Childhood represents a critical period for physical and cognitive development, laying the foundation for lifelong health and well-being. Among the myriad factors influencing childhood development, oral health and cognitive abilities hold particular significance. ¹ Poor oral health, exemplified by conditions such as dental caries, can lead to pain, discomfort, and difficulty in chewing, all of which may interfere with a child’s ability to concentrate and perform academically. Simultaneously, early brain development is highly sensitive to nutritional inputs, making it essential to explore strategies that address both oral health and cognitive growth synergistically.²

Recent advances in the fields of functional foods and nutraceuticals offer promising solutions to tackle these dual challenges. Probiotic strains, such as LLSPL-Bacillus coagulans, have been extensively studied for their ability to promote a balanced oral microbiome by suppressing harmful pathogens like Streptococcus mutans while encouraging the growth of beneficial bacteria.³ These beneficial effects contribute to the prevention of dental caries and the maintenance of overall oral health. Similarly, essential oils, such as cinnamon, clove oil and peppermint oil, are recognized for their antimicrobial and anti-inflammatory properties, further enhancing oral hygiene by targeting bacterial colonies and reducing oral inflammation.⁴

Beyond oral health, cognitive development during childhood is influenced by essential nutrients like docosahexaenoic acid (DHA), an omega-3 fatty acid integral to brain function. DHA plays a crucial role in neurogenesis, synaptic plasticity, and the maintenance of cell membranes in the brain. Numerous studies have demonstrated that adequate DHA intake is associated with improved memory, learning, and attention in children. However, ensuring consistent and adequate DHA consumption can be challenging, particularly for children with dietary limitations or preferences that exclude traditional sources like fatty fish.^5-7

Phosphatidylserine is a phospholipid that plays a critical role in maintaining the structural integrity of cell membranes, particularly in neurons. It supports cognitive functions by enhancing neurotransmitter activity and promoting efficient communication between brain cells. Research suggests that Phosphatidylserine supplementation may improve memory, attention, and overall cognitive performance, especially in aging individuals.⁸

This study aims to introduce and evaluate a novel candy formulation designed to combine these three bioactive ingredients—LLSPL-Bacillus coagulans, essential oils (cinnamon, clove oil and peppermint oil), and DHA, Phosphatidylserine (PS)—into a single, standardized, and child-friendly delivery system. The candy is formulated to target oral pathogens effectively, thus promoting oral health, while simultaneously providing a sufficient concentration of DHA to support cognitive development. The dual-purpose nature of this candy offers a unique opportunity to integrate oral care and brain health into a convenient, non-invasive, and enjoyable intervention for children.

The study focuses on children aged 4 to 12 years, a critical developmental stage characterized by rapid cognitive growth and susceptibility to oral health issues. It seeks to evaluate the impact of this candy on cognitive performance, through assessments of memory, attention, and learning.

By addressing these interrelated aspects of child health, this research not only provides valuable insights into the efficacy of combining probiotics, essential oils, and DHA in a single formulation but also contributes to the broader understanding of how functional foods can be leveraged to promote holistic health outcomes in children. If successful, this approach could pave the way for innovative solutions to improve both oral health and cognitive development, offering significant benefits for children and their families.

This study was designed to evaluate the impact of a novel candy formulation containing LLSPL-Bacillus coagulans, essential oils (cinnamon,  clove oil and peppermint oil), and DHA on the cognitive abilities of children aged 4 to 12 years. A structured methodology was developed to ensure the reliability and validity of the findings, as described below:

Study Design and Participants

This was a prospective, longitudinal study conducted over four months. Children aged 4 to 12 years were recruited from schools, following inclusion and exclusion criteria. Inclusion criteria required participants to be healthy children with no known cognitive impairments or medical conditions affecting oral or neurological health. Exclusion criteria included children with severe food allergies, ongoing treatments involving probiotics or omega-3 supplements, and unwillingness to participate.

Cognitive Assessment

To assess cognitive abilities, a test questionnaire based on the Montreal Cognitive Assessment (MoCA)⁹ framework was developed. The questionnaire included age-appropriate items adapted for younger populations, targeting domains such as memory, attention, visuospatial abilities, language, and executive functions. The test was validated by a multidisciplinary team comprising specialists in psychiatry and neurology to ensure its relevance and appropriateness for the target age group. The cognitive assessments were conducted by professionals certified in administering MoCA tests, ensuring standardized evaluation procedures.

Study Procedure

1. Baseline Assessment: At the start of the study, all participants underwent a baseline cognitive assessment using the validated MoCA-based questionnaire. This served as the reference point for evaluating changes in cognitive performance over the study period.
2. Intervention: Participants were provided with the candy formulation containing LLSPL-Bacillus coagulans, cinnamon,  clove oil, peppermint oil, and DHA. The candy was standardized to deliver consistent quantities of these active ingredients daily. Children were instructed to consume the candies under parental supervision to ensure adherence.
3. Follow-Up Assessments: Follow-up cognitive assessments were conducted at 2 months and 4 months after the baseline evaluation using the same MoCA-based questionnaire. The assessments aimed to measure changes in cognitive performance over time.

Data Collection and Analysis

• Primary Outcome: Changes in cognitive test scores across baseline, 2 months, and 4 months.
• Secondary Outcome: Participant adherence to the intervention and any reported side effects or adverse reactions.
• Data Handling: All data were recorded in structured case report forms and entered into a secured database. Descriptive statistics were used to summarize participant demographics and baseline characteristics. Changes in cognitive scores over time were analyzed using repeated-measures ANOVA to determine the statistical significance of differences between time points.
• Validation and Quality Control: The study adhered to standardized protocols for cognitive testing and data collection. All MoCA-certified professionals involved in the assessments underwent periodic training and inter-rater reliability checks to maintain consistency.

Ethical Considerations

The study was approved by the institutional ethics committee. Written informed consent was obtained from parents or guardians, and assent was taken from participating children. Participants were assured of their right to withdraw from the study at any time without consequences. Confidentiality of participant data was maintained throughout the study. This methodology ensured a robust evaluation of the intervention's effects on cognitive abilities, providing valuable insights into the potential of the candy formulation to enhance both oral health and cognitive development in children

Table 1: Descriptive Statistics for Age Distribution

Table 1 summarizes the key descriptive statistics for the age distribution of the study participants. It includes the number of observations (N), minimum age, maximum age, mean age, and standard deviation. The data set comprises 100 children aged between 4 and 12 years, with an average age of 7.66 years. The standard deviation of 1.976 indicates moderate variability in ages within the sample.

Table 2: Gender Distribution of Study Participants

Table 2 presents the gender distribution of the study participants. Among the 100 participants, 40% (40 individuals) are male, while 60% (60 individuals) are female. The total percentage sums up to 100%, reflecting the complete representation of the sample

Table 3: Comparison of Cognitive Parameters Between Male and Female Participants at Baseline

Table 3 compares the cognitive performance of male and female participants at baseline across several cognitive domains: Executive Functions & Visuospatial Skills, Language & Naming, Memory, Attention & Concentration, Abstraction, Orientation, and Total Score. For each parameter, the mean, standard deviation (SD), and standard error of the mean (SEM) are provided for both genders. The analysis includes the results of ANOVA tests, which assess the significance of any differences between males and females for each cognitive parameter. The F-values and p-values indicate whether any observed differences are statistically significant.

The findings show that males and females have similar scores in all cognitive domains. For example, males scored slightly higher on Executive Functions & Visuospatial Skills (mean = 3.60) compared to females (mean = 3.48), but the difference was not statistically significant (F = 3.957, p = 0.491). Similarly, while females scored slightly higher on Language & Naming (mean = 4.53) compared to males (mean = 4.40), this difference was also not significant (F = 1.479, p = 0.227). In other parameters such as Memory, Attention & Concentration, Abstraction, and Orientation, the scores were very similar between genders, with no significant differences found (all p-values > 0.05). The Total Score, a cumulative measure, also showed no significant difference between the genders (mean = 21.00 for males and mean = 21.15 for females, F = 0.059, p = 0.809).

In conclusion, the analysis indicates that there are no statistically significant gender differences in cognitive performance across the measured domains at baseline. This suggests that both male and female participants exhibit similar cognitive abilities in these areas at the start of the study.

Table 4: Comparison of Cognitive Parameters Between Male and Female Participants at 2^nd month

Table 4 presents a comparison of cognitive performance between male and female participants during the second month of the study across various cognitive domains. The parameters evaluated include Executive Functions & Visuospatial Skills, Language & Naming, Memory, Attention & Concentration, Abstraction, Orientation, and Total Score. The table includes the mean, standard deviation (SD), and standard error of the mean (SEM) for each cognitive domain for both males and females. The results of an Analysis of Variance (ANOVA) test are provided, showing the F-values and corresponding p-values to assess whether there are any significant differences between the genders.

The findings reveal that there are no significant differences in cognitive performance between males and females across any of the parameters. For instance, in Executive Functions & Visuospatial Skills, the males had a mean score of 4.05 (SD = 0.714), while females had a mean of 3.88 (SD = 0.761), but the difference was not statistically significant (F = 1.068, p = 0.304). Similarly, in Language & Naming, males had a mean score of 4.85 (SD = 0.700), and females scored slightly higher with a mean of 5.03 (SD = 0.663), but again, no significant difference was observed (F = 1.173, p = 0.281).

For the Memory parameter, both males (mean = 3.90, SD = 0.744) and females (mean = 3.92, SD = 0.869) showed nearly identical scores, with no significant difference (F = 4.305, p = 0.412). In Attention & Concentration, males (mean = 5.00, SD = 0.784) and females (mean = 4.90, SD = 0.796) also had similar scores, with no significant difference (F = 0.406, p = 0.526). The Abstraction domain showed a slight difference (mean = 1.73 for males and mean = 1.65 for females), but this was also not significant (F = 2.617, p = 0.109).

In the Orientation domain, males scored a mean of 4.08 (SD = 0.888) and females scored 4.22 (SD = 0.885), but this difference was not significant (F = 0.108, p = 0.744). The Total Score (combining all cognitive domains) was identical for both genders (mean = 23.60 for both males and females), with no significant difference (F = 0.004, p = 0.951).

As in the baseline results, the analysis of the second-month data also shows no significant gender differences in cognitive performance across all the measured parameters. This indicates that both male and female participants show similar cognitive abilities in these domains at the second month of the study.

Table 5: Comparison of Cognitive Parameters Between Male and Female Participants at 2^nd month

Table 5 describes in the fourth month of the study, cognitive performance between male and female participants was again compared across various cognitive domains, including Executive Functions & Visuospatial Skills, Language & Naming, Memory, Attention & Concentration, Abstraction, Orientation, and the Total Score. The results indicated minimal differences in performance between genders, with males and females showing nearly identical scores across all parameters. For instance, in Executive Functions & Visuospatial Skills, males scored a mean of 4.43 and females 4.28, with no significant difference (F = 0.010, p = 0.920). Similarly, in Language & Naming, males scored 5.28 and females scored 5.47 (F = 1.332, p = 0.251), and in Memory, males scored 4.30 and females 4.32 (F = 3.006, p = 0.086), with no significant difference. Other domains, including Attention & Concentration, Abstraction, and Orientation, also showed no significant gender differences. The Total Score for both genders was nearly identical, with males scoring 25.78 and females 25.93 (F = 0.122, p = 0.728). Overall, the analysis of the third-month data reflects that gender does not significantly influence cognitive performance, as both male and female participants performed similarly across all domains.

Table 6: Descriptive Statistics for Total Score (30 pts) Across Baseline, 2nd Month, and 4th Month

Table 6 shows the descriptive statistics for the Total Score across the three time points—Baseline, 2nd Month, and 4th Month—show a clear trend of improvement. At baseline, the mean score was 21.09, with a standard deviation of 1.016 and a standard error of 0.102. The 95% confidence interval for the mean ranged from 20.89 to 21.29, and the scores varied between 19 and 22. By the second month, the mean total score increased to 23.60, with a standard deviation of 1.449 and a standard error of 0.145. The confidence interval for the mean was between 23.31 and 23.89, with scores ranging from 20 to 26. The fourth month showed a further rise in the mean total score to 25.87, with a standard deviation of 1.454 and a standard error of 0.145. The confidence interval for the mean was from 25.58 to 26.16, with scores ranging from 22 to 29. Overall, across the three months, the total mean score was 23.52, with a standard deviation of 2.358 and a standard error of 0.136, and scores ranged from 19 to 29. This progression in the scores over time highlights a positive trend in performance, reflecting gradual improvement throughout the study period.

Table 7: ANOVA for Total Score (30 pts) Across Baseline, 2nd Month, and 4th Month

The ANOVA table provides the results of a one-way analysis of variance conducted to compare the Total Score (30 pts) across the Baseline, 2nd Month, and 4th Month time points. The analysis tests whether there are any statistically significant differences in the mean scores between these time points.

• Between Groups: The sum of squares between the groups is 1143.380, indicating the variability in the Total Score due to differences across the three time points. With 2 degrees of freedom (df), the mean square between groups is calculated as 571.690. This represents the average variability between the time points. The F-value is 326.837, which is a very large value, suggesting a substantial difference between the groups. The corresponding p-value is 0.000, which is highly significant and indicates that there are statistically significant differences in the Total Score across the three time points.
• Within Groups: The sum of squares within the groups is 519.500, representing the variability of scores within each time point. With 297 degrees of freedom, the mean square within groups is 1.749, which shows the average variability within each group. This is smaller compared to the variability between the groups, highlighting that the differences between the time points are more pronounced than the variability within each time point.
• Total: The total sum of squares is 1662.880, which represents the overall variability in the Total Score across all three time points combined. The total degrees of freedom is 299, accounting for all the observations in the study.

The significant F-value and p-value indicate that there are meaningful differences in the Total Score over time, confirming that the scores significantly changed between the Baseline, 2nd Month, and 4th Month. These results suggest that the intervention or changes over time had a substantial effect on the participants' scores.

Table 8: Distribution of Child ID by Total Score Categories Across Baseline, 2nd Month, and 4th Month

This table presents the distribution of Child ID categorized by Total Score performance across three time points: Baseline, 2nd Month, and 4th Month. The children’s scores are categorized into three groups: Excellent, Good, and Moderate. The table includes both the number (n) and percentage (%) of children falling into each category for each time point.

• Baseline: At baseline, the majority of children (88.4%) fell into the Moderate category, with 61 children. Eight children (11.6%) were categorized as Good, and none were classified as Excellent. In total, 69 children were evaluated at baseline, representing 100% of the sample at this time point.
• 2nd Month: By the second month, the distribution shifted significantly. A substantial proportion of children (41.3%) were classified as Excellent, with 92 children in this category. The same proportion (41.3%) were classified as Good. Only 17.5% of children (39 children) remained in the Moderate category. In total, 223 children were evaluated at this time point.
• 4th Month: At the fourth month, there was a notable shift in the performance categories. All children (100%) were classified as Excellent, with 8 children meeting this criterion and no children falling into the Good or Moderate categories.

The overall total across all time points shows that the distribution of children across the three categories is 33.3% Excellent, 33.3% Good, and 100% Moderate.

The Chi-Square value for the comparison of the distribution across the three time points is 136.758, with 4 degrees of freedom and a p-value ≤0.001, which indicates that the differences in the distribution of children across the categories over time are statistically significant.

The results indicate significant shifts in the Total Score categories over the study period, with a marked improvement in performance from Baseline to 2nd Month, followed by all children being classified as Excellent by the 4th Month. The Chi-Square test confirms that these changes are statistically significant, reflecting a meaningful progression in the children's performance.

This study explored the effects of HETAFU candy, formulated with probiotics, essential oils, and DHA, on the cognitive performance of children, with a specific focus on potential gender differences across various cognitive domains. The domains assessed included Executive Functions & Visuospatial Skills, Language & Naming, Memory, Attention & Concentration, Abstraction, and Orientation. Evaluations were performed at baseline, after two months, and after four months of regular HETAFU candy consumption.

The results demonstrated consistent improvements in cognitive performance across all domains for both male and female participants over time. Importantly, there were no significant gender-based differences at any point during the study, suggesting that the benefits of HETAFU candy are equally effective for both genders.

Effects of HETAFU Candy on Cognitive Domains

Baseline Performance

At the study's initiation, no significant differences were observed between males and females in any of the cognitive domains. This parity at baseline ensured a robust foundation for analysing the effects of HETAFU candy over time without confounding gender-related disparities. Both genders demonstrated comparable levels of cognitive ability in areas such as memory, language, and visuospatial skills.

Improvements Over Time

Regular consumption of HETAFU candy led to significant cognitive enhancements in all domains by the second and fourth months. The improvement patterns were similar for both males and females, indicating that the formulation’s active ingredients were equally effective regardless of gender.

• Executive Functions & Visuospatial Skills: Both genders showed marked improvements in tasks requiring problem-solving, planning, and spatial awareness. The combination of DHA and essential oils in HETAFU candy likely contributed to these enhancements by supporting neural development and connectivity. The well-documented role of DHA in promoting synaptic plasticity may explain the significant gains in these higher-order cognitive functions.⁶
• Language & Naming: Improvements in this domain were slightly more pronounced in females, though the differences were not statistically significant. Research suggests that females often exhibit advantages in verbal processing tasks, and the neuroprotective effects of DHA might have further enhanced this ability. Nevertheless, males also demonstrated substantial gains, underscoring the universal impact of HETAFU candy.⁶
• Memory and Attention: Both genders exhibited considerable improvement in memory retention and attention span. Probiotics, an essential component of HETAFU candy, are known to positively influence the gut-brain axis, reducing systemic inflammation and enhancing cognitive functions like memory and focus. These results align with findings from previous studies linking probiotic supplementation to improved attention and memory in children.⁵
• Abstraction and Orientation: Participants exhibited significant progress in abstract reasoning and spatial orientation tasks over time. This suggests that the bioactive compounds in HETAFU candy may play a role in enhancing cognitive flexibility and spatial reasoning, essential components of academic and everyday problem-solving skills.

Final Cognitive Outcomes After Four Months

By the end of the fourth month, participants demonstrated robust cognitive improvements across all domains, irrespective of gender. This reflects the cumulative benefits of consistent HETAFU candy consumption, highlighting its potential as a dietary supplement for enhancing brain function. The lack of significant gender differences further supports the hypothesis that the cognitive benefits of HETAFU candy are universally applicable.

The observed outcomes can be attributed to the synergistic effects of the candy’s ingredients:

• DHA enhances neuronal health and connectivity, critical for memory and executive functions.⁷
• Probiotics improve gut health, which indirectly influences brain function through the gut-brain axis.¹⁰
• Essential oils reduce oxidative stress and inflammation, supporting overall cognitive resilience.¹¹

Mechanisms Behind Cognitive Improvement

The unique combination of bioactive compounds in HETAFU candy likely exerts its effects through multiple mechanisms:

1. Gut-Brain Axis: Probiotics modulate gut microbiota, reducing neuroinflammation and enhancing neurotransmitter synthesis, particularly serotonin and dopamine, which are crucial for attention and memory.¹⁰
2. Neuroprotection: DHA, an omega-3 fatty acid, strengthens neuronal membranes and supports synaptic plasticity, directly impacting learning and memory.¹²
3. Antioxidant Properties: Essential oils reduce oxidative stress, which is known to impair cognitive functions, particularly in developing brains.¹¹

Interpretation of Gender Neutrality

The absence of significant gender differences suggests that the cognitive benefits of HETAFU candy are not mediated by gender-specific pathways. Instead, they reflect the universal impact of its ingredients on brain health. This finding is particularly encouraging as it implies that HETAFU candy can be equally effective in diverse populations.

Strengths and Limitations

Strengths:

1. Comprehensive Assessment: Cognitive domains were evaluated at multiple time points, providing a clear picture of temporal changes.
2. Novel Intervention: HETAFU candy offers a practical, non-invasive approach to improving cognitive performance, particularly in children who may struggle with conventional supplements.
3. Gender Inclusivity: The study design ensured equal representation of both genders, enabling robust conclusions about the universal effects of the candy.

Limitations:

1. Sample Size: A larger sample would enhance the generalizability of the findings, particularly to populations with different socio-economic and dietary backgrounds.
2. Confounding Factors: Factors such as variations in diet, physical activity, and home environment were not controlled, which could influence cognitive outcomes.
3. Long-Term Effects: The study duration was limited to four months, and long-term effects of HETAFU candy remain unexplored.

To conclude, HETAFU candy presents a promising, innovative approach to improving cognitive abilities in children, offering a child-friendly, palatable alternative to traditional supplements. By combining probiotics, DHA, and essential oils, the candy leverages the synergistic effects of these bioactive compounds to support cognitive development. Over the course of the four-month study, participants demonstrated significant improvements in cognitive function, with benefits observed across both genders, highlighting the universal applicability of the formulation. This suggests that HETAFU candy is effective regardless of gender-specific biological or neurological differences. DHA plays a pivotal role in neuronal development and synaptic plasticity, while probiotics likely contributed to improvements in memory, attention, and overall cognitive function through the gut-brain axis. Essential oils, with their antioxidative and anti-inflammatory properties, further enhanced the neuroprotective effects of the candy. The results of this study align with the growing body of evidence supporting the use of functional foods in pediatric health. Unlike traditional supplements, HETAFU candy addresses common barriers to supplementation, such as taste and ease of consumption, ensuring better adherence in children. This makes it a practical and feasible option for promoting cognitive health in children. As a safe, non-invasive intervention, HETAFU candy offers an accessible, enjoyable means of enhancing cognitive development, with potential for widespread use in paediatric healthcare. With further validation, it could become a key tool in dietary strategies aimed at optimizing brain function in children worldwide.

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