Abstract
Iron is a micronutrient that is important to humans, and its deficiency is considered a public health issue. Through this study, we were able to confirm the importance of microencapsulating iron sulphate through complex coacervation using sodium alginate and soy protein isolate as the wall material and its application in a food matrix. The phase diagram results demonstrate a strong affinity between these biopolymers, achieving an encapsulation efficiency of over 90% at a 1:1 ratio. The iron release profile from the microcapsules was as expected, with enhanced absorption in the intestinal phase and an overall accessibility of 59.05%. When the iron microcapsules were incorporated into a cookie, subsequent analysis revealed no significant differences in colour or texture compared with those of the control sample. These findings suggest that iron fortification via microencapsulation can be effectively integrated into widely accepted food products, such as cookies.
Introduction
According to the World Health Organization, iron deficiency anaemia is the most common form of anaemia, accounting for 90% of cases. In Brazil, anaemia affects children (up to 5 years old), women of childbearing age, and pregnant women. Approximately, 20.9% of Brazilian children and 29.4% of Brazilian women are affected, according to the Ministry of Health. The main characteristic of iron deficiency anaemia is low haemoglobin levels in the blood, resulting in the inability to transport oxygen through red blood cells to meet the physiological needs of the body (Brazil, 2004, 2006). Iron deficiency affects the global population, with an estimated 2–3 billion people affected by this condition worldwide. When iron supplementation is necessary, it must be administered at high doses, which often leads to side effects, such as unpleasant taste, odour, tooth darkening, intestinal issues, and interactions with other nutrients. Therefore, microencapsulation offers a promising alternative to reduce some of these side effects in individuals taking iron supplements (Dehnad et al., 2023).
In the food industry, microencapsulation is used to protect and store bioactive ingredients. The main objectives of this method include improving the stability and efficacy of bioactive compounds, reducing interactions with environmental factors, preventing sensory and nutritional losses, masking unpleasant flavours, and prolonging the shelf life of encapsulated materials (Abbas et al., 2023; Bastos et al., 2020; Pereira et al., 2018; Razavizadeh & Yeganehzad, 2024). According to Koohenjani and Lashkari (2022), the use of a double emulsion with iron does not impact the properties of creams compared with those containing free iron. The sensory characteristics remained unchanged, as did consumer acceptability. The double water/oil/water emulsion improved iron bioavailability by protecting the iron and allowing for controlled release in the small intestine.
One of the microencapsulation methods described in previous research is complex coacervation, which is the focus of this study. This process can be divided into three steps: the first step is emulsification, which can involve either a simple or double emulsion depending on the properties of the bioactive compound; the second step is coacervation; and the third step is crosslinking, which forms the outer shell (Yan & Zhang, 2014). Numerous previous studies have reported the use of complex coacervation; e.g., Silva et al. (2019) and Zhang et al. (2021) encapsulated different probiotics via complex coacervation, Farias et al. (2023) encapsulated β-carotene, and Bastos et al. (2020) encapsulated black pepper oil. Recent work by Barbosa and Garcia-Rojas (2022) and Koohenjani and Lashkari (2022) described the encapsulation of iron via a double emulsion without specific applications. However, no studies have reported the encapsulation of iron via the complex coacervation process.
Thus, this study explored the formation of a complex between soy protein isolate (SPI) and sodium alginate (NaAlg) for the encapsulation of iron sulphate and its use in fortifying cookies.
Materials and methods
Materials
SPI used was purchased from Relva Verde Alimentos EIRELI (São Paulo, Brazil). Iron sulphate heptahydrate (P.A., 215422), NaAlg, porcine pepsin (P6887), porcine pancreatin (P7545), and an extract of porcine bile (B3883) were also obtained from Sigma–Aldrich (St. Louis, USA). Polyglycerol polyricinoleate (PGPR) was obtained from Ingredients Online. Corn oil was purchased from a local supermarket in Volta Redonda, RJ. Ultrapure water (Master System R&D, Gehaka, Brazil), with a conductivity of 0.05 μS/cm, and P.A. grade reagents were used in all tests.
Heat treatment and dialysis of SPI
To improve protein solubility, the following analyses were conducted: heat treatment and dialysis of SPI. These analyses were carried out following to the methodology described by O’Flynn et al. (2021). A solution of SPI (5 mg/ml) was prepared, and the pH was adjusted to 9.0 with 0.5 mol/L NaOH. This solution was placed on a magnetic stirrer (NT 101, Novatecnica, Brazil) for 1 hr and then transferred to a thermal bath (Microprocessor Control MPC, Huber, Germany) at 90 °C for 20 min. Immediately afterwards, the mixture was centrifuged (Digicen 21 R, Oltoalresa, Spain) at 8000 × g for 10 min at room temperature. After this first process, the supernatant was removed and distributed onto dialysis membranes (D9527-100FT, Sigma–Aldrich, USA) to separate the protein from other solutions smaller than 12 kDa. SPI was subsequently freeze-dried (Terroni, Enterprise I, Brazil) for 48 hr.
Determination of ζ-potentials
The ζ-potentials of the biopolymers (SPI and NaAlg) were determined via a Zetasizer (Malvern Instruments, Nano ZS90, UK). The pH of the solutions was adjusted based on the results obtained by placing the samples at 0.1% (wt/wt) in an MPT-27 autotitrator (Malvern Instruments, UK). The analyses were conducted across pH values ranging from 2.0 to 9.0 at 25 °C. The strength of the electrostatic interaction (SEI) between oppositely charged PEs was estimated according to Equation 1:
where ZP1 and ZP2 are the absolute zeta potentials of the polymers at a given pH.
Influence of biopolymer concentration on the formation of coacervate complexes
Stock solutions of 1.5% (wt/wt) SPI and 1.5% (wt/wt) NaAlg were prepared using ultrapure water containing 0.02% (wt/wt) sodium azide. A total of four subgroups were prepared with concentrations of 0.1%, 0.5%, 1.0%, and 1.5% (wt/wt) at pH 4.0 using different ratios (1:1, 2:1:4:1, 6:1, 8:1, and 10:1) of total biopolymer concentrations and various proportions of wall material. These samples were evaluated after 48 hr at 4 °C, following the methodology proposed by Lan et al. (2021). The phase diagrams of the complexes formed by the proteins and polysaccharides in different proportions and total concentrations of biopolymers are as follows: □ cloudy solution with a precipitate, ■ cloudy solution, |$\circ $| solution with a smaller volume of precipitate, and ● solution with a larger volume of precipitate that is slightly cloudy.
Encapsulation of iron sulphate
The wall materials were used at concentrations of 0.5%, 0.75%, and 1.0% (wt/wt) with core-to-wall material ratios of 1:1, 2:2, and 4:1, respectively. Initially, water-in-oil (W1/O) emulsions were prepared using corn oil and PGPR at a concentration of 2% (wt/wt), along with different concentrations of the iron sulphate solution. Emulsification was performed at 12,500 rpm for 4 min using an Ultraturrax (IKA, T25D, Germany). In the second stage, to obtain the microcapsules, double emulsions (W1/O/W2) containing the SPI and NaAlg solutions were prepared. At this stage, the pH of the SPI and NaAlg solutions was adjusted to 4.0 via acetic acid at concentrations of 20% and 10% (vol/vol) with the aid of a benchtop pH meter, after which the solutions were subjected to and homogenised with UltraTurrax at 10,000 rpm for 3 min. The system was subsequently placed in an ice bath for 1 hr. To induce cross-linking, 5 ml of CaCl2 (30 mg/ml) was added, and the mixture was refrigerated at 10 °C for 24 hr. Later, the supernatant was removed, and the obtained precipitates were frozen in liquid nitrogen and dried in a freeze dryer for 48 hr.
Encapsulation efficiency
The amount of iron found in the external aqueous phase (W2) of the emulsion was measured via a Biomate 3S spectrophotometer (Thermo Fisher Scientific, USA). A total of 0.5 ml of the double emulsion was mixed with 0.55 ml of ferrozine and shaken for 30 min at room temperature after being centrifuged at 10,000 rpm for 10 min at 10 °C. The absorbance was measured at 562 nm post solution shaking. The encapsulation efficiency was determined via Equation 2:
where “Amax” is when all iron has been released, “At” is the absorbance of the emulsion of the external aqueous phase (W2) at a given time, and ‘A0’ is the absorbance of the emulsion without iron sulphate (Prichapan et al., 2021).
Fourier transform infrared spectroscopy
The samples were analysed to obtain infrared spectra. These analyses were conducted at room temperature via a Fourier transform infrared (FTIR) spectrometer (Vertex 70, Bruker, Germany), and the results were recorded in the range of 4,000–500 cm−1.
Optical microscopy
The microencapsulation process was monitored via an optical microscope; an aliquot of the fresh sample was placed between the slide and the coverslip and observed through an optical microscope (K220, Kasvi, Brazil) coupled to a Moticam camera (5 MP, Kasvi, Brazil).
In vitro gastrointestinal simulation of microcapsules
The in vitro gastrointestinal simulation was based on the INFOGEST methodology (Brodkorb et al., 2019). During the process, 0.2 ml samples were collected from the gastric (30, 60, 90, and 120 min) and intestinal phases (150, 180, 210, and 240 min). Digestion was carried out with 0.3 mg of iron microcapsules and 1 ml of ultrapure water. In the oral phase, 1.04 ml of salivary fluid at pH 7, 0.13 ml of salivary amylase and 6.5 μl of calcium chloride were added to 1,300 g of sample. The samples were subsequently shaken at 100 rpm in a shaker (Tecnal, TE-424, Brazil) for 2 min at 37 °C. After oral digestion, 2.008 ml of gastric fluid was added to the final volume of the oral phase, and the pH of the solution was adjusted to 3.0 with a bench pH meter with hydrochloric acid (6 M HCl). In addition, 1.3 μl of calcium chloride and 0.26 ml of porcine pepsin were added, and the mixture was shaken at 100 rpm in a shaker (Tecnal, TE-424, Brazil) for 2 hr at 37 °C. In the intestinal phase, 2.21 ml of intestinal fluid was added, the pH was adjusted again to 7 with sodium hydroxide (6 M NaOH), and then 10.4 μl of calcium chloride, 1.3 pancreatin, and 0.65 ml of bile were added. The bile solution was dissolved in intestinal fluid at a concentration of 10 mM. Pancreatin was also solubilized in the intestinal fluid at a concentration of 100 U/ml. The intestinal phase was shaken at 100 rpm in a shaker (Tecnal, TE-424, Brazil) for 2 hr at 37 °C.
Bioaccessibility of iron microcapsules
Bioaccessibility was measured after simulating the digestion process. The collected samples were centrifuged at 16,000 rpm for 30 min at 4 °C; the supernatant obtained after centrifugation represented the micellar portion. The extraction and quantification of ferrous sulphate were conducted to assess the encapsulation efficiency. The bioaccessibility (B) of iron sulphate was calculated via Equation 3:
where CDigesta is the iron sulphate concentration in the total digestion and CMicellar is the iron sulphate concentration in the micellar part.
Iron release
Iron was released according to Prichapan et al. (2021). The amount of iron found in the external aqueous phase (W2) of the emulsion was measured via a Biomate 3S spectrophotometer. A total of 0.5 ml of the double emulsion (centrifuged at 10,000 rpm for 10 min at 10 °C) was mixed with 0.55 ml of ferrozine and shaken for 30 min at room temperature. After shaking, the absorbance was measured at 562 nm. The release of iron was calculated via Equation 4:
where “Amax” is when all iron has been released, “At” is the absorbance of the emulsion of the external aqueous phase (W2) at a given time, and “A0” is the absorbance of the emulsion without iron sulphate.
Formulation and enrichment of cookies with iron sulphate microcapsules
Cookies were used as a food matrix for the incorporation of iron microcapsules. Three cookie formulations were prepared: (M1) without the addition of microcapsules or free iron, (M2) with the addition of free iron, and (M3) with the addition of iron microcapsules. To prepare the cookies, margarine, sugar, salt, and sodium bicarbonate were mixed for 3 min. The microcapsules and wheat flour were subsequently added, as shown in Supplementary Table 1. Finally, water was added, and the mixture was homogenized for 2 min. The cookies were shaped into petri dishes 60 mm in diameter and 15 mm thick. During baking, the cookies were heated at 170 °C in an electric oven (Mondial, FR-20, Brazil) for 10 min and then cooled to room temperature before analysis (Farias et al., 2023).
Texture analysis: hardness and fracturability
Texture analysis was conducted 24 hr after cookie processing via a TA-XTPlus texturometer (Stable Micro Systems, Surrey, UK). The probe used was 2 mm long (Stable Micro Systems, 2 mm Cyl. Stainless, Number 12239—P/2) on an HDP/90 platform. The following parameters were set for the test: pretest speed = 1.0 mm/s, test speed = 0.5 mm/s, posttest speed = 10.0 mm/s, and distance = 10 mm, and the compression force was measured. The texture results are reported in Newtons (N) and millimetres (mm), respectively, representing the arithmetic mean of 5 determinations across three samples from the same experiment, following the methodology of Filipčev et al. (2014).
Colour analyses
A colorimeter (MiniScan EZ, Hunterlab, USA) was used for colour analysis, and the analysis was performed in decaplica (n = 10). The samples were analysed via the CIE-L*a*b* System (Commision Internationale L'Eclairage), and the luminosity (L*), red intensity (+a*), and yellow intensity (+b*) parameters were obtained according to the methodology of Gusmão et al. (2018).
Statistical analysis
Data evaluation was carried out via Tukey analysis and analysis of variance via the SISVAR 5.6 program, with a significance level of p < .05. The experiments were performed in triplicate, and the results are presented as the means ± SDs.
Results and discussion
Formation of the coacervate complex: zeta potential
Figure 1 shows the ζ-potential and SEI values of the SPI and NaAlg biopolymers as a function of pH. At the values studied (2.0–5.0), the polymers have opposite charges up to pH 5.0: SPI positive and NaAlg negative. At pH 5.5, we observe the isoelectric point (pI) of the protein, where its value is equal to zero. For the formation of the SPI—NaAlg complex, the pKa value of NaAlg (pKa 3.38) and the pI of the protein influence. The highest SEI value was observed at pH 4.0, which is close to the pKa value of NaAlg, and the pH and pKa values are close to each other; these values contribute to the ability of the carboxylic groups of NaAlg to gain hydrogen (protonation), balance the protonated molecules and deprotonate, which favours the self-aggregation of biopolymers (Farias et al., 2023). The pH chosen for the formation of coacervate complexes between the SPI and NaAlg biopolymers was 4.0 due to the interaction observed between them through the ζ-potential.
ζ-potential and electrostatic interaction force (SEI) of soy protein isolate and sodium alginate.
Formation of coacervate complexes: phase diagram
The phase diagram (Figure 2) shows the formation of a coacervate complex of SPI and NaAlg at different concentrations of biopolymers and ratios. In the diagram, concentrations are between 0.1% and 1.5% (wt/wt) and ratios between 1:1 and 10:1; initially, we observed that the lower the ratio and the lower the concentration are, the less turbidity and slight precipitation there are. When the concentration of biopolymers was increased and the ratio was maintained at 1:1, a greater volume of precipitate and slight turbidity was observed. With the phase diagram, it is possible to evaluate the best pH range and limit for the coacervated complexes of SPI and NaAlg, Lan et al. (2021) and Constantino and Garcia-Rojas (2023). A ratio of 1:1 (SPI:NaAlg) and a concentration of 1.0% biopolymers were chosen to continue the work, since there were no changes in the percentage of biopolymers to 1.5%, and the biopolymer ratio (1:1) resulted in the best volume formation of the coacervate complex.
State diagram of complexes formed by concentrations of soy protein isolate (SPI) and sodium alginate (NaAlg) in different ratios (1:1 and 10:1) and total concentration of biopolymers at pH 4.0. □ represents a cloudy solution with less precipitate, ■ represents a solution with light precipitate without turbidity, |$\circ $| represents a solution with smaller volume precipitate, and ● represents a solution with a greater volume of precipitate and slightly turbid.
Encapsulation efficiency
The passage details the results of nine different treatments used for encapsulating iron, as summarized in Table 1. All of the treatments achieved encapsulation efficiencies >90%, with no significant differences observed among them. This efficiency is notably higher than the 60% threshold reported in other studies using complex coacervation, such as those by Bastos et al. (2020), Farias et al. (2023), and Soares et al. (2019).
Encapsulation efficiency of samples to encapsulate iron sulphate, with contractions of 1%, 0.75%, and 0.5% (wt/wt) and ratios of 1:1, 2:1, and 4:1 (SPI:NaAlg).
| Samples | Wall material | Iron (mg) | Core/wall material | EE (%) | ||
|---|---|---|---|---|---|---|
| SPI (g) | NaAlg (g) | Total | ||||
| A | 0.050 | 0.050 | 0.1 | 5.0 | 1:1 | 99.43a |
| B | 0.050 | 0.050 | 0.1 | 2.5 | 2:1 | 99.49a |
| C | 0.050 | 0.050 | 0.1 | 1.25 | 4:1 | 98.2a |
| D | 0.038 | 0.038 | 0.076 | 3.75 | 1:1 | 98.98a |
| E | 0.038 | 0.038 | 0.076 | 1.87 | 2:1 | 99.17a |
| F | 0.038 | 0.038 | 0.076 | 0.938 | 4:1 | 96.86a |
| G | 0.025 | 0.025 | 0.05 | 2.5 | 1:1 | 99.61a |
| H | 0.025 | 0.025 | 0.05 | 1.25 | 2:1 | 97.69a |
| I | 0.025 | 0.025 | 0.05 | 0.625 | 4:1 | 98.18a |
| Samples | Wall material | Iron (mg) | Core/wall material | EE (%) | ||
|---|---|---|---|---|---|---|
| SPI (g) | NaAlg (g) | Total | ||||
| A | 0.050 | 0.050 | 0.1 | 5.0 | 1:1 | 99.43a |
| B | 0.050 | 0.050 | 0.1 | 2.5 | 2:1 | 99.49a |
| C | 0.050 | 0.050 | 0.1 | 1.25 | 4:1 | 98.2a |
| D | 0.038 | 0.038 | 0.076 | 3.75 | 1:1 | 98.98a |
| E | 0.038 | 0.038 | 0.076 | 1.87 | 2:1 | 99.17a |
| F | 0.038 | 0.038 | 0.076 | 0.938 | 4:1 | 96.86a |
| G | 0.025 | 0.025 | 0.05 | 2.5 | 1:1 | 99.61a |
| H | 0.025 | 0.025 | 0.05 | 1.25 | 2:1 | 97.69a |
| I | 0.025 | 0.025 | 0.05 | 0.625 | 4:1 | 98.18a |
Note. In all treatments, a 5 g CaCl2 solution (30 mg/ml) was added. The superscript letters in the means are Tukey’s statistical results and their differences mean that the samples differ statistically (p < .05).
SPI = soy protein isolate; NaAlg = sodium alginate; EE (%) = encapsulation efficiency.
Encapsulation efficiency of samples to encapsulate iron sulphate, with contractions of 1%, 0.75%, and 0.5% (wt/wt) and ratios of 1:1, 2:1, and 4:1 (SPI:NaAlg).
| Samples | Wall material | Iron (mg) | Core/wall material | EE (%) | ||
|---|---|---|---|---|---|---|
| SPI (g) | NaAlg (g) | Total | ||||
| A | 0.050 | 0.050 | 0.1 | 5.0 | 1:1 | 99.43a |
| B | 0.050 | 0.050 | 0.1 | 2.5 | 2:1 | 99.49a |
| C | 0.050 | 0.050 | 0.1 | 1.25 | 4:1 | 98.2a |
| D | 0.038 | 0.038 | 0.076 | 3.75 | 1:1 | 98.98a |
| E | 0.038 | 0.038 | 0.076 | 1.87 | 2:1 | 99.17a |
| F | 0.038 | 0.038 | 0.076 | 0.938 | 4:1 | 96.86a |
| G | 0.025 | 0.025 | 0.05 | 2.5 | 1:1 | 99.61a |
| H | 0.025 | 0.025 | 0.05 | 1.25 | 2:1 | 97.69a |
| I | 0.025 | 0.025 | 0.05 | 0.625 | 4:1 | 98.18a |
| Samples | Wall material | Iron (mg) | Core/wall material | EE (%) | ||
|---|---|---|---|---|---|---|
| SPI (g) | NaAlg (g) | Total | ||||
| A | 0.050 | 0.050 | 0.1 | 5.0 | 1:1 | 99.43a |
| B | 0.050 | 0.050 | 0.1 | 2.5 | 2:1 | 99.49a |
| C | 0.050 | 0.050 | 0.1 | 1.25 | 4:1 | 98.2a |
| D | 0.038 | 0.038 | 0.076 | 3.75 | 1:1 | 98.98a |
| E | 0.038 | 0.038 | 0.076 | 1.87 | 2:1 | 99.17a |
| F | 0.038 | 0.038 | 0.076 | 0.938 | 4:1 | 96.86a |
| G | 0.025 | 0.025 | 0.05 | 2.5 | 1:1 | 99.61a |
| H | 0.025 | 0.025 | 0.05 | 1.25 | 2:1 | 97.69a |
| I | 0.025 | 0.025 | 0.05 | 0.625 | 4:1 | 98.18a |
Note. In all treatments, a 5 g CaCl2 solution (30 mg/ml) was added. The superscript letters in the means are Tukey’s statistical results and their differences mean that the samples differ statistically (p < .05).
SPI = soy protein isolate; NaAlg = sodium alginate; EE (%) = encapsulation efficiency.
Table 1 highlights that samples A, B, and G presented the highest encapsulation efficiency values. Despite these high efficiencies, there were no significant differences between the samples. The optimal proportion of wall to core material was identified as 1:1, which yielded the best results. However, variations in the amount of encapsulated iron were noted. Based on these findings, Sample A was selected for further work due to its higher iron content, which aligns with previous work, particularly that of Soares et al. (2019). This choice ensures that a greater quantity of iron is available in the microcapsule, optimizing the effectiveness of the encapsulation process.
Fourier transform infrared spectroscopy
FTIR is responsible for chemically characterising the materials used in the microcapsule. Figure 3 shows the results for samples of corn oil (CO), SPI, NaAlg, and the iron microcapsule (MIC). Through the bands, we were able to identify the characteristic groups of each wall material used. In the SPI, we observed the amide bands of Groups I, II, and III: hydrogen bonds (1632 cm−1), elongation of NH2 (1527 cm−1) and stretch bonding of C–N and N–H (1157 cm−1), which are characteristic of proteins. In alginate (NaAlg), bands can be observed at 1593 cm−1, which corresponds to carboxylic acid (–COO−). Additionally, the –CO bond belonging to the acid group (RCOOH) was observed at peak 1407 cm−1. These results support those of Farias et al. (2023) and Bastos et al. (2020). The spectrum of the microcapsule is the coacervate complex of SPI and NaAlg. Figure 3 shows the interaction between them and the characteristic bands of the amino groups and the carboxyl groups (SPI and NaAlg, respectively). These characteristics are indicative of the interaction and formation of the coacervate complex.
Fourier transform infrared of corn oil (CO), soy protein isolate (SPI), sodium alginate (NaAlg), and iron microcapsule (MIC).
Optical microscopy
Figure 4 shows the double emulsion (W1/O/W2) particles with iron. In this figure, we also verify that the ratio (1:1) chosen for manufacturing the microcapsules is effective. In another study that used a double emulsion (W1/O/W2), Choi et al. (2009) reported similar results.
Optical microscopy 40× objective of the double emulsion (W1/O/W2) containing iron using soy protein isolate and sodium alginate as wall material in a 1:1 ratio.
In vitro gastrointestinal simulation of microcapsules
The passage details the gastrointestinal simulation used to analyse the release of iron from microcapsules made of SPI and NaAlg. The digestion process begins in the oral phase, where salivary fluid and amylase at pH 7.0 are involved for approximately 2 min. However, as shown in Figure 5 no iron release occurs during this phase because the microcapsule remains intact. The release of iron starts in the stomach phase. Here, the pepsin enzyme begins to break down the SPI, initiating partial release of the iron. However, the presence of NaAlg, a pepsin inhibitor, prevents complete degradation of the microcapsule. In the intestinal phase, the microcapsule is fully broken down, and the remaining iron is released. The coacervate complex of SPI and NaAlg is further digested by trypsin in this phase, leading to complete release of the encapsulated iron, substantiating results reported in studies by Bastos et al. (2020) and Farias et al. (2023).
Release of iron encapsulated in soy protein isolate and sodium alginate microcapsules.
This study reported a bioaccessibility of iron of 59.05%, which is a measure of the amount of iron available for absorption during digestion. Iron absorption occurs primarily in the duodenum of the small intestine and is influenced by its chemical form and antinutritional factors. The bioaccessibility found in this study aligns with values reported in the literature, such as those reported by Barbosa and Garcia-Rojas (2022) at 49.54% and those reported by Buyukkestelli and EL (2019), which range between 41.17% and 52.97%.
Texture and colour analysis of the cookies
The passage describes the results of hardness and fracturability analyses for cookies fortified with iron microcapsules. According to the data presented in Supplementary Table 2, the addition of microcapsules did not significantly affect the texture of the cookies, as confirmed by ANOVA and Tukey’s test. This finding suggests that incorporating microcapsules does not alter the hardness or fracturability of the cookies. This supports the findings of Filipčev et al. (2014), who reported similar results in gingerbread cookies, where microcapsule addition led to comparable or lower hardness values.
The passage also discusses colour analysis results, presented via the CIE L*a*b* colour model, which evaluates colour perception on the basis of human vision. The values for L* (lightness), a* (red–green axis), and b* (yellow–blue axis) are positive, indicating a greater intensity of yellow (b*) than red (a*) across all samples, regardless of microcapsule addition. The predominance of yellow suggests that the visual appearance remains consistent. The L* value represents luminosity, whereas b* and a* indicate the intensities of yellow (+) vs. blue (−) and red (+) vs. green (−), respectively. These interpretations are consistent with the findings of Popov-Raljić et al. (2013), which supports the stability of cookies’ visual properties even with microcapsule fortification.
Conclusion
In conclusion, this study has successfully demonstrated the potential of using SPI and NaAlg for iron encapsulation, as confirmed by phase diagram analysis and a high encapsulation efficiency exceeding 90% with a 1:1 wall material ratio. The in vitro gastrointestinal simulation further validated the controlled release of encapsulated iron, particularly in the intestinal phase, indicating that the microencapsulation system effectively protects and delivers iron where it is most needed, with a release rate above 50%. When incorporated into cookies, the addition of microcapsules did not significantly affect the texture or colour of the final product, as confirmed by texture and colour analyses.
The lack of significant differences between the control samples and those with microcapsules suggests that this fortification method can be seamlessly integrated into popular food products. Given that cookies are known worldwide, iron microencapsulation provides a practical and effective approach to food fortification, potentially contributing to the broader goal of addressing iron deficiency.
Data availability
Data are available on request from the authors.
Author contributions
Marina Stogmüller Ferreira da Silva (Conceptualization, Methodology, Validation, Investigation, Writing—original draft, Writing—review & editing), Bruno Sérgio Toledo Barbosa (Methodology, Validation, Investigation, Writing—original draft, Writing—review & editing), and Edwin Elard Garcia-Rojas (Conceptualization, Methodology, Resources, Writing—review & editing, Supervision, Project administration, Funding acquisition).
Funding
The authors thank the CNPq, Brazil (313928/2021-5); CAPES, Brazil (Código 001) and FAPERJ, Brazil (E-26/210.052/2023; E-26/201.030/2021) for the financial support.
Conflicts of interest
All authors declare that they have no conflict of interest.
Acknowledgements
The authors would like to express their gratitude for the technical support provided by Dr. Paulo Cezar da Cunha Júnior (UFRRJ) in the colour analysis of the samples.
Ethics statement
Ethics approval was not required for this research.
