appearance to 3 being worst appearance. Sample 456 Sample 623 Sample 982

Results Table 1.1 Force of control and hazelnut flour cupcakes using texture analyzer Control: no hazelnut flour (g) 50% hazelnut flour (g) 100% hazelnut flour (g) Trial 1 22.6 23.3 16.4 Trial 2 27.8 13.8 11.4 Trial 3 17.2 16.7 18.6 Average 22.5 a 17.9 a 15.5 a Standard deviation 5.30 4.87 3.69 Averages bearing same superscript are not statistically significant, P>0.05 Average force (g) from texture analyzer 30 25 20 15 10 5 0 Control 50% hazelnut flour 100% hazelnut flour Variations Figure 1.1 Average force of control and hazelnut flour cupcakes using texture analyzer Table 2.1 Water activity values of control and hazelnut flour cupcakes using water activity meter Control: no hazelnut 50% hazelnut flour 100% hazelnut flour flour Trial 1 0.879 0.847 0.838 Trial 2 0.690 0.847 0.790 Trial 3 0.827 0.886 0.860 Average 0.799 a 0.860 a 0.829 a Standard deviation 0.098 0.023 0.036 Averages bearing same superscript are not statistically significant, P>0.05

Water activity value 1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 Control 50% hazelnut flour 100% hazelnut flour Variations Figure 2.1 Average water activity values for control and hazelnut flour cupcakes Table 3.1 Volume of control and hazelnut flour cupcakes using seed volume apparatus Control: no hazelnut flour (cm 3 ) 50% hazelnut flour (cm 3 ) 100% hazelnut flour(cm 3 ) Trial 1 140 80 65 Trial 2 75 70 50 Trial 3 85 100 75 Average 100 a 83.3 a 63.3 a Standard deviation 35 15.3 12.6 Averages bearing same superscript are not statistically significant, P>0.05

160 140 120 Volume (cm³) 100 80 60 40 20 0 Control 50% hazelnut flour 100% hazelnut flour Variations Figure 3.1 Average volume of control and hazelnut flour cupcakes using seed volume apparatus Table 4.1 Hedonic scale rating average of palatability of control and hazelnut flour cupcakes (1 is dislike extremely and 9 like extremely) Control: no hazelnut 50% hazelnut flour 100% hazelnut flour flour Trial 1 5.38 5.75 3.38 Trial 2 6.50 6.38 3.75 Trial 3 6.88 5.75 4.00 Average 6.25 a 5.96 a 3.71 b Standard deviation 0.78 0.36 0.31 Averages bearing different superscripts are statistically significant, P<0.05

3.71 5.96 6.25 Variable 2 Variable 1 Control b a Hedonic scale value 8 7 6 5 4 3 2 1 0 a a b Control 50% hazelnut flour 100% hazelnut flour Variations Figure 4.1 Average palatability of control and hazelnut flour cupcakes using 9-point hedonic scale

Table 4.2 Texture average results of control and hazelnut flour cupcakes on 3 point scale (1 being best texture to 3 being worst) Control: no hazelnut 50% hazelnut flour 100% hazelnut flour flour Trial 2 1.5 1.5 3 Trial 3 1.25 1.75 3 Average 1.375 a 1.625 a 3 b Standard deviation 0.177 0.177 0 ***Texture of cupcakes was not evaluated for Trial 1 Averages bearing different superscripts are statistically significant, P<0.05 1.375 1.625 3 Control Variable 1 Variable 2 a b

3.5 3 b Texture value 2.5 2 1.5 1 0.5 a a 0 Control 50% hazelnut flour 100% hazelnut flour Variations Figure 4.2 Average texture of control and hazelnut flour cupcakes using 3-point scale Table 4.3 Appearance results of control and hazelnut flour cupcakes on 3 point scale (1 being best appearance to 3 being worst appearance Control: no hazelnut 50% hazelnut flour 100% hazelnut flour flour Trial 2 1.25 1.75 3 Trial 3 1.5 1.75 3 Average 1.375 a 1.75 a 3 b Standard deviation 0.177 0 0 ***Appearance of cupcakes was not evaluated for Trial 1 Averages bearing different superscripts are statistically significant, P<0.05

1.375 1.75 3 Control Variable 1 Variable 2 a b 3.5 3 b Appearance value 2.5 2 1.5 1 0.5 a a 0 Control 50% hazelnut flour 100% hazelnut flour Variations Figure 4.3 Average appearance of control and hazelnut flour cupcakes using 3-point scale

Table 4.4 Palatability 9-point hedonic scale tally of results for all trials Sample 982 Sample 456 (50% hazelnut flour) (100% hazelnut Sample 623 (Control) flour) Like Extremely 0 0 0 Like Very Much 6 2 5 Like Moderately 5 2 8 Like Slightly 5 2 4 Neither Like nor 1 5 3 Dislike Dislike Slightly 4 2 3 Dislike Moderately 3 0 1 Dislike Very Much 0 3 0 Dislike Extremely 0 8 0 Table 4.5 Texture tally of results for Trial 2 and Trial 3 Sample 982 (50% hazelnut flour) Sample 456 (100% hazelnut flour) Sample 623 (Control) Worst texture 0 16 0 Neither 10 0 6 Best texture 6 0 10 *** Trial 1 was not included in this count because texture of cupcakes was not evaluated for Trial 1 Table 4.6 Appearance tally of results for Trial 2 and Trial 3 Sample 982 (50% hazelnut flour) Sample 456 (100% hazelnut flour) Control Worst appearance 0 15 1 Neither 12 0 4 Best appearance 4 1 11 *** Trial 1 was not included in this count because appearance of cupcakes was not evaluated for Trial 1

Discussion Objective and subjective tests were conducted for all variables (Control, Variable 1: 50% hazelnut flour, and Variable 2: 100% hazelnut flour) in order to test the null hypothesis that using hazelnut flour to replace all-purpose flour will not have an effect on texture, appearance, and palatability of white cupcakes. The objective methods used were the Texture Analyzer, Water Activity System, and Seed Volume Apparatus. The objective data results are provided in Table 1.1, Table 2.1, and Table 3.1, which include force from the Texture Analyzer, water activity values from the Water Activity System, and volume from the Seed Volume Apparatus for three trials of all cupcake variations. Each of these tables include average and standard deviation of the three trials as well. Figure 1.1 presents the average force of control and two variations of hazelnut flour cupcakes. The Tukey-Kramer comparison test shows that there is no significant difference between the variables in terms of texture analyzer data. Table 2.1 includes water activity data for each trial and all variations, along with average and standard deviation. By visual inspection, it becomes apparent that water activity is not greatly affected by the amount of hazelnut flour used in the cupcakes. This is verified by the Tukey-Kramer comparison test which results in no significant difference between the three groups tested. Figure 2.1 displays the average water activity of all trials for the control group and two variations of hazelnut flour (50% hazelnut flour and 100% hazelnut flour). Table 3.1 includes volume data for all trials corresponding to each variation. The average volume is presented in Figure 3.1. It becomes evident that the volumes of hazelnut flour cupcakes were smaller than the volumes of the control. This physical property was studied by Anil when implementing hazelnut testa in bread-making and finding out that the volumes of hazelnut testa breads were reduced using the control breads as a comparison point (Anil, 2007). Nevertheless, similar to texture analyzer and water activity data results, the seed volume apparatus data resulted in no significant difference between the three variables based on the Tukey-Kramer comparison test. It is worth noting that Figure 1.1, Figure 2.1, and Figure 3.1 all include error bars that correlate to the calculated standard deviation corresponding to texture analyzer data, water activity values, and volume consequently. In terms of the subjective evaluation, a degree of significance was found between certain variables based on taste, appearance, and texture of the cupcakes. Taste of the control group and hazelnut flour cupcakes was evaluated using a 9-point hedonic scale that was converted to its corresponding numbering system for statistical analysis of data. Table 4.1 includes the average results for each trial and the average and standard deviation corresponding to all trials. Figure 4.1 displays the average hedonic scale value of all trials and each variation. The Tukey-Kramer comparison test shows that the control is not significantly different from Variable 1 (50% hazelnut flour). However, the control group is significantly different from Variable 2 (100% hazelnut flour). Consequently, Variable 1 is also significantly different from Variable 2. This means that the panelists were able to differentiate between the samples based on the use of hazelnut flour regarding taste of the cupcakes. Taste preference was heavily weighted towards the control group with no hazelnut flour (all-purpose flour) and Variable 1 (50% hazelnut flour and 50% all-purpose flour). Table 4.2 and Table 4.3 present average results corresponding to texture and appearance in that particular order. It is important to mention that texture and appearance was only evaluated for Trials 2 and 3. Originally, a triangle test had been planned prior to Trial 1 but it was realized that it didn t apply to the variables being tested because all three variables differed in amounts and type of flour. As a result, texture and appearance subjective data was not obtained for Trial 1. A change was made to the sensory scorecard that

incorporated specific questions on texture and appearance to evaluate the cupcake samples for Trial 2 and Trial 3. A 3-point scale was used to rank the cupcakes in terms of texture and appearance, 1 for best of the quality tested and 3 for worst of the quality tested. Figure 4.2 displays the average values of texture for each sample from the panelists evaluation. Similarly, Figure 4.3 presents average values in terms of appearance. The Tukey-Kramer comparison test results on both texture and appearance show that there was a significant difference between the control and Variable 2 (100% hazelnut flour); Variable 1 (50% hazelnut flour) and Variable 2 (100% hazelnut flour). The control group and Variable 1 were not significantly different based on texture and appearance. An overall count was also made from the subjective evaluation results with the purpose of obtaining a general view regarding panelist acceptability of hazelnut flour cupcakes based on palatability, texture, and appearance. Table 4.4 includes the results of the 9-point hedonic palatability scale and shows the number of people that liked and disliked the cupcakes made using the two varying amounts of hazelnut flour. It can be seen that 6 panelists liked very much Sample 982, which contained 50% hazelnut flour and 50% all-purpose flour. Additionally, a total of 8 people disliked very much Sample 456, the variation with 100% hazelnut flour. This provides an indication that the best option for consumer acceptability in terms of hazelnut flour is Variation 1, a combination of hazelnut flour and all-purpose flour. Such variation may work for non-celiac gluten-sensitive consumers that are looking to reduce the amount of gluten in their diets to alleviate gastrointestinal discomfort (Hafstrom et al, 2001). Table 4.5 and Table 4.6 show negative acceptability from panelists in terms of texture and appearance of Variation 2 (100% hazelnut flour). Variation 1 which consisted of 50% hazelnut flour obtained good results on average since 6 panelists found it to have the best texture and 4 panelists picked it for best appearance. Although the hazelnut flour variations were not found to be significantly different based on the objective data results from the Texture Analyzer, the Water Activity System, and the Seed Volume Apparatus, the subjective results statistical analysis showed that the use of hazelnut flour to replace all-purpose flour in white cupcakes did have an effect on palatability, texture, and appearance of cupcakes. This indicates that the null hypothesis cannot be accepted. The use of hazelnut flour in the making of cupcakes had an effect on both force and volume of the cupcakes, as it can be seen in the values on Table 1.1 and Table 3.1. Force and volume decreased for hazelnut flour variations. Water activity remained about the same throughout. This condition of low volume was noted by Demirkesen in an experiment that involved the use of chestnut flour in gluten free bread formulations. In this study, they found deterioration in quality parameters (low volume and darker color) of the bread with increased chestnut flour variations (Demirkesen et al, 2010). This was also observed in the variation with a higher amount of hazelnut flour, Sample 456 (100% hazelnut flour), which resulted in lower volumes for all trials. The decrease in force (firmness) of the cupcakes with hazelnut flour may be due to absence of gluten along with gliadin and glutenin, the associated proteins of gluten. Schamne concluded that the control had greater elasticity and structural network for greater firmness than the muffins made with rice flour, maize, and cassava starches because of the presence of glutenin (Schamne, et al, 2010). Sources of error may have been made during the execution of the Seed Volume Apparatus testing because two team members took turns running the test and hitting the seeds down, which means that one person may have used more force than the other. Also, an error could have occurred during the baking process because of differing baking skill levels among team members and factors such as temperature of butter and milk and length of mixing time.

Another source of error could be from either the Water Activity System or the Texture Analyzer not being calibrated properly. Future work could entail testing numerous variables of different ratios of hazelnut flour to all-purpose flour to determine the best recipe for lowering the gluten content of the cupcakes. Also, the use of gums, such as xanthan gum, guar gum, and locust bean gum or even a blend of these gums can be tested in terms of physical properties in the formulation of gluten-free muffins. The incorporation of gums can improve overall quality of final product by binding, thickening, and emulsifying the gluten-free ingredients and helping to maintain good volume and texture. The results of this experiment confirmed that in order to decrease the amount of gluten in cupcakes and to maintain quality, an appropriate ratio of hazelnut flour to all-purpose flour must be established. This will allow gluten-sensitive persons to lower their gluten intake without sacrificing taste.

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