1. Introduction
Carotenoids are among the major classes of pigments found in tree leaves, fruits, and vegetables. Consumption of these compounds has been considered to bring health benefits. According to the literature, carotenoids may act as reducing agents for cancer [
1,
2,
3], cardiovascular diseases [
4,
5] and macular degeneration [
6], as well as antioxidants [
7] and provitamin A [
8]. The nutritive and coloring properties of carotenoids make them be an ideal food additive to develop functional products with desirable appearance.
The interest in recovering carotenoids from agro-industrial waste has increased, considering their high potential to enhance the valorization of by-products. Several studies [
9,
10,
11,
12] have reported the extraction of carotenoids from different vegetable wastes. However, guaraná residues, with potential uses in the production of soft drinks and food ingredient industries, have been rarely explored as a possible source of carotenoids in foods.
Carotenoids are highly susceptible to harsh external conditions, such as high temperature and shear, presence of oxygen, light, acidity, and pro-oxidant agents [
13,
14,
15]. Encapsulation has emerged as a key approach for making the incorporation of these compounds in processed foods feasible. In the food industry, spray drying is the most widely used technology for the entrapment of bioactive molecules in the form of microparticles. The technique consists of atomizing a dispersion or emulsion, containing the component of interest and drying adjuvants, followed by its dehydrating forming microparticles. This encapsulation facilitates transport, prolongs shelf-life, and reduces the risk of carotenoid degradation during food processing [
16,
17].
Several additives are used as carrier materials to facilitate the drying, handling and application of these bioactive compounds. In addition, the property of these carrier materials can guarantee the stability of carotenoids against oxidation, storage conditions and processing in the food industry. Tuyen et al. [
18] reported the effect of different concentrations of maltodextrin as carrier material and different inlet temperatures on color preservation, total carotenoid content and antioxidant activity of spray-dried gac powder. In addition, Hojjati et al. [
19] investigated the influence of different concentrations of soluble soy polysaccharides on the properties of microcapsules loaded with canthaxanthin obtained by spray drying. Indeed, the encapsulation of canthaxanthin in this study resulted in a more significant storage stability of the samples. Highlighting the variety of carrier materials, Etzbach et al. [
20] evaluated the effect of using different carriers such as maltodextrin, modified starch, inulin, alginate, gum arabic and cellobiose for spray drying golden blackberry juice rich in carotenoids. In this study, cellobiose’s proposed alternative carrier showed a high capacity to protect carotenoids from degradation processes by exposure to light, high temperature and oxygen, possibly due to a more compact particle wall and larger particle sizes.
The development of starchy products includes the investigation of new formulations and new processing techniques. However, typical technologies used to favor gelatinization and texturization of starchy products apply a combination of heat and shear over these products, which can be harmful to carotenoids. During gelatinization, starch swells, destabilizing its crystalline structure and loses birefringence. The main starch fractions, amylose and amylopectin, disperse and lead to the production of a paste [
21,
22]. After disruption of their granular structure, during processing, starch is subjected to mechanical stresses, mainly shear provoked by processing.
The rheological properties of starch solutions can vary according to factors such as amylose and amylopectin content, the presence of functional groups, and granularity. In the case of the current research, these factors can be influenced by adding microparticles loaded with carotenoids into the oat paste, which may change the paste’s rheological behavior. On the other hand, the processing of oat paste, which involves heat and shear conditions, can lead to the degradation of thermosensitive compounds such as carotenoids. From that, the proposal focused on understanding the effects of these conditions on microparticles rich in carotenoids, also evaluating the effectiveness of microencapsulation by spray drying. Oats have wide applications, and the carotenoids from the guarana by-product are still atypical but with great potential for exploration and use by the food industry.
To simulate a pasting environment where temperature and shear are applied, a rheometer equipped with a pasting cell was used to produce an oatmeal paste enriched with encapsulated carotenoids. Thus, this study was aimed to prepare a carotenoid-rich extract from guaraná peels, encapsulate this extract, investigate the influence of microparticles on the pasting properties of oatmeal, and evaluate the effect of thermo- and mechanical stress on carotenoid stability under different conditions.
2. Material and Methods
2.1. Materials
The Executive Commission for Cocoa Cultivation Planning, CEPLAC (Taperoá, Bahia, Brazil), provided the guaraná fruits. The fruit peels were removed from seeds and pulp followed by being washed with water. The reddish peels were dried in a convection oven (Marconi, MA035/1152) at 50 °C for 18 h [
23]. They were then milled and stored under dark at −20 °C until analysis.
β-carotene (CAS 7235-40-7) and lutein (CAS 127-40-2) standards were purchased from Sigma-Aldrich (Saint Louis, USA). Analytical-grade ethanol (CAS 64-17-5), hexane (CAS 110-54-3), and acetone (CAS 67-64-1) were obtained from Fisher Scientific (Waltham, MA, USA).
The sunflower oil used during the guaraná peel extract preparation was from Cargill (brand Liza). The microparticles rich in carotenoids produced by spray drying (SD) were prepared using gum arabic as carrier material, obtained from Nexira, Brazil. Oat flakes (the Oat Quaker Company, Chicago, IL, USA) were purchased from a local supermarket at Columbus, OH, USA.
2.2. Carotenoid-Rich Extract Preparation
Carotenoid-rich extract preparation from guaraná peels was performed following [
24], using ethanol as solvent, at a ratio of 1:10 (peel: solvent,
w/
v), during 4 h at 50 °C. The concentration of 3% of sunflower oil was added to the extract to reduce the carotenoid degradation detected during preliminary experiments. The final concentration was determined considering the liquid–liquid equilibrium for the mixture composed of oil and ethanol [
25]. As follows, the material was concentrated using a rotary evaporator (TE-211 Tecnal, Piracicaba, Brazil) at 48 ± 2 °C to 20% of the initial volume. The concentrated extract was named guaraná peel extract (GPE).
2.3. Microencapsulation by Spray Drying
Formulations were prepared with a ratio of 1:2, 1:3, and 1:4 of concentrated ethanolic extract:gum arabic in aqueous solution (20%
w/v),
v/
v, using Ultra-Turrax
® IKA T25 (Labotechnic, Staufen, Germany) at 11,200×
g for 3 min. The mixture was atomized according to Rocha et al. [
26], with modifications. The spray dryer (Model MSD 1.0, Labmaq do Brasil, Ribeirão Preto, Brazil) was used coupled with a spray nozzle of 1.2 mm, with an inlet air temperature of 100 °C, an air drying speed of 2.5 m/s, a feed flow of 10 mL/min, and air pressure of 8.4 kgf/cm
2.
2.4. Total Carotenoid Content
The microparticles were blended with hexane for 1 min and ultrasonicated for 20 min using an ultrasound bath, Branson 1800 (Branson Ultrasonics Corporation, Danbury, CT, USA) to extract carotenoids. Absorbances of extracts were measured at 450 nm wavelength and recorded using a UV–visible spectrophotometer (Thermo Scientific, Waltham, MA, USA, Genesys 10S). The β-carotene standard was used for quantification, and the results were expressed as µg β-carotene/g sample [
27]. The test was conducted in triplicate.
The retention after encapsulation was calculated as a ratio of the total carotenoid amount in the microparticles to that in the feed materials before atomization.
2.5. Stability Study of Microparticles
The samples were placed in glass vials covered with aluminum foil and stored in desiccators containing saturated solutions of magnesium chloride MgCl
2 (relative humidity, RH of 32.8%). The desiccators were kept at a temperature of 25 °C, and the storage period lasted for 90 days under the specified conditions [
28]. The samples were evaluated every 15 days in terms of carotenoid content and color. Particle size distribution, mean diameter, moisture content, and water activity were analyzed at the initial time and after 90 days of storage, in triplicate.
2.5.1. Carotenoid Degradation Kinetics
Carotenoid stability was determined by comparing the total carotenoid concentration at the initial time and over the storage of encapsulated and non-encapsulated carotenoid-rich GPE. Previous studies [
29,
30] have hypothesized that a first-order kinetics describes adequately the reaction of carotenoids degradation. Thereby, to investigate the stability of our samples, the degradation constant (
k) and the half-life (t
1/2) were determined following the first-order kinetic model, according to Equations (1) and (2).
where
C = carotenoid concentration at time
t (μg/g);
C0 = initial carotenoid concentration (μg/g);
t = time (days).
2.5.2. Color
The microparticles color defined by the parameters L* (Brightness), a* (red–green), and b*(yellow–blue) were determined using a HunterLab Mini Scan XE colorimeter (Reston, VA, USA).
Chroma (color saturation) was calculated according to the Minolta procedure [
31]:
The total color difference (Δ
E) was calculated according to Equation (4).
where
= total color difference;
= final L*;
= initial L*;
= final a*;
= initial a*;
= final b*;
= initial b*.
2.5.3. Mean Diameter
Mean diameter of particles were measured using the SALD-201V laser diffraction particle analyzer, Shimadzu (Kyoto, Japan). Ethanol was used as a dispersing liquid. The measurements were conducted at 25 °C.
2.5.4. Moisture Content and Water Activity
The moisture content of the microparticles was determined in a moisture analyzer model MB 35 from Ohaus (Parsippany, OH, USA), in triplicate. The results are expressed in percentage. The determination of water activity (aw) was achieved by direct reading, on an Aqualab hygrometer, model CX-2T, from Decagon Devices Inc., Pullman, WA, USA. The readings were performed at 25 °C.
2.6. Dynamic Vapor Sorption (DVS) of Microparticles and Oat Flakes
Water vapor sorption isotherms of the microparticles and oat flakes were determined at 25 °C using a Dynamic Vapor Sorption instrument (Surface Measurement Systems Ltd., Allentown, PA, USA). Under a continuous airflow (200 mL/min), the system was pre-equilibrated at 5% relative humidity (RH). The samples were exposed sequentially to different relative humidities (RH) from 30 to 95%. The RH transitions and mass variations of the sample were monitored continuously. The moisture sorption isotherms were determined using the DVS Analysis Macro V6.1 software, in duplicate.
2.7. Thermal Properties of Oat Flakes and Oat Flakes Containing Microparticles
The thermal behavior of oat flakes and oat flakes containing microparticles was described by the parameters ‘onset of gelatinization’ (To), ‘peak gelatinization temperature’ (Tp), and ‘gelatinization enthalpy’ (J/g) and measured by a Multi-Cell Differential Scanning Calorimeter (MC-DSC, TA Instruments, New Castle, DE, USA) equipped with the TRIOS software (TA Instruments, New Castle, DE, USA). The samples dispersed in water (with a moisture content of 80%) were weighed into ampoules and sealed. An empty pan was used as the reference. The samples were equilibrated at 5 °C and then heated to 140 °C at a rate of 1 °C/min. Each sample was run in triplicate, and the average results are shown.
2.8. Pasting Properties
The oatmeal paste was prepared using the Discovery Hybrid Rheometer 3 (DHR-3, TA Instruments Ltd., New Castle, DE, USA) and a pasting cell geometry to simulate operating conditions where temperature and shear are applied during the process. The experiment was conducted by mixing (i) oat flakes and distilled water (80% w/w); (ii) oat flakes blended with microparticles (6% w/w), and distilled water (80% w/w); (iii) oat flakes, GPE (6% w/w), and distilled water (80% w/w).
The samples were prepared using the following procedure: (1) conditioning the sample at 25 °C for 2 min; (2) heating ramp to selected temperatures of 70, 80 or 90 °C at 5 °C/min with a shear rate of 50 or 100 1/s; (3) flow peak hold at the selected temperature and shear conditions for 120 s; (4) cooling ramp to 25 °C at 5 °C/min; (5) oscillation frequency at 25 °C from 0.01 to 10 Hz with a 0.5% strain. The testing parameters and treatments are shown in
Table 1. Each sample was run in triplicate.
2.9. Retention of β-Carotene, Lutein, and Total Carotenoid Content in the Oatmeal Paste
Total carotenoid content of oatmeal paste containing encapsulated and free GPE was analyzed using spectrophotometry, as described in
Section 2.4. The main carotenoids extracted from guaraná peels were β-carotene and lutein [
24]. The contents of the incorporated carotenoids were quantified in the oatmeal paste and the raw material by HPLC to evaluate the effect of the process on the retention of these compounds, in duplicate. Carotenoid extraction was carried out following Kopec et al. [
32], with some modifications according to the solvents used. Sequential extractions were performed using methanol and a mixture of ethanol: acetone: hexane (1:1:1,
v/
v/
v). The extract was then injected into the HPLC instrument Agilent 1260 ultra-high-performance liquid chromatograph with a diode array detector (UHPLC-DAD), using a C30 column (YMC Inc., Meridian, ID, USA, 4.6 × 250 mm, 3 μm) [
33]. The contents of β-carotene and lutein were calculated from their peak areas in comparison to standards with known concentration, using a calibration curve. Carotenoid retention was determined by comparing their content before and after the pasting process and expressed as a percentage.
2.10. Statistics
The data were analyzed using ANOVA and Tukey’s test in SAS statistical software (version 8.02, Statistic Analysis System). Significant differences were defined at p < 0.05. All data were expressed as the means ± standard deviation (SD). The pasting process was performed in duplicate.
4. Conclusions
Microencapsulation of carotenoid-rich extract from guaraná peels enhanced its stability over storage. The sample SD1:2 exhibited promising features among other formulations, including high carotenoid content, suitable particle size, and intense color. The addition of microparticles rich in bioactive compounds into oatmeal paste increased the onset and peak temperature of starch but decreased its enthalpy. Further, it reduced the viscosity of the system, whereas the opposite trend was observed for the samples with free GPE. Overall, the gelatinization of oat flakes starch was affected by the decrease in accessible water, due to the presence of the microparticles.
Testing conditions, such as temperature and shear, favored heat transfer and oxygen incorporation in the system, reducing the total carotenoid content of the samples. In addition, the treatment significantly altered the lutein content, when compared to β-carotene. However, pigment loss decreased by encapsulation.
The recovery of bioactive components from by-products, such as β-carotene (provitamin A) and lutein (recognized as an agent that prevents macular degeneration), reveals the importance of the present research on adding value to a waste of a food process encouraging sustainable development. In addition, the study of applications of these materials, as suggested in the current work, represents a suitable trend for obtaining functional foods. The range of oat products and derivatives is wide, including oatcake, oatmeal, and porridge, which can be enriched with bioactive compounds to add health benefits. In addition, the use of oats as an ingredient in newer sectors, such as plant proteins, has grown and has opened up possibilities for applications and, consequently, new processes.