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Article

Development of Functional Muffins with Fruits of the Chilean Forest (Calafate and Maqui) and Supplemented with Prebiotic Fiber

by
Paula García-Milla
1,2,
Rocío Peñalver
1 and
Gema Nieto
1,*
1
Department of Food Technology, Nutrition and Food Science, Veterinary Faculty, University of Murcia, Regional Campus of International Excellence “Campus Mare Nostrum”, Campus de Espinardo, 30100 Murcia, Spain
2
Nutrition and Dietetics Program, Faculty of Health Sciences, Universidad Autónoma de Chile, Providencia 7500975, Chile
*
Author to whom correspondence should be addressed.
Appl. Sci. 2024, 14(17), 7757; https://doi.org/10.3390/app14177757
Submission received: 29 July 2024 / Revised: 26 August 2024 / Accepted: 29 August 2024 / Published: 2 September 2024
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Abstract

:
Inadequate nutrient intake, coupled with increased oxidative stress, leads to an imbalance responsible for the current major diseases. Many foods have traditionally been used as traditional medicine, including native berries from southern Chile. Both Maqui and Calafate possess high antioxidant activity, which grants them medicinal power and makes them an excellent alternative for improving health. The aim of this study is to create a functional food with therapeutic properties capable of counteracting oxidative stress and thereby contributing to improving people’s health. To achieve this, a muffin with inulin, Maqui, and Calafate has been developed. The results show that the incorporation of inulin alone increases the fiber content and antioxidant capacity of the muffins; however, Maqui and Calafate contribute significantly more. Furthermore, contents of phenolic compounds are elevated, and an increase in the folic acid content is observed in the samples compared to the control. We can conclude that producing products with inulin, Maqui, and Calafate can be used to enhance the nutritional value and increase the antioxidant activity of bakery products, providing nutrients while also delivering unique characteristics in color, aroma, and flavor, making them appealing to consumers.

1. Introduction

Reactive oxygen species (ROS) are daily products of human metabolism and perform specific functions in our body, such as gene expression, immune protection, and cellular respiration, among others [1]. However, the uncontrolled production of these species leads to increased oxidative stress that can result in damage to DNA, proteins, and lipids, which may result in an inflammatory response that is responsible for several contemporary diseases, such as diabetes mellitus, cancer [2,3], and other types of disorders, such as Alzheimer’s and Parkinson’s diseases [4,5]. According to Sies H (2015), there are specific forms of oxidative stress, including nutritional or dietary oxidative stress and postprandial oxidative stress [6]. Inadequate nutrient intake, often characterized by the consumption of pro-oxidant foods with low levels of vitamins E and C, as well as the deficiency of carotenoids, polyphenols, and other micronutrients such as selenium, leads to nutritional stress [7]. In addition, postprandial stress is a form of nutritional oxidative stress arising from sustained postprandial diet-induced hyperlipidemia and/or hyperglycemia. This type of stress is associated with a higher susceptibility of LDL lipids to oxidation and, consequently, with increased risks of atherosclerosis, diabetes mellitus, and obesity [7,8]. It is essential for a person to maintain a balanced diet that provides antioxidants such as vitamins A, C, E, and CoQ10 and minerals such as selenium and copper in order to counteract the effects of free radicals and prevent their deregulation [9], since the introduction of dietary antioxidants or exogenous antioxidants could maintain the balance between reactive oxygen species and antioxidants.
Many foods that have traditionally been used as medicines contain these vitamins and minerals, exhibiting high antioxidant and anti-inflammatory properties.
Native berries from Southern Chile are traditionally used as medicine by the indigenous communities who live in this region. Soil and climatic conditions and the distinct nature of this geographic region give rise to fruits with high concentrations of phytochemicals [10,11].
According to recent studies, the plant and fruit species of Southern Chile have high phenolic contents and show high biological activities, with the high antioxidant activity of fruits such as Maqui, Murta, Calafate, strawberry, and Arrayán being highlighted [12,13]. The number of polyphenolic compounds present in these berries ranges from 13 to 36, with “Calafate” having been reported as the fruit with the greatest abundance of compounds, among which we can mention quercetin-3-rutinoside and the presence of gallic, chlorogenic, caffeic, coumaric and ferulic acids, myricetin, and kaempferol [12,14].
Numerous studies have focused attention on these native berries due to their nutritional properties. Undoubtedly, they have become an alternative to enhance health, being consumed as plain fruit or nutraceuticals and including them in preparations such as jams, juices, and yogurt [12,15].
On the other hand, foods with high contents of antioxidants, such as green tea [16] and turmeric [17], could modify the gut microbiota to the point of correcting the so-called intestinal dysbiosis. In this way, antioxidant activity has been linked to the gut microbiota, which plays a fundamental role in inflammatory processes associated with chronic and neurodegenerative diseases [18,19]. In addition, it has been shown that dietary fiber modulates the microbiota, conferring benefits similar to those of antioxidants.
Inulin, a naturally occurring fructan usually obtained from chicory, has multiple health benefits [20]. Among these, it is notable for its role in regulating intestinal function, promoting satiety, controlling lipid and blood glucose profiles, and modulating the gut microbiota [21,22].
Additionally, it is important to highlight that inulin has an antioxidant capacity that could benefit the overall health of those who consume it. Furthermore, inulin is used in the food industry as a specific additive or as a substitute for sugar or oil, thereby improving the nutritional characteristics of products [23].
It is important to highlight that the consumption of foods with a high antioxidant capacity, as well as those with a prebiotic effect, such as inulin, could be beneficial for people with neurodegenerative diseases such as Parkinson’s disease. Oxidative stress has been observed to promote brain damage, stemming from mitochondrial dysfunction and the production of inflammatory cytokines, which lead to abnormalities in the blood–brain barrier. According to research, antioxidant therapy could be an alternative to reduce damage and delay the progression of the disease [4].
Given the growing interest in the consumption of diet-derived antioxidants, the present study aims to create a functional food with therapeutic properties and the ability to ameliorate oxidative stress, therefore contributing to the improvement of people’s health, including people with neurodegenerative diseases such as Parkinson’s disease. For this purpose, a muffin has been supplemented with food items such as inulin, Maqui berry and Calafate berry.

2. Materials and Methods

Four products were developed based on a basic Chilean sponge cake recipe. The products included the control, a muffin with inulin, a muffin with inulin and Calafate, and finally a muffin with inulin and Maqui. The products and their ingredients are detailed in Table 1. For the preparation of the products, the dry or powdered ingredients were mixed. In a separate container, the egg whites were beaten until stiff peaks formed, and then the liquid ingredients, such as milk and oil, were added. After that, the dry mixture was combined with the wet mixture and stirred until a homogeneous mixture was obtained. This was then placed in a preheated oven at 180 °C for 30 min.
Each of the developed products underwent techniques and protocols previously published to determine their chemical composition.

2.1. Determination of the Nutritional Composition

2.1.1. Moisture

The moisture content was determined according to the AOAC method (1995). The procedure involved weighing 3 g of the sample in a petri dish, which was then left for 24 h in an oven at 110 °C. The results were expressed as a percentage using the formula % Moisture = 100 − Dry matter [24,25].

2.1.2. Inorganic Matter

Inorganic matter was obtained by the complete incineration of 1 g of fresh sample in a muffle furnace at 525 °C. Its determination was carried out according to the following formula: % ash = ((W2 − W0) / (W1 − W0)) × 100, where W0 is the weight of the empty crucible, W1 is the weight of the crucible plus the sample, and W2 is the weight of the crucible plus the ash [25].

2.1.3. Determination of Energy

For the determination of energy, the energy value of macronutrients such as protein, carbohydrates, and fats were obtained by summing them, according to the Atwater conversion values (WHO, 1985) [24,26]

2.1.4. Determination of Protein

Following the Kjeldahl protocol described by AOAC 955.04 [26], 2 g of fresh sample was weighed and 1 catalyst tablet plus 15 mL of concentrated sulfuric acid were added. Then, digestion occurred at 450 °C for 1 h. Once the sample had cooled, distillation was performed with 38% NaOH until a color change was observed in the sample. After that, the sample was titrated with 0.1 N HCl.
For the calculation, the following formula was used: % of crude protein
((V2 − V1) × 0.1)/W × 1.4 × F
where V1 is the volume in mL of HCl solution required; V2 is the volume in mL of HCl required for the sample; normality is considered at 0.1; and W is the weight of the sample in grams.
F is the conversion factor (6.25: meats, fish, eggs, legumes, and vegetable proteins; 5.7: cereals and soy derivatives; 6.38: milk and dairy products; 5.55: gelatin; and 5.95: rice).

2.1.5. Determination of Carbohydrates

Carbohydrates were obtained according to the following formula: Total carbohydrates (%) = 100 − (fat + ash + protein + moisture + dietary fiber) [24,27]

2.1.6. Determination of Fat

Following the AOAC protocol (1990) [25], 1 g of the previously dried sample (from the moisture determination) was weighed and placed in a Tecator Soxhlet fat extractor with 50 mL of ether. To avoid sample contamination, tweezers and gloves were used throughout the entire procedure. Once the procedure was completed, the samples were placed in an oven to dry and remove any remaining moisture. After that, they were left to cool in a desiccator and then weighed.
Total fat was calculated according to the following formula:
Fat (%) = ((W1 − W2)/W) × 100
where, W1 is the weight of the cup with ether extract or fat residue; W2 is the weight of the empty cup; and W is the weight of the sample.

2.1.7. Determination of Folate

A total of 1 g of the sample was mixed with 25 mL of extraction buffer containing 2 g of sodium ascorbate/100 mL and 2-mercaptoethanol 10 mmol/L, pH 7.85. This was performed in a nitrogen atmosphere.
The samples were boiled for 10 min, and once cooled, the pH was adjusted to 4.9 with 60 mmol/L HCl. Then, a 5 mL aliquot was incubated for 3 h at 37 °C in a nitrogen atmosphere with 1 mL of pig kidney conjugase and 1 mL of α-amylase preparation (20 mg/mL in 1 g of sodium ascorbate/100 mL). Afterward, the samples were boiled at 100 °C to inactivate the enzyme, cooled, filtered with a 0.45 μm pore filter, and then connected to an anion exchange cartridge (SAX) attached to a 12-port vacuum manifold.
The analysis of the samples was performed on an HPLC/MS system. The complete protocol is described in the article “Formulation and Physical–Chemical Analysis of Functional Muffin Made with Inulin, Moringa, and Cacao Adapted for Elderly People with Parkinson’s Disease” https://doi.org/10.3390/antiox13060683 (accessed on 29 August 2024), following the protocol described by Peñalver R [28] based on Vahteristo et al. (1996) [29,30,31].
Data analysis was conducted with the MassHunter Qualitative Analysis Navigator software (Agilent Technologies, Rev. B.08.00).

2.1.8. Determination of Dietary Fiber

Following the AOAC 985.29 (1990) [32] protocols, 1 g of the sample was weighed and 50 mL of 0.005 m phosphate buffer at pH 6 was added. The mixture was then heated to 95 °C, and 0.2 mL of α-amylase was added. After 30 min, the samples were cooled to room temperature to adjust the pH to 7.5.
Then, in a 60 °C water bath, 1 mg of protease was added, and the mixture was stirred for 30 min. Afterward, the samples were cooled to adjust the pH to 4.5, and 0.3 mL of amyloglucosidase enzyme was added. The samples were left for 30 min at 60 °C.
For sample filtration, 280 mL of 95% ethanol at 60 °C was added, allowing the fiber to precipitate for at least 1 h. The samples were then filtered using a Fibertec™ 1023 analyzer. In a crucible containing 0.5 g of Celite, the samples were placed, and the residue was left in an oven. Once dry, they were weighed, and the crucibles were removed to determine the protein and ash content.

2.1.9. Antioxidant Capacity

The oxygen radical absorption capacity (ORAC) technique was developed following the protocol described by Ehlenfeldt and Prior (2001) [33].
A black 96-well plate was filled with 200 μL of distilled water. The blanks consisted of 20 μL of phosphate buffer and 20 μL of Trolox samples and standards.
After preparing the plates, the process was initiated in the Synergy HT plate reader following purging with water and the necessary reagents. The GEN 5 program was employed to establish the protocol. First, 200 μL of fluorescein was added to each well. After 15 min, 20 μL of AAPH (0.216 g in 10 mL phosphate buffer) was introduced into each well. Measurements were taken every minute for an hour and a half with 485 nm excitation and 528 nm emission. The entire procedure was conducted at 37 °C. Supporting software was utilized to calculate the area under the curve for all readings, and the data were extrapolated using Trolox standard curves with known concentrations [29].
The Ferric Reducing Antioxidant Power (FRAP) technique was performed following the protocol outlined by Benzie and Strain (1999) [33]. A FRAP solution was prepared by mixing 20 mL of acetate buffer, 2 mL of TPTZ, and 2 mL of FeCl3·6H2O. In each cuvette, 1 mL of the FRAP solution and 100 μL of the sample were added. Absorbance readings were taken at 593 nm over a 4 min period, with the FRAP solution serving as the blank. A standard Trolox curve was generated to compare the results.
The ABTS method was conducted according to the procedure described by Re et al. (1999) [34]. One mL of ABTS solution and 100 μg of the sample were combined, stirred for 30 s, and then the absorbance was measured at 734 nm after 2 min. The blank consisted of an ABTS solution with an absorbance of approximately 0.7, and was adjusted using water and MnO2
The DPPH method was based on the procedures outlined by Brand-Williams et al. (1995) and Sánchez-Moreno et al. (1998) [34,35]. The DPPH reagent was made by dissolving 0.0063 g of DPPH in 250 mL of ethanol and stored away from light. A mixture of 3.9 mL of DPPH reagent and 100 μL of the sample or standard solution was prepared. The absorbance was measured at 515 nm after 30 min of mixing, while keeping the preparation away from light. Methanol was used for zeroing, and the blank consisted of the DPPH reagent without any samples.

2.2. Colorimetry of Products

For the colorimetry technique, a tristimulus colorimeter was used to measure the colors red, green and blue. To do this, a white table was used with opaque white paper on which the samples were placed. The lighting in the room was natural lighting.
The color measurement was performed based on the color coordinates of the Hunter color system [36].
The techniques and protocols used in the methodology were previously described in the article “Formulation and Physical–Chemical Analysis of Functional Muffin Made with Inulin, Moringa, and Cacao Adapted for Elderly People with Parkinson’s Disease” https://doi.org/10.3390/antiox13060683 (accessed on 29 August 2024).

2.3. Statistical Analysis

Statistical evaluations were conducted using the STATA software version 16 for data analysis. The ANOVA parametric test was employed to assess variance, while the Bonferroni test was utilized for multiple comparisons, with a significance threshold set at p < 0.005.

3. Results

3.1. Composition of the Product

The addition of inulin has been reported in diverse products offered by the food industry; however, most of the described uses are based on its function as a thickener, emulsifier, and gelling agent. It can also be used in food preparations as a substitute for sugar and/or fats [37].
The preparation of the muffins described in Table 1 was based on the recommendations of Rodriguez J (2014), where part of the product’s fats was replaced by chicory-derived inulin [38].

3.2. Determination of Macronutrients

The caloric contribution of the standard muffin (control) is 367.02 ± 13.60 calories per 100 g of product. When inulin was used to substitute oil (MI sample), it was observed a reduction of about 28 calories when compared to the control, with a caloric contribution of 339.91 ± 4.06 calories per 100 g of product. However, the samples with added Chilean berries (Calafate sample [MICa] and Maqui sample [MIMa]) had a contribution of 302.56 ± 44.75 and 303.83 ± 4.71, respectively, resulting in a reduction of up to 65 calories with respect to the control (Table 2).
Protein contents remained similar between the samples, with a no more than 0.67 g/100 g of difference between the sample with the lowest protein contribution and the sample with the highest protein contribution. However, it can be seen that the MI and MIMa samples had higher protein values compared to the control (Table 2). When calculating the protein percentage, we found that proteins accounted for 7% of the caloric value in the case of the control, while in the samples MI, MICa, and MIMa, the percentage reached 8.3%, 8.5%, and 9.3%, respectively.
On the other hand, it was found that carbohydrates (Carbs) increased by 11 g on average in the samples that contained inulin (MI, MICa, and MIMa), with the highest value observed in the Calafate sample (MICa) at 54.40 ± 7.43 g of Carbs/100 g of product, followed by the Maqui sample (MIMa) at 52.53 ± 0.91 g of Carbs/100 g of product, and finally the inulin sample, which did not contain fruits of Southern Chile (MI), at 46.53 ± 0.53 g of Carbs/100 g of product. These samples were compared to the control, the value of which was 43.23 ± 4.67 g of Carbs/100 g of product.
Despite the results from the analysis of variance regarding the nutritional composition of the samples that did not show statistically significant values, it is worth mentioning that the fat contribution had a tendency (p = 0.0621) to decrease in samples containing inulin (MI, MICa, and MIMa) compared to the control. This was more evident in the Calafate sample with 7.10 ± 5.40 g/100 g, followed by the Maqui sample with 7.24 ± 0.77 g/100 g, which were compared to the control that presented 18.69 ± 3.67 g/100 g (Table 2).

3.3. Dietary Fiber

Table 3 shows the analysis of the insoluble, soluble, and total fiber contents of the samples analyzed. It can be noted that, in general, the muffins contained more soluble than insoluble fiber. When comparing between samples, we can notice that values for soluble fiber were similar between samples; the same occurred in the case of insoluble fiber. However, when looking at total fiber, we can mention that incorporating inulin into the product enhanced the total fiber content of foods.

3.4. Determination of Antioxidant Activity

The ABTS, DPPH, ORAC, and FRAP assays were used to assess antioxidant activity. In Table 4, it can be noted an increase in the antioxidant activity of all samples containing inulin (MI, MICa, and MIMa) in comparison to the control. The Calafate sample (MICa) exhibited the highest antioxidant activity, followed by the Maqui sample (MIMa) and finally the sample without Chilean berries that only contained inulin (MI).
Although the results displayed in Table 4 are quite noticeable regarding the difference between the inulin samples and the control, it is important to emphasize that only the ORAC and FRAP assays have shown a statistically significant difference. In relation to these assays, the samples containing Maqui and Calafate exhibited the highest antioxidant activities, the most representative of which is the Calafate sample, as assessed by the ORAC assay, with a value of 100.89 ± 2.00 μmol TE/g, followed by the Maqui sample, with 90.69 ± 5.41 μmol TE/g, and the sample containing inulin without berries, with a value of 67.92 ± 5.84 μmol TE/g (p < 0.005). On the other hand, the values obtained using the FRAP assay showed similar antioxidant activities in the Calafate and Maqui samples, with values of 12.03 ± 4.48 μmol TE/g and 12.03 ± 1.13 μmol TE/g, respectively, followed by the sample without Chilean berries, with 3.18 ± 0.10 μmol TE/g (p < 0.005).
Since the MI sample (muffin with inulin) is a preparation that consisted of using the same procedure as the control but incorporating inulin as a substitute for a portion of fat (oil), observing a statistically significant difference in antioxidant activity compared to the control was not expected. However, it was found that its antioxidant activity increased by 23.3 μmol TE/g in the ORAC assay and 1.02 μmol TE/g in the FRAP assay when compared with the control (Table 4). In view of these results and considering the possibility of conducting ABTS and DPPH assays on inulin, a sample of this fiber was analyzed separately from the batter in order to measure its antioxidant activity using the aforementioned assays. Although results showed rather low values, antioxidant activity is indeed present in inulin (Table 5), conferring greater antioxidant properties to the products that contain this ingredient as compared to the control.

3.5. Phenolic Compounds

An analysis of phenolic compounds was performed, with the results showing a statistically significant increase in all samples containing inulin (MI, MICa, and MIMa) compared to the control. However, the sample that exhibited the highest content of phenolic compounds was the preparation with Calafate (MICa), with 8.91 ± 0.07 mg gallic acid/g sample, followed by the Maqui sample (MIMa), with 8.75 ± 0.18 mg gallic acid/g sample, and finally the sample with inulin without berries (MI), with 3.28 ± 0.22 mg gallic acid/g sample (p = 0.000) (Table 6).

3.6. Folate Content

Table 7 shows the concentration of each of the four folate monoglutamates, as well as the total folate monoglutamate concentration and total folate concentration in the samples. In the aforementioned table, it can be noted a reduction in the total folate content in the samples containing inulin (MI, MICa, and MIMa). However, the Calafate sample (MICa) has a total folate value of 587.09 ± 1.17 µg folic acid equivalents/100 g, followed by the Maqui sample (MIMa) with 548.68 ± 6.35 µg folic acid equivalents/100 g and finally the sample that only contained inulin (MI) with 288.25 ± 3.33 µg folic acid equivalents/100 g. Despite these findings, it was observed that the content of folic acid was higher in the samples added with inulin, with the Maqui sample exhibiting the highest value at 45.84 ± 0.63 μg/100 g, followed by the Calafate sample with 44.77 ± 0.67 μg/100 g, and finally the sample only containing inulin without addition of fruits from Southern Chile at 34.46 ± 1.25 μg/100 g, showing statistically significant differences with respect to the control that had 18.98 ± 1.52 μg/100 g (p = 0.000). Furthermore, the value of 5-MTHF also experienced an increase in the samples containing inulin, with the highest concentration being found in the Calafate sample (MICa) with 101.55 ± 1.04 μg/100 g, followed by the Maqui sample (MIMa) with 91.43 ± 1.01 μg/100 g and finally the sample with inulin without berries (MI) with 73.16 ± 1.11 μg/100 g (p = 0.000).
It is worth mentioning that among all vitamers, the Calafate and Maqui samples exhibited enhanced and higher concentrations compared to the sample that only contained inulin, with 5F-THF being the vitamer with the highest concentration, equivalent to 398.11 ± 1.86 μg/100 g in the Calafate sample (MICa) (p = 0.000) (Table 7).

3.7. Colorimetry

The color analysis of the samples, performed using a colorimeter, showed a statistically significant difference in all color analyses, as shown in Table 8. The control had a higher brightness, total color, and presence of yellow shades; while the product with the lowest brightness corresponded to the Maqui sample (MIMa), followed by the Calafate sample (MICa), which was predictable due to the shades exhibited by the fruits of the Chilean forests (Figure 1). In addition, there were fewer shades of yellow in these samples (MICa and MIMa), as well as lower saturation, when compared to the control.
The sensory analysis of the product included the participation of a group of people with Parkinson’s disease from the Murcia community. A total of 23 individuals whose average age was 59.1 ± 18 years were involved in the sensory evaluation procedure. Participants consisted of 11 men (48%) and 12 women (52%). The descriptive analysis of the sensory evaluation is displayed in Table 9. The evaluation was conducted according to a five-point hedonic scale, where five meant “like very much” and one meant “dislike very much”.
According to the results, the control product obtained the highest score in overall color, followed by the sample with added inulin. Similar results were obtained regarding the appearance of the samples, where the products with added Maqui and Calafate had the lowest scores.
In relation to aroma, scores were similar, varying between 3.13 and 3.17 for the inulin samples, with the control obtaining 3.74. Although average values in general did not reach five, purchase intention and overall acceptability values were above three points, which is quite acceptable. The product with added Maqui and fiber obtained the highest purchase intention score. It should be noted that overall acceptability was not substantially different between products with added inulin (MI, MIMa, and MICa), showing values between 3.61, 3.61, and 3.57, respectively.
Out of the subjects who participated in the tasting, 91% indicated that they “liked” the product when asked about overall flavor of the products tasted, followed by 5% who reported they “neither liked nor disliked”. No participant expressed a sense of dislike for any of the products tasted; however, 5% did not answer this question (Figure 2).
It should be noted that users reported an occasional consumption of muffins (61%), followed by a consumption frequency of one to three times a week (13%), four to five times a week (9%), and every day (9%). Only 4% mentioned they never consume this type of product (Figure 3).
One hundred percent of users indicated that they were willing to buy an improved product that provides health benefits.

4. Discussion

According to published studies, inulin might be used to replace the fat content in the production of bakery and confectionery products, with replacement percentages ranging from 19% to 100% [38,39]. For the purposes of this study, we conducted initial tests that consisted of the preparation of products supplemented with 40%, 50%, 60% and 70% inulin. Product taste tests included the participation of a group of experts of Murcia University. The expert group was composed of nutrition professionals and food engineers who were part of research teams dedicated to developing new products, with experience in sensory evaluation and product tasting.
After this evaluation, it was found that the best concentration of inulin was 60%, since it did not alter sensory characteristics, particularly the texture perceived on the palate, and it may enhance the nutritional value of the product. It should be noted that as more inulin is incorporated, the texture is perceived as “harder” or “gummier” due to the formation of gels resulting from the addition of this dietary fiber.
Once the muffin with added inulin was prepared, two Chilean berries (Calafate and Maqui) were included with the aim of providing greater antioxidant activity, thus developing a functional food with improved therapeutic properties. According to Ruiz A et al. (2014), Berberis microphylla G. Forst, also known as “Calafate”, along with other 20 species of edible berries and fruits, contain high levels of anthocyanins and hydroxycinnamic acid. As shown by the findings of their study, Calafate has a total flavonol concentration of 1.33 ± 0.54 μmol/g, with pulp and skin having the highest concentrations of flavonol compared to seeds [20]. The authors reported that polyphenolic compounds were present in glycosylated form, with 3-glycoside conjugates being the most abundant anthocyanins. The total anthocyanin content of Calafate berry was 17.81 ± 0.98 μmol g−1; similar results were found in Maqui berries (17.88 ± 1.15 μmol g−1). These fruits exhibit an antioxidant activity that is primarily due to the presence of polyphenols, particularly anthocyanin-based compounds, which represent 80% of the total compounds. This turns Maqui and Calafate into fruits with high antioxidant activities [14,40].
Inulin is a natural fructan extracted from plants such as chicory or Jerusalem artichoke. It is an indigestible polysaccharide due to the types of glycosidic bonds present in its structure [20]. However, inulin is not only an indigestible polysaccharide or a dietary fiber; research studies have found that it has antioxidant activity. A group of researchers who assessed the in vitro antioxidant activity of inulin by comparing it to that of vitamin C, which was used as a control, found that as the concentration of inulin increased from 0.25 to 10 mg/mL, the DPPH radical scavenging activity of inulin increased linearly (R2 = 0.985, p < 0.05). However, the DPPH-scavenging ability of inulin was significantly lower than that of vitamin C at doses ranging from 0.25 to 10 mg/mL (p < 0.05) [41]. In the context of our study, we conducted a DPPH measurement of inulin in concentrations in accordance with the protocol described in the Section 2 (100 μL), finding an activity of 9.092518 ± 10.43 μmol TE/g. On the other hand, the ABTS assay yielded similar results for the research team of Hong Mei et al., where antioxidant activity increased linearly (R2 = 0.988, p < 0.05) as inulin levels increased; however, this was also lower when compared to that of vitamin C [20]. The results obtained in the ABTS assays conducted in our study indicated an activity of 262.5728 ± 34.74 μmol TE/g. Although inulin is not a potent natural antioxidant, it is safe to say that to the extent this dietary component has a greater presence, there could be benefits associated with antioxidant activity, in addition to benefits already attributed to inulin-derived dietary fiber. This was concluded by a study that evaluated its antioxidant activity and its role in preventing the impairment of human colonic muscle cells. The study results showed a significantly higher antioxidant activity when compared to that of simple sugars. In addition, inulin protected the human colonic mucosa from LPS-induced damage, suggesting that inulin may protect the gut from oxidative stress-related damage [42].
Inulin has been widely used in the food industry and incorporated into several products; however, its use in bakery and confectionery products is probably the most well-known. In our study, we have used inulin as a replacement for oil at a percentage of 60%, since higher concentrations altered the structure of the dough, turning it dryer, tougher or rubberier. Rodríguez J and her team, who examined replacement levels of 0%, 35%, 50%, 70%, and 100%, concluded that using inulin instead of oil significantly decreased (p < 0.05) batter viscosity. It was determined that the best concentration was 70%, since the resulting products were softer and classified as acceptable by an untrained sensory panel [43]. However, in another study that addressed the use of inulin as a fat substitute (in concentrations of 50%, 75%, and 100%) to be used in bakery products, it was concluded that as the percentage of inulin increased, the volume of muffins decreased. Moreover, a descriptive analysis revealed significant effects on the appearance of the product, determining that the best replacement concentration was 50%, which resulted in comparable muffins whose crumb was slightly firmer [44].
On the other hand, Rodríguez JP (2016) reached the conclusion that using inulin as a replacer at a concentration of 45% would result in a reduction of 22 calories, equivalent to 36% of calories from fats and 14% of the total caloric value. In our study, it was found that the average calorie reduction was 51.5 calories, with the lowest reduction being observed in the inulin sample that did not contain berries (MI), at 27.11 calories; followed by the sample with inulin and Maqui (MIMa), at 63.19 calories; and finally, the greatest calorie reduction was seen in the sample with added inulin and Calafate (MICa). On the other hand, our products showed an average reduction of 55.93 g in terms of the fat contribution, and when the reduction in fats was ranked from the lowest to the highest, the MI, MIMa, and MICa samples had 4.75 g, 11.45 g and 11.59 g, respectively. These results differ from those obtained by Rodríguez JP, who achieved a reduction in fats from 4 g to 2.4 g in the development of his inulin product [45]. It should be noted that reductions in fat contents and calories have also been found by Doménech G et al. (2015), whose study demonstrated a statistically significant reduction in both macronutrients in all products with added inulin, which included cookies, croissants, muffins, and sponge cakes [46].
The results of our dietary fiber analyses did not show statistically significant differences; however, it was found an increase in the total fiber content of the products supplemented with inulin. In general, studies that have examined the addition of inulin in bakery and confectionery products are primarily focused on the organoleptic characteristics that inulin confers to the dough, as well as on the ability of inulin to contribute to fat reduction in the final product. However, it is quite hard to find studies focused on the total dietary fiber content [43,47,48,49]. In spite of this difficulty, we have found a paper which states that fructan loss caused by the baking process in bakery products ranges from 35% to 47% and depends on the harvest of the product, as well as on the companies that produce the inulin powder from Jerusalem artichokes, while fructose production after hydrolysis ranges from 11% to 45.8% [50].
It is worth noting that despite the relevance of studying prefabricated products with a high content of antioxidants or that are beneficial to people’s health, no studies on products similar to those analyzed in this research work have been found. After conducting a search in PubMed, we only found studies on the antioxidant activity of Maqui extracts or the antioxidant activity of Maqui and/or Calafate fruits, but no research on the value that these berries provide to bakery and confectionery products was found. However, research has been conducted on the preparation of beverages and juices, mainly on Maqui-based beverages. One of these studies was about a liquor into which Maqui berry had been incorporated. The results obtained from the FRAP assay showed an increase in antioxidant activity during the first 30 days that remained stable in the following days until the end of the study (5.35 ± 0.26 and 13.99 ± 0.74 mM Trolox for the control product and the Maqui product, respectively) [51], while in our study including muffins supplemented with Maqui, we obtained a value of 12.03 ± 1.13 μmol TE/g in the analysis conducted using the same assay (FRAP). On the other hand, the ORAC assay conducted in the study that discussed the Maqui-based liquor showed an increase throughout maceration, reaching final values of 25.37 ± 3.37 and 48.41 ± 3.08 mM Trolox for the control product and the Maqui product, respectively [52], while our product obtained 90.69 ± 5.41 μmol TE/g for the Maqui product. Most studies on beverages and juices have reported and identified the presence of anthocyanins, flavones, and other compounds, which demonstrates that the use of berries from Southern Chile enhance the nutritional value and antioxidant activity of juices [52,53,54].
With respect to folates, in the year 1951, Decree 1934 was enacted in Chile, which established that flour must be fortified with iron, calcium, and vitamins of the B-complex. Subsequently, in 1999, the regulation was modified, requiring mills to implement the mandatory addition of folic acid to wheat flour in a concentration of 2.2 mg/kg [55,56]. This measure contributes to the improvement of nutritional value and folate contribution of wheat flour-based products. A study published in the book Nutrition and an active life: from knowledge to action by the Pan American Health Organization (PAHO), in which researchers analyzed bread by purchasing 1 kg of bread at 50 different bakeries from the Metropolitan Region of Chile, reported that the folate content of 100 bread samples was 202 ± 94 µg/100 g of product. From the 100 samples, it was found that nine contained less than 37 μg of folic acid/100 g; researchers drew the conclusion that these nine bread samples were prepared with unfortified flour [57].
Müller H conducted research that analyzed bakery products, cereals, and legumes. The study revealed either similar results or values well below those shown in the Chilean bread analysis. It was found that bakery products had a folate content of 14 µg/100 g when made from rye flour (whole grain rye flour) and 88 µg/100 g in the case of crispbread [58].
It should be mentioned that the products analyzed in this study were prepared using wheat flour obtained from a market located in the city of Murcia, Spain. Despite this fact, our analyses showed that the product supplemented with inulin and Maqui obtained a value of 45.84 ± 0.63 μg/100 g, followed by the product with inulin and Calafate of 44.77 ± 0.67 μg/100 g, and finally the product that did not contain berries at 34.46 ± 1.25 μg/100 g, compared to the control at 18.98 ± 1.52 μg/100 g. These values are above those found in the analysis of Chilean bread.

5. Conclusions

According to the results obtained, the inclusion of inulin in bakery products such as muffins resulted in an increased fiber content and antioxidant activity. Therefore, this dietary fiber could improve health by exerting its known effect as a prebiotic fiber but also by acting as an antioxidant.
On the other hand, Maqui and Calafate berries have a high antioxidant power, which enhances the functional nutritional value of muffins.
We can conclude that the preparation of products with added inulin, Maqui and Calafate may be useful to improve nutritional and functional value of baked goods, not only providing nutrients but also conferring unique characteristics in terms of color, aroma, flavor, and antioxidants capacity.
This research allows us to affirm that the incorporation of inulin and berries from southern Chile makes it possible to obtain a product that could improve health without affecting its organoleptic characteristics, thus achieving excellent acceptability. The high antioxidant capacity could help reduce reactive oxygen species, improving neurological health and preventing or delaying some diseases.
Recent research on the use of Calafate and Maqui focuses on their nutritional characteristics. However, this study confirms that these fruits can be incorporated into bakery and pastry preparations with excellent acceptability.

Author Contributions

Validation, G.N.; Investigation, P.G.-M., R.P. and G.N.; Writing—original draft, P.G.-M. and G.N.; Writing—review & editing, R.P. and G.N.; Supervision, G.N.; Funding acquisition, G.N. All authors have read and agreed to the published version of the manuscript.

Funding

This research received no external funding.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

The data presented in this study are available on request from the corresponding author.

Conflicts of Interest

The authors declare no conflict of interest.

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Figure 1. Shades of the products analyzed.
Figure 1. Shades of the products analyzed.
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Figure 2. Descriptive analysis of overall muffin likeability, as reported by volunteers participating in the sensory analysis of the samples. N/I: not informed.
Figure 2. Descriptive analysis of overall muffin likeability, as reported by volunteers participating in the sensory analysis of the samples. N/I: not informed.
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Figure 3. Descriptive analysis of the muffin consumption frequency reported by volunteers participating in the sensory analysis of the samples. N/I: not informed.
Figure 3. Descriptive analysis of the muffin consumption frequency reported by volunteers participating in the sensory analysis of the samples. N/I: not informed.
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Table 1. Sample formulation (n = 4).
Table 1. Sample formulation (n = 4).
SamplesFlour (g)Baking Powder (g)Sugar
(g)		Eggs
(units)		Semi-Skimmed Milk
(g)		Olive Oil
(g)		Inulin (g)Calafate
(g)		Maqui
(g)
Control1505100111550000
MI15051001115203000
MICa15051001115203060
MIMa15051001115203006
Control: standard muffin; MI: muffin with inulin as a fat replacer; MICa: muffin with inulin (MI) + Calafate berries; MIMa: muffin with inulin (MI) + Maqui berries.
Table 2. Nutritional composition of the samples (n = 4) (grams/100 g).
Table 2. Nutritional composition of the samples (n = 4) (grams/100 g).
Samples
ControlMIMICaMIMa
Moisture23.44 ± 0.16 (ns)23.55 ± 0.25 (ns)23.67 ± 1.02 (ns)24.01 ± 1.27 (ns)
Energy367.02 ± 13.60 (ns)339.91 ± 4.06 (ns)302.56 ± 44.75 (ns)303.83 ± 4.71 (ns)
Proteins6.48 ± 0.19 (ns)7.08 ± 0.55 (ns)6.46 ± 1.18 (ns)7.13 ± 0.35 (ns)
Carbohydrates43.23 ± 4.67 (ns)46.53 ± 0.53 (ns)54.40 ± 7.43 (ns)52.53 ± 0.91 (ns)
Fat18.69 ± 3.67 *13.94 ± 0.45 *7.10 ± 5.40 *7.24 ± 0.77 *
Ash0.93 ± 0.13 (ns)1.15 ± 0.11 (ns)1.01 ± 0.09 (ns)1.05 ± 0.11 (ns)
Control: standard muffin; MI: muffin with inulin as a fat replacer; MICa: muffin with inulin (MI) + Calafate berries; MIMa: muffin with inulin (MI) + Maqui berries. ANOVA test: * statistical trend (p 0.0621); ns: not statistically significant.
Table 3. Determination of fiber in the samples (n = 4) (grams/100 g).
Table 3. Determination of fiber in the samples (n = 4) (grams/100 g).
Samples
ControlMIMICaMIMa
Insoluble Fiber0.87 ± 0.41 (ns)0.95 ± 0.05 (ns)0.85 ± 1.15 (ns)1.10 ± 0.40 (ns)
Soluble Fiber6.35 ± 0.68 (ns)6.80 ± 0.18 (ns)6.49 ± 0.90 (ns)6.34 ± 0.34 (ns)
Total Fiber6.45 ± 1.09 (ns)7.59 ± 0.22 (ns)8.80 ± 2.05 (ns)7.70 ± 0.75 (ns)
Control: standard muffin; MI: muffin with inulin as a fat replacer; MICa: muffin with inulin (MI) + Calafate berries; MIMa: muffin with inulin (MI) + Maqui berries. ANOVA test; ns: not statistically significant.
Table 4. Intergroup analysis of the antioxidant activity of the samples (n=4), measured as % in ABTS, and DPPH assays and as μmol TE/g in ORAC and FRAP assays.
Table 4. Intergroup analysis of the antioxidant activity of the samples (n=4), measured as % in ABTS, and DPPH assays and as μmol TE/g in ORAC and FRAP assays.
SamplesAssay
DPPH
(%)		ABTS
(%)		ORAC (μmol TE/g)FRAP (μmol TE/g)
Control579.92 ± 33.032446.39 ± 723.3444.58 ± 6.97 **2.16 ± 0.28 **
MI661.14 ± 104.152835.81 ± 723.3467.92 ± 5.84 **3.18 ± 0.10 **
MICa742.67 ± 70.673600.12 ± 969.93100.89 ± 2.00 **12.03 ± 4.48 **
MIMa905.74 ± 360.794130.49 ± 1321.3390.69 ± 5.41 **12.03 ± 1.13 **
Control: standard muffin; MI: muffin with inulin as a fat replacer; MICa: muffin with inulin (MI) + Calafate berries; MIMa: muffin with inulin (MI) + Maqui berries. ** statistically significant (p < 0.05) in analysis of variance.
Table 5. Antioxidant activity of inulin as measured by the ABTS and DPPH assays, represented as the mean ± standard deviation (sample analyzed in triplicate).
Table 5. Antioxidant activity of inulin as measured by the ABTS and DPPH assays, represented as the mean ± standard deviation (sample analyzed in triplicate).
Mean/Standard DeviationABTS (μmol TE/g)DPPH (μmol TE/g)
Inulin262.5728 ± 34.749.092518 ± 10.43
Table 6. Phenolic compound contents of the samples (n = 4) (mg gallic acid/g sample).
Table 6. Phenolic compound contents of the samples (n = 4) (mg gallic acid/g sample).
ParametersSamples
ControlMIMICaMIMa
Folin (mg gallic acid/g sample)2.35 ± 0.03 **3.28 ± 0.22 **8.91 ± 0.07 **8.75 ± 0.18 **
Control: standard muffin; MI: muffin with inulin as a fat replacer; MICa: muffin with inulin (MI) + Calafate berries; MIMa: muffin with inulin (MI) + Maqui berries. ** statistically significant (p = 0.000) in the ANOVA.
Table 7. Folate vitamers (FA, THF, 5M-THF, and 5F-THF, expressed as µg/100 g FW) and total folate (expressed as µg of folic acid equivalents/100 g FW). (n = 4).
Table 7. Folate vitamers (FA, THF, 5M-THF, and 5F-THF, expressed as µg/100 g FW) and total folate (expressed as µg of folic acid equivalents/100 g FW). (n = 4).
Samples (n = 4)Folic AcidTHF5M-THF5F-THFTotal
Control18.98 ± 1.52 **148.00 ± 10.15 **70.97 ± 0.16 **374.27 ± 2.85 **612.22 ± 5.95 **
MI34.46 ± 1.25 **8.43 ± 1.83 **73.16 ± 1.11 **172.20 ± 3.87 **288.25 ± 3.33 **
MICa44.77 ± 0.67 **42.66± 0.58 **101.55 ± 1.04 **398.11 ± 1.86 **587.09 ± 1.17 **
MIMa45.84 ± 0.63 **49.11 ± 3.32 **91.43 ± 1.01 **362.30 ± 2.65 **548.68 ± 6.35 **
THF: tetrahydrofolate; 5-MTHF: 5-methyltetrahydrofolate; 5-FTHF: 5-formyltetrahydrofolate. Control: standard muffin; MI: muffin with inulin as a fat replacer; MICa: muffin with inulin (MI) + Calafate berries; MIMa: muffin with inulin (MI) + Maqui berries. ** statistically significant (p = 0.000) in analysis of variance.
Table 8. Color assessment of the samples (n = 4) according to the coordinates of the Hunter color system.
Table 8. Color assessment of the samples (n = 4) according to the coordinates of the Hunter color system.
Samples (n = 4)Colorimetry
CLabh
Control29.12 ± 2.35 **61.55 ± 2.11 **4.70 ± 0.6628.72 ± 2.30 **80.74 ± 0.61 **
MI28.30 ± 1.83 **60.66 ± 0.85 **5.70 ± 0.5027.73 ± 1.79 **78.37 ± 0.87 **
MICa6.63 ± 0.59 **35.29 ± 0.87 **5.43 ± 0.303.81 ± 0.60 **34.85 ± 2.83 **
MIMa8.42 ± 0.33 **34.06 ± 0.66 **5.54 ± 0.176.34 ± 0.29 **48.89 ± 0.44 **
L: brightness; a: green shades; b: yellow shades; h: saturation; C: color. Control: standard muffin; MI: muffin with inulin as a fat replacer; MICa: muffin with inulin (MI) + Calafate berries; MIMa: muffin with inulin (MI) + Maqui berries. ** statistically significant (p = 0.000) in analysis of variance.
Table 9. Descriptive analysis of the sensory evaluation performed by a group of people with Parkinson’s disease (n = 23).
Table 9. Descriptive analysis of the sensory evaluation performed by a group of people with Parkinson’s disease (n = 23).
SamplesAppearanceAromaTextureFlavorOverall ColorPurchase IntentionOverall Acceptability
Average ± SD
Control4.00 ± 0.743.74 ± 0.863.32 ± 0.994.00 ± 0.934.09 ± 0.923.36 ± 1.263.95 ± 0.72
MI4.04 ± 1.113.13 ± 1.253.39 ± 0.993.83 ± 1.114.09 ± 0.923.22 ± 1.443.61 ± 1.03
MICa3.26 ± 1.213.17 ± 1.073.18 ± 1.303.23 ± 1.383.26 ± 1.323.30 ± 1.433.61 ± 1.20
MIMa3.48 ± 1.343.13 ± 1.223.26 ± 1.103.39 ± 1.273.48 ± 1.383.39 ± 1.343.57 ± 1.20
Control: standard muffin; MI: muffin with inulin as a fat replacer; MICa: muffin with inulin (MI) + Calafate berries; MIMa: muffin with inulin (MI) + Maqui berries.
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García-Milla, P.; Peñalver, R.; Nieto, G. Development of Functional Muffins with Fruits of the Chilean Forest (Calafate and Maqui) and Supplemented with Prebiotic Fiber. Appl. Sci. 2024, 14, 7757. https://doi.org/10.3390/app14177757

AMA Style

García-Milla P, Peñalver R, Nieto G. Development of Functional Muffins with Fruits of the Chilean Forest (Calafate and Maqui) and Supplemented with Prebiotic Fiber. Applied Sciences. 2024; 14(17):7757. https://doi.org/10.3390/app14177757

Chicago/Turabian Style

García-Milla, Paula, Rocío Peñalver, and Gema Nieto. 2024. "Development of Functional Muffins with Fruits of the Chilean Forest (Calafate and Maqui) and Supplemented with Prebiotic Fiber" Applied Sciences 14, no. 17: 7757. https://doi.org/10.3390/app14177757

APA Style

García-Milla, P., Peñalver, R., & Nieto, G. (2024). Development of Functional Muffins with Fruits of the Chilean Forest (Calafate and Maqui) and Supplemented with Prebiotic Fiber. Applied Sciences, 14(17), 7757. https://doi.org/10.3390/app14177757

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