Use of Non-Pharmacological Supplementations in Children and Adolescents with Attention Deficit/Hyperactivity Disorder: A Critical Review

Attention deficit hyperactivity disorder (ADHD) is a common neurodevelopmental disorder in children and adolescents, with environmental and biological causal influences. Pharmacological medication is the first choice in ADHD treatment; recently, many studies have concentrated on dietary supplementation approaches to address nutritional deficiencies, to which part of non-responses to medications have been imputed. This review aims to evaluate the efficacy of non-pharmacological supplementations in children or adolescents with ADHD. We reviewed 42 randomized controlled trials comprised of the following supplementation categories: polyunsaturated fatty acids (PUFAs), peptides and amino acids derivatives, single micronutrients, micronutrients mix, plant extracts and herbal supplementations, and probiotics. The reviewed studies applied heterogeneous methodologies, thus making it arduous to depict a systematic overview. No clear effect on single cognitive, affective, or behavioral domain was found for any supplementation category. Studies on PUFAs and micronutrients found symptomatology improvements. Peptides and amino acids derivatives, plant extracts, herbal supplementation, and probiotics represent innovative research fields and preliminary results may be promising. In conclusion, such findings, if confirmed through future research, should represent evidence for the efficacy of dietary supplementation as a support to standard pharmacological and psychological therapies in children and adolescents with ADHD.


Introduction
Attention deficit hyperactivity disorder (ADHD) is a neurodevelopmental disorder affecting about 5% of children and 2.5% of adults worldwide. It is characterized by dysregulated cognition and behaviors, resulting in inattention, excessive motor activity, and impulsivity [1].
Drug treatments for ADHD mainly act to potentiate the action of catecholamines, which are neurotransmitters involved in the prefrontal cortex responsible for the maintenance of attention and cognitive control [2]. and sensory functions [32]. Evidence in humans is preliminary; however, chronic intake of probiotics is associated in healthy adults with altered brain connectivity during affective and attentive tasks [32].
Given the number and heterogeneity of studies conducted on nutritional supplementation in ADHD, the present review aims at offering a systematization for the results of studies conducted from January 2010 to March 2020 and evaluating the efficacy of non-pharmacological dietary supplementations in ADHD.
We defined the present review as "critical" to highlight our aim to offer clinicians an analytical and up-to-date point of view in the clinical management of children and adolescents with ADHD, specifically in a nutritional supplementation framework alone or in combination with pharmacological treatment. Moreover, this review did not have a systematic or quantitative objective.

Materials and Methods
The present review is reported according to the preferred reporting items for systematic reviews and meta-analyses (PRISMA) [33]. We searched PubMed from January 2010 to March 2020 using the following string: (probiotic OR prebiotic OR vitamin OR mineral OR phytonutrient OR amino acid OR supplementation OR non-pharmacological) AND (ADHD OR attention deficit hyperactivity disorder) NOT review [All Fields] NOT adult NOT infant. The search manually completed with relevant articles. We included randomized controlled trials involving children and adolescents with ADHD receiving non-pharmacological supplementation. We focused on studies addressing cognitive or behavioral outcomes; we excluded papers not written in English, researches addressing other diagnoses than ADHD, studies with no administration of non-pharmacological supplementations, studies that did not report behavioral/cognitive outcomes, and studies on preschool or adult subjects. Specifically, our search did not include infants (<5 years) given ADHD onset which typically corresponds to school age [1]. Moreover, since this review aim was to offer evidences regarding clinical management of children and adolescents with ADHD, we decided to exclude adults in the PubMed search.

Results
Our search found 414 articles. After title and abstract screening, articles retained were 44; we excluded two articles after full text review. In total, 42 articles are included in the present review. The PRISMA flow chart is shown in Figure 1.
In the following text and tables, results are presented according to supplementation categories as follows: a. PUFAs; b. peptides and amino acids derivatives; c. single micronutrient (Zinc or Vitamin D); d. micronutrients mix; e. plant extracts or herbal supplementations; f.
probiotics. Nutrients 2020, 12, x FOR PEER REVIEW 4 of 32 4 Figure 1. Preferred reporting items for systematic reviews and meta-analyses (PRISMA).Flow diagram of the study selection process.
In the following text and tables, results are presented according to supplementation categories as follows:
ADHD-RS and CGI-S: -Significant time x treatment effect: MPH + supplementation group showed greater improvements compared to supplementation alone. CGI-S: -Slow decrease in Omega-3/6 groups, compared with a rapid decrease and subsequent slight increase in the MPH-containing groups. -Adverse events were numerically less frequent with Omega-3/6 or MPH + Omega-3/6 than MPH alone.  No effect of fish oil supplementation was observed for other SDQ subscales or SDQ total score, the ADHD rating scale, or family functioning (FAD).
Cognitive measures: Fish oil supplementation did not lead to any changes on the stop signal task, trail-making task, or flanker task.  CTRS-R: teacher-rated behavioral indexes (specifically, the authors considered the ADHD index).
No between-group difference was found in the changes of the various measures of ADHD symptoms throughout the study period. No between-group or within-group difference was found in the changes of the various measures of symptoms throughout the study period. In a subgroup of 17 children with learning difficulties an increased erythrocyte DHA was more strongly associated with improved word reading (r = 0.683), improved spelling (r = 0.556), an improved ability to divide attention (r = 0.676), and lower parent ratings of oppositional behavior (r = 0.777), hyperactivity (r = 0.702), restlessness (r = 0.705), and overall ADHD symptoms (r = 0.665).
Milte et al., 2015 [49] 87 ADHD or parent-rated symptoms higher than the 90th percentile on the CPRS-R (67 males), including parent-reported learning difficulties. No between-group or within-group difference was found in the changes of the various measures of symptoms throughout the study period. An increased proportion of erythrocyte EPA + DHA was associated with improved spelling (r = 0.365) and attention (r = −0.540) and reduced oppositional behavior   Teacher and Parent rated ADHD-RS-IV: no significant between-groups outcome results. However, those in the ALC group experienced fewer adverse events than the placebo group regarding headaches and irritability.     CGI-I and C-GAS: assessment of severity and improvement as reported by clinician. Clinician-rated ADHD-RS-IV.
CPRS-R: parent-rated behavioral indexes. CTRS-R: teacher-rated behavioral indexes. SDQ: parent-and teacher-rated emotional and behavioral indexes. BRIEF: teacher-rated behavioural measures of executive skills in everyday environment.
CGI-I: the number of responders in supplementation group was 20 (51%) versus 11 (27%) on placebo. Clinician-rated ADHD-RS-IV. Improvement in inattention and hyperactivity symptoms, aggression, emotional dysregulation, conduct problem and problem behaviour in ADHD who received supplementation compared with placebo group. Darling  More of those who stayed on supplementation (84%) were identified as "Much" or "Very Much" improved overall relative to baseline functioning, compared to 50% of those who switched to psychiatric medications and 21% of those who discontinued treatment.
79% of those still taking micronutrients, 42% of those using medications, and 23% of those who discontinued treatment were considered remitters based on parent-reported ADHD. Clinician-rated ADHD-RS-IV. CGI-I and C-GAS: assessment of severity and improvement as reported by clinician. CPRS-R: parent-rated behavioral indexes. Parent-rated CMRS for a measure of emotion dysregulation. SDQ: parent-rated emotional and behavioral indexes.
Varying predictors were found across outcomes: lower pre-treatment folate and B12 levels, being female, greater severity of symptoms and co-occurring disorders in pre-treatment condition, more pregnancy complications and fewer birth problems were identified as possible predictors of greater improvement for outcome measures.

(e)
Authors  Primary outcomes: Parent and Teacher ADHD-RS to measure behavior. Secondary outcomes: C-GAS: assessment of symptoms severity as reported by clinician. Physiological parameters.
Side effect checklist.
Parent and Teacher ADHD-RS: -A significant improvement was found in inattention score and parent total rating score in supplementation group. -Response rate was higher in supplementation group compared to placebo based only on parent rating. C-GAS: No significant between-group difference after treatment. No between-group significant difference in side effects. 1 × 10 10 colony-forming units\day for 4 weeks before delivery + for 6 months after delivery.
Randomization between treatment and placebo groups is equal, except for one study [46], but there is still too much heterogeneity around the number of treatment groups.

Methodologies
We identified three studies [15,17,21] using Acetyl-L-carnitine (ALC), l-carnosine, and PS (Table 1b). It is not possible to depict a systematic comparison of those studies, given the heterogeneous supplementation approaches. Specifically, two works [15,17] compared subjects who tok a placebo to a group that took supplementation as an add-on to MPH. On the contrary, one study [21] used PS alone in the active treatment group and placebo in control group. All studies comprised uniform samples, except Ghajar et al. [17], who also included children with ODD and applied equal randomization in treatment and placebo groups. Two studies [15,17] considered similar outcome measures, including parent and teacher questionnaires regarding children's behavioral and cognitive symptoms. Hirayama et al. [21] used a go/no-go task and ADHD diagnostic criteria. Although these studies focused on different supplementations, they applied similar methodologies, especially regarding sample, randomization, and outcomes. The use of neuropsychological and physiological parameters could be improved and integrated to better understand response to treatment mechanisms. Moreover, future researches could address PS efficacy as an add-on to MPH.

Results
Regarding these studies results, ALC alone did not improve ADHD symptoms [15], but it reduced adverse effects when taken in addition to MPH. Moreover, l-carnosine seemed to improve behavioral problems according to parents [17]. PS had effects on ADHD symptoms, short-term auditory memory, and cognitive tasks [21]. In conclusion, l-carnosine and PS seem to have some effects on ADHD symptoms or cognitive domains but it remains unclear their specific role and mechanisms of action.

Results
A link between zinc consumption and attention improvement was found in two studies [55,56]: zinc treatment improved inattention scores on parent questionnaires [55] and had effects on attention deficit disorder subtype of ADHD [56]. One study [54] did not find significant differences between zinc supplementation and placebo in the outcome measures; however, children taking zinc supplementation in addition to dextroamphetamine showed better drug dose optimization. All studies regarding vitamin D supplementation highlighted positive outcomes on parent-rated behavioral indexes or ADHD symptoms [25,57,58].

Methodologies
Several studies focused on broad combinations of vitamins and minerals [59][60][61][62][63] (Table 1d). One used a mixture of vitamin D and magnesium [60], two provided "daily essential nutrient formula", which contained 13 vitamins, 17 minerals, and 4 amino acids [59,61]. The remaining studies represent further analyses [63] and follow-up research [62] of the Rucklidge et al. paper [61]. All studies were comprised of children with several comorbidities and applied equal randomization between groups. All studies used multiple ratings, except for Hemamy et al. [60], and one work using magnetic resonance imaging (MRI) data as an outcome (however, this last did not find any significant effect) [59].

Results
These studies highlighted improvements in several behavioral, emotional, and cognitive ADHD symptoms. Furthermore, the follow-up study [62] offers additional evidences regarding possible efficacy of micronutrients mix. However, these studies are characterized by low numerosity and high heterogeneity of samples and treatments.

Methodologies
Seven studies focused on plant or herbal extracts containing a mix of micronutrients, vitamins, and macronutrients (Table 1e). It is not possible to depict a systematic comparison of those studies, given the heterogeneous supplementation approaches. Specifically, one work used Ginkgo biloba plant extract as an add-on to MPH [69]. Others examined the efficacy of an herbal compound [64], Korean red ginseng (KRG) [65] or tocotrienol-rich fractions [70], as a single treatment compared to placebo, ningdong granule [66], sweet almond syrup [67], or Ginkgo biloba [68], compared to MPH. These studies focused on patients with ADHD symptoms without comorbidities and they applied an equal randomization, except for one work [64]. Each study used multiple ratings as outcomes, one of which [65] considered neurophysiological assessment and another cognitive tasks measures [64].

Results
All but two studies [68,70] found beneficial effects related to supplementation, in terms of improved symptomatology and/or less adverse effects as compared to MPH. Improvement in attention indexes was found in studies using a blend of herbs [64], Korean red ginseng [65], and Ginkgo biloba as an add-on to MPH [69]. Parent-, teacher-, or clinician-rated behavioral improvement was reported with KRG [65], ningdong granule [66], sweet almond syrup [67], and Ginkgo biloba as an add-on to MPH [69]. No adverse effects of supplementations were highlighted except for one study that reported increased appetite with sweet almond syrup [67], whereas half of the studies reported adverse effects of MPH [66][67][68]. One study [65] showed that KRG reduced the electroencephalography theta/beta ratio, a marker of cognitive processing capacity, significantly more than placebo. These results are heterogeneous and preliminary, and thus future homogeneous investigations that consider physiological parameters could offer more systematic evidences regarding herbal or extract supplementations. Furthermore, conflicting results [e.g., [68][69] between identical supplementation may be due to the different objectives: in one case authors aimed at comparing effects of supplementation with MPH [68], in the other case effects of MPH and non-pharmacological treatment together were compared to placebo [69].

Probiotics: Methodologies and Results
Only one paper focused on probiotics supplementation, through a different study design compared to the other reviewed studies [71] (Table 1f). A six-months-lasting probiotic supplementation was administered soon after childbirth and a follow-up assessment was conducted after 13 years. The quantity of Bifidobacterium species bacteria in the feces of children later diagnosed with ADHD or Asperger syndrome was found to be lower as compared to healthy children. ADHD or Asperger syndrome was diagnosed in 6/35 (17.1%) children in the placebo and none in the probiotic group (p = 0.008). This last study offers preliminary suggestions regarding probiotics supplementation as a preventive treatment, however further randomized clinical trials are needed to offer more systematic evidence regarding this treatment efficacy. Figure 2 summarizes the main findings reported by the included works regarding nutritional supplementations on ADHD behavioral or cognitive symptoms; the vertical axis indicates the number of studies.

Negative results
Non-significant results

Discussion
We investigated the recent literature about the efficacy of non-pharmacological treatments for ADHD in children and adolescents, alone or in combination with pharmacological treatment.

Discussion of Methodologies
It is relevant that nearly half of the reviewed studies used supplementation with PUFA (mostly EPA and DHA as omega3 PUFAs and, for some, omega6; see Table 1a). This supplementation approach could be linked to previous evidence suggesting the involvement of lower blood levels of DHA in children and adolescents with ADHD [12].
The other reviewed studies addressed the efficacy of peptides and amino acids derivatives (Table 1b), micronutrients (alone or in combination; see Table 1c,d), and plant or herbal extracts (Table 1e); one study investigated the association between early-life probiotics supplementation and ADHD or Asperger syndrome diagnoses at puberty (Table 1f). These supplementation approaches are less frequently reported in the scientific literature as compared to PUFA supplementation.
Each reviewed study used different combinations and doses of drugs and/or non-pharmacological supplementations. Therefore, it is not possible to draw systematic conclusions on optimal type or dose of compound that could be useful in the treatment of ADHD symptomatology.
Regarding outcome measures, Table 2 depicts a summary of sources of information that were considered in the reviewed studies. Table 2 highlights the need to consider homogeneous outcome variables in future research to obtain more systematic evidence related to the same outcomes. Moreover, objective neurophysiological outcomes should be more consistently evaluated together with clinical evidence.

Discussion of Results
The majority of reviewed papers reported improvements but no specific effect of different supplements was found, thus suggesting a non-specific beneficial influence of micro-and macro-nutrients on a broad spectrum of functions and symptoms. A possible explanation of this result could be ascribed to general environmental and dietary influences that have been previously associated to the severity of ADHD symptoms in children and adolescents, such as low socioeconomic status, parents' education, and unhealthy diet [72,73]. In this framework, it is still unclear whether ADHD onset and persistence over time represent the cause or the effect of unhealthy dietary patterns that could lead to nutritional deficits [72,73]. In any case, this review suggests that non-pharmacological supplementation, prescribed on the basis of individual nutritional deficiencies, could constitute a valid clinical path. It is not clear whether supplementation has a role for patients with no dietary imbalance. Moreover, the substances that are contained in various supplementations could benefit brain functioning but may also influence overall physiological functioning in children and adolescents, given a non-specific effect of these compounds. Clinicians should support alternative or additional treatment options only after appropriate blood tests and medical examinations.
In any case, the supplementation approach seems to be valid in combination with pharmacological treatment, as highlighted by positive results of MPH combination with PUFAs [35][36][37]52,56], peptides or amino acid derivatives [15,17], zinc [55,56], vitamin D [25,57,58], vitamin D and magnesium [60], and sweet almond syrup [67]. In these studies, ameliorations were found in behavioral symptoms as reported by parents and clinicians, together with less adverse events compared to pharmacotherapy alone. Hence, drugs and ad hoc nutritional supplementation could represent a valid therapeutic approach.
Other studies focused on children and adolescents who were not under pharmacological treatment for reasons including low compliance, adverse effects or non-response. This second group of studies found mixed results, in terms of finding beneficial effects of supplementation alone and of finding no effect at all. However, the majority of these studies reported a beneficial effect of supplements over placebo.
Specifically, 8 out of 14 studies regarding PUFA supplementation alone found symptoms amelioration over placebo in attention, psychosocial functioning, emotional problems, behavior as reported by parents and teachers, and working memory [38,39,41,[44][45][46]51,53]. The only study addressing phosphatidylserine supplementation found positive effects of treatment over placebo in behavioral and cognitive symptomatology as reported by clinicians and through a go/no-go computerized task [21]. However, one study prescribed zinc supplementation against placebo and found no improvement in behavior, memory, or attention [54]. Five studies used plant or herbal extracts versus placebo. Two found significant beneficial effects of a patented blend of herbs (compound herbal preparation) and Korean red ginseng on attention and symptomatology as reported by clinicians [64,65].
Two studies found similar effects of ningdong granule or sweet almond syrup as compared to MPH treatment in behavioral measures reported by parents and teachers, with fewer side effects related to herbal supplementation than MPH [66,67]. Lastly, one study reported greater parent-and teacher-rated behavioral amelioration effects of MPH as compared to Ginkgo biloba supplementation alone [68] and another [70] did not find efficacy using tocotrienol-rich fractions compared to placebo. The only study concerning early-life probiotic supplementation revealed positive effects compared to placebo preventing ADHD onset later in life [71]. Studies regarding micronutrients mix supplementation found beneficial effects over placebo in general functioning, emotional dysregulation, aggression, and attention [59,61,62]. Importantly, a follow-up work by Rucklidge et al. identified various factors related to response to treatment with micronutrients mix, such as lower pre-treatment folate and B12 levels, being female, greater severity of symptoms and co-occurring disorders in pre-treatment condition, more pregnancy complications, and fewer birth problems [63]. This work highlighted the role of biological and environmental variables related to response to non-pharmacological treatment. This last area of research needs further research, given the high heterogeneity of results due to confounding biological and environmental variables.

Limitations
There were limitations within the articles described in this review. Results should be interpreted in the light of high heterogeneity related to various methodological factors. Indeed, the included works considered heterogeneous treatments, trial durations, methodologies (e.g., supplementation used as unique or combined treatment), and outcomes, even within the same category of supplements. Hence, it was not possible to carry out a meta-analysis of research results, which instead would be auspicable to provide clinicians with more systematic evidence. Moreover, samples were not uniformly involving only children with ADHD diagnoses; other comorbidities or typically developing children were included in some samples. Studies in this research field are also susceptible of cultural influences such as local dietary habits, thus making results difficult to generalize.
Although the majority of studies used similar parent and/or teacher assessment measures as primary outcomes, in many cases clinicians' evaluation or neurophysiological / neuropsychological assessments were lacking. Only three studies [38,59,65] used neurophysiological data, like magnetic resonance imaging or electroencephalography. These kinds of assessments should be included in future research.
Lastly, the majority of studies found beneficial effects, but this may be due to the fact that only studies that found effects were published. However, a formal evaluation of bias was not conducted due to the non-systematic nature of this review.

Conclusions
This review suggest that supplementation approaches may be effective in add-on to pharmacotherapy in improving some behavioral and neuropsychological indicators in children and adolescents with ADHD. The heterogeneity of results suggests that supplementation should be personalized based on each patient's dietary issues. Several supplementation components that are still poorly investigated and may be effective. Moreover, some nutritional supplementations could represent an alternative treatment or rehabilitation in situations of non-response or poor compliance or lack of tolerability of drug treatments, a field that must still be investigated further.

Conflicts of Interest:
The authors declare no conflict of interest.