Bilberry (Eye)

Presbyopia is a global problem with an estimated 1.3 billion patients worldwide. In the area of functional food applications, dietary supplements or herbs, there are very few reports describing the positive effects of their use. In the available literature, there is a lack of studies in humans as well as on an animal model of extracts containing, simultaneously, compounds from the polyphenol group (in particular, anthocyanins) and iridoids, so we undertook a study of the effects of a preparation composed of these compounds on a condition of the organ of vision. Our previous experience on a rabbit model proved the positive effect of taking an oral extract of Cornus mas in stabilizing the intraocular pressure of the eye. The purpose of this study was to evaluate the effect of an orally administered ternary compound preparation on the status of physiological parameters of the ocular organ. The preparation contained an extract of the chokeberry Aronia melanocarpa, the honeysuckle berry Lonicera caerulea L., and the bilberry Vaccinium myrtillus (hereafter AKB) standardized for anthocyanins and iridoids, as bioactive compounds known from the literature. A randomized, double-blind, cross-over study lasting with a "wash-out" period of 17 weeks evaluated a group of 23 people over the age of 50, who were subjects with presbyopia and burdened by prolonged work in front of screen monitors. The group of volunteers was recruited from people who perform white-collar jobs on a daily basis. The effects of the test substances contained in the preparation on visual acuity for distance and near, sense of contrast for distance and near, intraocular pressure, and conjunctival lubrication, tested by Schirmer test, LIPCOF index and TBUT test, and visual field test were evaluated. Anthocyanins (including cyanidin 3-O-galactoside, delphinidin 3-O-arabinoside, cyanidin 3-O-glucoside, cyanidin 3-O-rutinoside, cyanidin 3-O-arabinoside) and iridoids (including loganin, sweroside, loganic acid) were identified as substances present in the extract obtained by HPLC-MS. The preliminary results showed that the composition of AKB applied orally does not change visual acuity in the first 6 weeks of

Comment in Surv Ophthalmol. 2004 Nov-Dec;49(6):618; author reply 618. doi: 10.1016/j.survophthal.2004.08.009. We have systematically reviewed placebo-controlled trials of V. myrtillus-extracted anthocyanosides for evidence of positive effects on night vision. Searches of computerized databases and citations in retrieved articles identified 30 trials with outcome measures relevant to vision in reduced light. Of these, 12 were placebo-controlled. The 4 most recent trials were all randomized controlled trials (RCTs) and were negative in outcome. A fifth RCT and 7 non-randomized controlled trials reported positive effects on outcome measures relevant to night vision. Negative outcome was associated with more rigorous methodology but also with lower dose level and extracts from geographically distinct sources that may differ in anthocyanoside composition. Healthy subjects with normal or above average eyesight were tested in 11 of the 12 trials. The hypothesis that V. myrtillus anthocyanosides improves normal night vision is not supported by evidence from rigorous clinical studies. There is a complete absence of rigorous research into the effects of the extract on subjects suffering impaired night vision due to pathological eye conditions. Evidence from methodologically weaker trials and auxiliary evidence from animal studies, trials of synthetic anthocyanosides, and a recent randomized controlled trial of Ribes nigrum (black currant) anthocyanosides may warrant further trials of V. myrtillus anthocyanosides in subjects with impaired night vision.

BACKGROUND/OBJECTIVES: Presbyopia is an age-related decline in near vision associated with lens stiffening and neuroretinal changes, while evidence for the effects of berry-derived phytochemicals remains limited. We investigated whether AKB, a double-standardised berry extract (anthocyanins ≥ 25%, iridoids ≥ 4.5%) from Aronia melanocarpa, Lonicera caerulea, and Vaccinium myrtillus, influences visual performance and circulating biomarkers potentially relevant to ocular homeostasis. METHODS: In a randomised, double-blind, placebo-controlled, two-period crossover trial, 23 adults aged >50 years received AKB (400 mg twice daily) or placebo for 6 weeks, separated by a 5-week washout. RESULTS: Compared with placebo, AKB was associated with improvements in selected visual-function outcomes, including near contrast sensitivity and visual-field parameters, together with directionally favourable changes in VEP and OCT readouts. AKB supplementation was also associated with lower circulating αA-/αB-crystallin and ALDH1A1 levels and higher circulating TRPV4 levels, whereas systemic antioxidant enzymes and advanced glycation end-products remained unchanged. Given the small sample size and the indirect nature of the biomarker assessment, these findings should be considered preliminary. CONCLUSIONS: Overall, short-term AKB supplementation was associated with modest, exploratory changes in selected functional and systemic biomarker outcomes, but larger and longer-term studies are needed to confirm clinical relevance and clarify underlying mechanisms.

Chronic diseases cause early death and financial strain worldwide. Cardio-metabolic health, crucial for preventing cardiovascular disease and type 2 diabetes, may benefit from bilberry's antioxidant and anti-inflammatory properties. This meta-analysis reviews studies of bilberry's impact on lipid profiles, glycemic indices, body composition, and inflammatory and oxidative factors. Inclusion criteria were randomized clinical trials assessing bilberry supplementation in adults for at least 1 week. A comprehensive review of literature was performed in PubMed, Web of Science, Scopus, and Google Scholar until July 21, 2024, without any time limitations. Mean changes and their SDs were used to calculate overall effect sizes, with the Hozo et al. method converting SEs, 95% CIs, and IQRs to SDs. A random-effects model accounted for between-study variations. Eleven RCTs, including 409 individuals, were incorporated into the present systematic review, and 8 were included in the meta-analysis. Combining five effect sizes from the five trials on long-term effects of bilberry administration compared with controls resulted in a non-significant decrease in FBG (WMD: -0.08 mmol/L, 95% CI: -0.22 to 0.07, p = 0.30). For HbA1c, the meta-analysis of three RCTs showed a marginally significant reduction (WMD: -1.63%, 95% CI: -3.36% to 0.11%, p = 0.06). The results of the meta-analysis on lipid profile showed a decreasing trend, although this reduction was not statistically significant for TC (WMD: -0.11 mmol/L, 95% CI: -0.30% to 0.08%, p = 0.27) or TG (WMD: -0.07 mmol/L, 95% CI: -0.32% to 0.19%, p = 0.62). However, a significant change in TG was reported in trials with a crossover design and RCTs with 4 weeks of intervention or less. Although HDL level did not show any significant change (WMD: -0.02 mmol/L, 95% CI: -0.10% to 0.07%, p = 0.70), the meta-analysis of five RCTs evaluating the long-term effects of bilberry supplementation revealed a significant change in LDL following bilberry supplementation (WMD: 0.07 mmol/L, 95% CI: 0.01%-0.14%, p = 0.01). Furthermore, no significant reduction was observed in SBP (WMD: -2.75 mmHg, 95% CI: -6.38% to 0.89%, p = 0.13) or DBP (WMD: -1.00 mm

Strategies for successful aging, including the use of food supplements, are part of the approach to support skin youthfulness. To demonstrate the efficacy of fermented bilberry extract (FBE) against skin aging and uneven complexion, a clinical trial was carried out on 66 subjects with visible "crow's feet" wrinkles, mild-to-moderate skin slackness, and uneven skin tone. The wrinkle depth, skin smoothness (Ra) and roughness (Rz), skin firmness (R0) and elasticity (R2), skin coloration (ITA°), and skin antioxidant capacity were measured before and after 28 (D28), 56 (D56), and 84 (D84) days of product use (either FBE or a placebo). These parameters were also integrated with a clinical evaluation, carried out by a dermatologist, and a self-assessment questionnaire to align the measured efficacy with the visual or perceived efficacy. At D84, the wrinkle depth had decreased by 10.6%, Ra had improved by 7.9%, Rz had decreased by 7.3%, R0 had improved by 13.3%, R2 had improved by 12.4%, and skin antioxidant capacity had increased by 20.8%. ITA° increased by 20.8% and was accompanied by a decrease in the skin's redness component by 16.8% and an increase in the lightness component by 2.2%. The variation of all the above-mentioned parameters was statistically significant between the FBE and PL groups. Our findings demonstrate the efficacy of FBE in improving skin aging and complexion evenness.

BACKGROUND: Type 2 Diabetes Mellitus (T2DM) is a major risk factor for the development of cardiometabolic diseases. T2DM prevention is largely based on weight-loss and whole diet changes, but intervention with dietary plant bioactives may also improve metabolic health. OBJECTIVE: To assess whether supplementation with bilberry and grape seed extract for 12 weeks improves cardiometabolic outcomes in individuals at risk of developing T2DM, and to determine whether individual treatment response is associated with differences in gut microbiota composition and levels of phenolic metabolites in blood and feces. METHODS: In the randomized, double-blind, placebo-controlled, cross-over PRECISE intervention study, 14 participants, aged ≥45 years, with a BMI >28 kg/m2, and having an increased risk of T2DM, received a supplement containing 250 mg of bilberry plus 300 mg of grape seed extract, or 550 mg of a control extract, per day, for 12 weeks each. Blood samples were obtained for the assessment of HbA1c, fasting glucose, oral glucose tolerance tests, insulin, glucagon levels, total, LDL and HDL cholesterol, and phenolic acids. We also assessed advanced glycation end products in the skin, ambulatory 24 hours blood pressure, 7-day dietary intake by weighed food diaries, fecal levels of phenolic metabolites using LC-MS/MS and gut microbiota composition using 16S rRNA gene sequencing analysis. RESULTS: The combined bilberry and grape seed extract did not affect glucose and cholesterol outcomes, but it decreased systolic and diastolic ambulatory blood pressure by 4.7 (p < 0.001) and 2.3 (p = 0.0009) mmHg, respectively. Eight out of fourteen participants were identified as blood pressure 'responders'. These responders had higher levels of phenylpropionic and phenyllactic acids in their fecal samples, and a higher proportional abundance of Fusicatenibacter-related bacteria (p < 0.01) in their baseline stool samples. CONCLUSION: Long-term supplementation with bilberry and grape seed extract can improve systolic and diastolic blood pressure in individuals at risk of T2DM. Individual responsiveness was correlated with the presence of certain fecal bacterial strains, and an ability

Berry consumption has beneficial effects on blood pressure. Intestinal microbiota transform berry phytochemicals into more bioactive forms. Thus, we performed a systematic review of randomized clinical trials to determine whether berry polyphenols in foods, extracts or supplements have effects on both the profile of gut microbiota and systolic and diastolic blood pressure in humans. PubMed, Cochrane Library, Scopus, and CAB Abstracts (EBSCOhost) were searched for randomized clinical trials in humans published from 1 January 2011 to 29 October 2021. Search results were imported into Covidence for screening and data extraction by two blinded reviewers, who also performed bias assessment independently. The literature search identified 216 publications; after duplicates were removed, 168 publications were screened with 12 full-text publications assessed for eligibility. Ultimately three randomized clinical trials in humans met the eligibility criteria. One randomized clinical trial showed a low risk of bias while the other two randomized clinical trials included low, high or unclear risk of bias. Together the randomized clinical trials showed that berry consumption (Aronia berry, strawberries, raspberries, cloudberries and bilberries) for 8-12 weeks had no significant effect on both blood pressure and the gut microbiota. More randomized clinical trials are needed to determine the effects of berry consumption on the profile of gut microbiota and blood pressure in humans.

Untargeted liquid chromatographic-high-resolution mass spectrometric (LC-HRMS) metabolomics for potential exposure marker (PEM) discovery in nutrikinetic studies generates complex outputs. The correct selection of statistically significant PEMs is a crucial analytical step for understanding nutrition-health interactions. Hence, in this paper, different chemometric selection workflows for PEM discovery, using multivariate or univariate parametric or non-parametric data analyses, were comparatively tested and evaluated. The PEM selection protocols were applied to a small-sample-size untargeted LC-HRMS study of a longitudinal set of serum samples from 20 volunteers after a single intake of (poly)phenolic-rich Vaccinium myrtillus and Vaccinium corymbosum supplements. The non-parametric Games-Howell test identified a restricted group of significant features, thus minimizing the risk of false-positive retention. Among the forty-seven PEMs exhibiting a statistically significant postprandial kinetics, twelve were successfully annotated as purine pathway metabolites, benzoic and benzodiol metabolites, indole alkaloids, and organic and fatty acids, and five (i.e. octahydro-methyl-β-carboline-dicarboxylic acid, tetrahydro-methyl-β-carboline-dicarboxylic acid, citric acid, caprylic acid, and azelaic acid) were associated to Vaccinium berry consumption for the first time. The analysis of the area under the curve of the longitudinal dataset highlighted thirteen statistically significant PEMs discriminating the two interventions, including four intra-intervention relevant metabolites (i.e. abscisic acid glucuronide, catechol sulphate, methyl-catechol sulphate, and α-hydroxy-hippuric acid). Principal component analysis and sample classification through linear discriminant analysis performed on PEM maximum intensity confirmed the discriminating role of these PEMs. © 2021. Springer-Verlag GmbH Germany, part of Springer Nature.

BACKGROUND: Bilberries from Sweden, rich in polyphenols, have shown cholesterol-lowering effects in small studies, and the cholesterol-lowering properties of oats, with abundant beta-glucans and potentially bioactive phytochemicals, are well established. Both may provide cardiometabolic benefits following acute myocardial infarction (AMI), but large studies of adequate statistical power and appropriate duration are needed to confirm clinically relevant treatment effects. No previous study has evaluated the potential additive or synergistic effects of bilberry combined with oats on cardiometabolic risk factors. Our primary objective is to assess cardioprotective effects of diet supplementation with dried bilberry or with bioprocessed oat bran, with a secondary explorative objective of assessing their combination, compared with a neutral isocaloric reference supplement, initiated within 5 days following percutaneous coronary intervention (PCI) for AMI. METHODS: The effects of Bilberry and Oat intake on lipids, inflammation and exercise capacity after Acute Myocardial Infarction (BIOAMI) trial is a double-blind, randomized, placebo-controlled clinical trial. A total of 900 patients will be randomized post-PCI to one of four dietary intervention arms. After randomization, subjects will receive beverages with bilberry powder (active), beverages with high-fiber bioprocessed oat bran (active), beverages with bilberry and oats combined (active), or reference beverages containing no active bilberry or active oats, for consumption twice daily during a 3-month intervention. The primary endpoint is the difference in LDL cholesterol change between the intervention groups after 3 months. The major secondary endpoint is exercise capacity at 3 months. Other secondary endpoints include plasma concentrations of biochemical markers of inflammation, metabolomics, and gut microbiota composition after 3 months. DISCUSSION: Controlling hyperlipidemia and inflammation is critical to preventing new cardiovascular events, but novel pharmacological treatments for these conditions are expensive and associated with negative side effects. If bilberry and/or oat, in addition to standard medic

Global prevalence of type 2 diabetes (T2D) is rising and may affect 700 million people by 2045. Totum-63 is a polyphenol-rich natural composition developed to reduce the risk of T2D. We first investigated the effects of Totum-63 supplementation in high-fat diet (HFD)-fed mice for up to 16 wk and thereafter assessed its safety and efficacy (2.5 g or 5 g per day) in 14 overweight men [mean age 51.5 yr, body mass index (BMI) 27.6 kg·m-2] for 4 wk. In HFD-fed mice, Totum-63 reduced body weight and fat mass gain, whereas lean mass was unchanged. Moreover, fecal energy excretion was higher in Totum-63-supplemented mice, suggesting a reduction of calorie absorption in the digestive tract. In the gut, metagenomic analyses of fecal microbiota revealed a partial restoration of HFD-induced microbial imbalance, as shown by principal coordinate analysis of microbiota composition. HFD-induced increase in HOMA-IR score was delayed in supplemented mice, and insulin response to an oral glucose tolerance test was significantly reduced, suggesting that Totum-63 may prevent HFD-related impairments in glucose homeostasis. Interestingly, these improvements could be linked to restored insulin signaling in subcutaneous adipose tissue and soleus muscle. In the liver, HFD-induced steatosis was reduced by 40% (as shown by triglyceride content). In the subsequent study in men, Totum-63 (5 g·day-1) improved glucose and insulin responses to a high-carbohydrate breakfast test (84% kcal carbohydrates). It was well tolerated, with no clinically significant adverse events reported. Collectively, these data suggest that Totum-63 could improve glucose homeostasis in both HFD-fed mice and overweight individuals, presumably through a multitargeted action on different metabolic organs.NEW & NOTEWORTHY Totum-63 is a novel polyphenol-rich natural composition developed to reduce the risk of T2D. Totum-63 showed beneficial effects on glucose homeostasis in HFD-fed mice, presumably through a multitargeted action on different metabolic organs. Totum-63 was well tolerated in humans and improved postprandial glucose and insulin responses to a high-carbohydrate breakfast test.

Erratum in Phytother Res. 2022 Apr;36(4):1836. doi: 10.1002/ptr.7459. Bilberry (Vaccinium myrtillus L.) is one of the richest natural sources of anthocyanins which are powerful antioxidants and reported to have antiinflammatory, antidyslipidemic, antihypertensive, and hypoglycemic effects. The objective of this study was to assess the effect of bilberry supplementation on biomarkers of glycemic control, lipid profile, antioxidant, and inflammatory status in patients with type 2 diabetes in a randomized, double-blind, placebo-controlled cross-over study. Twenty patients were randomized to receive either bilberry supplementation (1.4 g/day of extract) daily for 4 weeks followed by 6 weeks of washout and then an additional 4 weeks of matching placebo or vice versa. Blood pressure, metabolic parameters, antioxidant status, and oxidative stress were measured before and after each period. Results showed no effect on body weight, blood pressure, or lipid profile. HbA1c was reduced by 0.31 ± 0.58% during bilberry supplementation, but this change was not significantly different from that with placebo. Antioxidant status, oxidative stress, and inflammatory status showed no significant differences across treatments. This short-term study of bilberry supplementation did not show significant effects on cardiovascular risk factors or antioxidant status, but the tendency for improved glycemic control may suggest a longer treatment period may be effective in diabetic patients. © 2021 John Wiley & Sons, Ltd.

Age-related macular degeneration (AMD) is one of the most widespread degenerative disorders in elderly people. A 90-day, open-label clinical study was conducted in 40 patients, aged 50 years or older, with early-stage dry-type AMD to evaluate the safety and efficacy of Macumax®, a novel mixture of a phyto-mineral nutritional supplement containing ZeaLutein® (consisting of lutein, zeaxanthin, and piperine), extracts of bilberry, saffron, and zinc monomethionine. Subjects received one capsule of the supplement twice daily for 90 days. The treatment measures included physical examination, vital signs, and assessment of subjective and objective symptoms at baseline and after treatment. For efficacy assessment, baseline values were compared with the values after treatment at 30-day intervals, on days 30, 60, and 90. The safety of the treatment was assessed during all the visits. Overall, the patients showed improvement in the subjective symptoms, such as vision scores after treatment compared with baseline. The changes in diminished and distorted vision scores were found to be significant from day 60 (P < .05). In the case of objective symptoms, only 40% of the subjects (P < .05) had abnormal Amsler's grid aberration scores on day 90 compared with 77.5% of subjects at the beginning of the study. No adverse events were observed during the study. This pilot study provides evidence that Macumax® supplementation is safe and maintained eye health without further progression of the disease in patients with early-stage dry-type AMD. Clinical Trial Registration number: CTRI/2016/02/006676.

SCOPE: Diet rich in bilberries is considered cardioprotective, but the mechanisms of action are poorly understood. Cardiovascular disease is characterized by increased proatherogenic status and high levels of circulating microvesicles (MVs). In an open-label study patients with myocardial infarction receive an 8 week dietary supplementation with bilberry extract (BE). The effect of BE on patient MV levels and its influence on endothelial vesiculation in vitro is investigated. METHODS AND RESULTS: MVs are captured with acoustic trapping and platelet-derived MVs (PMVs), as well as endothelial-derived MVs (EMVs) are quantified with flow cytometry. The in vitro effect of BE on endothelial extracellular vesicle (EV) release is examined using endothelial cells and calcein staining. The mechanisms of BE influence on vesiculation pathways are studied by Western blot and qRT-PCR. Supplementation with BE decreased both PMVs and EMVs. Furthermore, BE reduced endothelial EV release, Akt phosphorylation, and vesiculation-related gene transcription. It also protects the cells from P2X7 -induced EV release and increase in vesiculation-related gene expression. CONCLUSION: BE supplementation improves the MV profile in patient blood and reduces endothelial vesiculation through several molecular mechanisms related to the P2X7 receptor. The findings provide new insight into the cardioprotective effects of bilberries. © 2020 The Authors. Published by Wiley-VCH GmbH.

Metabolic syndrome is a cluster of interrelated conditions that is associated with an increased risk of cardiovascular disease (CVD) and type 2 diabetes mellitus (T2DM). Oxidative stress may impair normal physiological functions, leading to various illnesses. T2DM is considered to be associated with increased oxidative stress, inflammation, and dyslipidemia, which may play a significant role in the development of cardiovascular complications, cancer and vision loss through cataracts and retinopathy. While conventional therapies are a cornerstone for the management of the major risk factors of metabolic syndrome, increasing antioxidant defense by increasing intake of antioxidant-rich foods may improve long term prospects in CVD, obesity and T2DM. Bilberry (Vaccinium myrtillus L.) is one of the richest natural sources of anthocyanins which give berries their red/purple/blue coloration. Anthocyanins are powerful antioxidants and are reported to play an important role in the prevention of metabolic disease and CVD as well as cancer and other conditions. This review focuses on the potential effects of bilberry supplementation on metabolic and cardiovascular risk factors. Although there is evidence to support the use of bilberry supplementation as part of a healthy diet, the potential benefits from the use of bilberry supplementation in patients with T2DM or CVD needs to be clarified in large clinical trials.

Metabolic syndrome (MetS) is a global challenge for atherosclerosis. It was hypothesized that a four-week consumption of anthocyanin supplements by MetS patients who had three or more risk factors linked with metabolic syndrome would have a greater improvement in cardiometabolic biomarkers and would also reduce the risk of thrombosis. A total of 55 participants in two groups of Normal healthy and MetS (age 25-75y) were given 320 mg anthocyanin supplements twice daily for 4 weeks. Platelet coagulant activities, lipid profiles, fasting blood glucose, and inflammatory and oxidative stress biomarkers were measured before and after supplementation to evaluate the atheroprotective effects of anthocyanins in the study subjects. Four weeks of anthocyanin supplementation significantly decreased cardiometabolic risk factors including the average serum fasting blood glucose (FBG) (by 13.3%, P < .05) and lipid profiles by significant reduction in triglyceride (by 24.9%, P < .05) and LDL-C (by 33.1%, P < .05) in the MetS group. Anthocyanin supplementation also decreased high sensitivity C-reactive protein (hs-CRP) level (by 28%, P < .05) in females. However, no significant differences in serum UA (uric acid) and HDL-C were observed between anthocyanin pre- and post-treatment in both groups. Moreover, Anthocyanin supplements decreased ADP-induced platelet activation configuration expressed as P-selectin by 40% (P < .05). There was a positive correlation between decreased hs-CRP values and the levels of LDL-C and FBG in the MetS group (P < .05). These results support the hypothesis that anthocyanin supplementation exerts anti-atherogenicity effects by improving cardiometabolic risk factors and reducing thrombogenicity in the MetS population.

This randomized, double-blinded, crossover study measured the acute effect of ingesting a mixed flavonoid-caffeine (MFC) supplement compared to placebo (PL) on energy expenditure (EE) and fat oxidation (FATox) in a metabolic chamber with premenopausal women (n = 19, mean ± SD, age 30.7 ± 8.0 year, BMI 25.7 ± 3.4 kg/m2). The MFC supplement (658 mg flavonoids, split dose 8:30, 13:00) contained quercetin, green tea catechins, and anthocyanins from bilberry extract, and 214 mg caffeine. Participants were measured twice in a metabolic chamber for a day, four weeks apart, with outcomes including 22 h EE (8:30-6:30), substrate utilization from the respiratory quotient (RQ), plasma caffeine levels (16:00), and genotyping for the single-nucleotide polymorphism (SNP) rs762551. Areas under the curve (AUC) for metabolic data from the MFC and PL trials were calculated using the trapezoid rule, with a mixed linear model (GLM) used to evaluate the overall treatment effect. The 22 h oxygen consumption and EE were significantly higher with MFC than PL (1582 ± 143, 1535 ± 154 kcal/day, respectively, p = 0.003, trial difference of 46.4 ± 57.8 kcal/day). FATox trended higher for MFC when evaluated using GLM (99.2 ± 14.0, 92.4 ± 14.4 g/22 h, p = 0.054). Plasma caffeine levels were significantly higher in the MFC versus PL trial (5031 ± 289, 276 ± 323 ng/mL, respectively, p < 0.001). Trial differences for 22 h EE and plasma caffeine were unrelated after controlling for age and body mass (r = -0.249, p = 0.139), and not different for participants with the homozygous allele 1, A/A, compared to C/A and C/C (p = 0.50 and 0.56, respectively). In conclusion, EE was higher for MFC compared to PL, and similar to effects estimated from previous trials using caffeine alone. A small effect of the MFC on FATox was measured, in contrast to inconsistent findings previously reported for this caffeine dose. The trial variance for 22 h EE was not significantly related to the variance in plasma caffeine levels or CYP1A2*1F allele carriers and non-carriers.