This was a randomized double-blinded placebo-controlled trial conducted from August 2017 to July 2020 at the King Chulalongkorn Memorial Hospital (KCMH), Thailand. The study was conducted in accordance with the Declaration of Helsinki and the Ethics Committee of the Faculty of Medicine, Chulalongkorn University approved the study protocol (IRB no. 240/60). The participants signed informed consent forms prior to enrollment. This trial was registered at http://www.clinicaltrials.gov as NCT03968003. CONSORT was done as the health research reporting checklist for randomized trial. Obese children aged between 7 and 15 years who had BMI above median plus 2 standard deviations (SDs) from the WHO growth reference14 were recruited from the Pediatric Nutrition and the Pediatric Obesity clinics from the KCMH, as well as from the social media (Chula Kids Club). All the children who met the inclusion criteria together with their parents were approached by the researchers and those agreed to the monthly follow-up and consented to the study were enrolled. The exclusion criteria were syndromic obesity, endocrine causes of obesity (e.g. hypothyroidism, growth hormone deficiency), concomitant use of medications that influence appetite or body weight (e.g. corticosteroids), and attending other concurrent weight reduction programs.
Participants were randomly allocated to 3 groups: inulin, placebo, and dietary fiber advice group. Randomization was performed by means of computer-generated permuted blocks size of 6. A research assistant who did not participate in data collection and analysis, generated the random allocation sequence and prepared the sealed envelopes. Other researchers enrolled participants and blindly assigned them to each group. To ensure concealment, the study products were weighed, packaged, and signed by consecutive numbers according to the randomization list by non-involved personnel. The inulin group consumed 13 g of extracted inulin powder from Thai Jerusalem artichoke around 30 min prior to dinner daily. The participants were recommended to consume the powder by mixing the powder to 150 ml of warm water and then stirring up the powder until dissolved. The placebo group consumed 11 g of isocaloric maltodextrin (Oligocarb; Ma-Jusmin Company Limited, Bangkok, Thailand) in the same manner. Blinding of the placebo to match the light brown color of inulin extract were processed through the study. Both supplements were provided in identical foil sachets. All participants were followed-up monthly for 6 months. During the monthly visit, the participants were asked to return all empty, half-empty, and full sachets, and the information was recorded. The parents/guardians were asked to do a parental checklist and verified with the researchers during the follow-up visits. The third group received structured advice (with portion size pictures) to consume appropriate amounts of dietary fiber for age15,16. Research assistants performed telephone contacts every 1st and 3rd week to check and monitor compliance and side effects.
Apart from the interventions, all groups received the same dietary advice about low energy (1,000–1,200 kcal/day) and low fat (25% of total energy from fat) diet. All participants were instructed to exert non-weight-bearing exercise for 60 min per day at least 4–5 days per week, to maintain a physical active lifestyle, and to reduce screen time.
Inulin extraction process
Inulin was prepared from Jerusalem artichoke (Helianthus tuberosus). Dry power of Jerusalem artichoke was purchased from local distributors in Nakhon Ratchasima, Thailand. The product was approved for human consumption by the Food and Drug Administration of Thailand (No. 30–1-14,358–1-001). The inulin extract was prepared based on the patented protocol developed by our team (Patent no. 15858, Inventor: Chonnikant Visuthranukul and Supakarn Chamni, Chulalongkorn University and National Science and Technology Development Agency, Thailand). Briefly, Jerusalem artichoke powder was decocted in water at 60-80ºC for 45–90 min. The resulting slurry was filtered, mixed with absolute ethanol, and incubated at room temperature overnight for inulin precipitation. Inulin was collected by filtration. All remained moisture contents were dried at 40-50ºC and pulverized to obtain the fine and homogenous inulin powder. The inulin content was evaluated based on the standard methods for dietary fiber including energy (ASTM Method D 240–76), total dietary fiber (AOAC (2016) 985.29), soluble dietary fiber (AOAC (2016) 991.42, 991.43), insoluble dietary fiber (AOAC (2016) 991.42), and fructans (Inulin + Oligofructose) (s) AOAC (2005) 997.08 + J. AOAC, 2000, 83(4); 1020–1025)17,18. The tests were performed by certified laboratory of the Institute of Nutrition, Mahidol University, Thailand. The degree of polymerization (DP) of resulting inulin was determined by MALDI-TOF Mass Spectrometry and Gel Permeation Chromatography coupled to multiangle laser light scattering. Jerusalem artichoke inulin indicating great distribution of fructan polymerization containing molecular mass 3460 m/z, which referred to the large DP of the resulting inulin extract. The microbial and heavy metal contaminants, and pesticide residues were examined and confirmed by the Department of Science Service, Thailand. The inulin extract was kept in aluminum foil sachets and stored in a dry container at ambient temperature until use.
Assessment of dietary intake, physical activity, and exercise
Dietary intake was assessed by a dietician, using 3-day dietary records (two weekdays and one weekend day). Fiber and other nutrient intakes were calculated using the Institute of Nutrition, Mahidol University Calculation-Nutrients (INMUCALs) Version 319. Adherence to the instruction of physical activity and exercise was evaluated by a physical activity questionnaire at every visit. The intensity of physical activity was defined by day/week and duration of physical activity in minutes/hours. Aerobic dance or bicycle riding with speed was classified as high intensity exercise. Brisk walking was classified as moderate intensity whereas walking from one place to another for at least 10 min was categorized as low intensity. Sedentary activity was defined as a type of lifestyle involving little or no physical activity which was interviewed and assessed by a research assistant. Screen time, such as watching television and playing games on the computer, smart phone, and tablet, was assessed separately from sedentary activity.
Anthropometry, body composition, and clinical evaluation
Trained personnel performed the anthropometric measurement. Weight and height were measured without shoes and with light clothing using a stadiometer to the nearest 0.1 kg and to the nearest 0.1 cm, respectively. Waist circumference was measured at the umbilicus level after normal exhalation with participants in standing position. Hip circumference was measured at the maximum circumference of the hips. BMI was calculated as weight in kilograms divided by the square of height in meters (kg/m2), and BMI z-score was calculated based on WHO 2007 growth reference using WHO Anthroplus program20. Body composition was measured by bioelectrical impedance analysis (BIA) using the InBody 770® (InBody Co., Ltd., Chungcheongnam-do, Korea). This multi-frequency with 8-point tactile electrodes was evaluated in Korean children aged 6–18 years21 with 99% reproducibility. Fat mass index (FMI) and fat-free mass index (FFMI) were calculated in the same manner as BMI22.
Blood pressure was measured by blood pressure monitor (Dinamap®). The presence of acanthosis nigricans was documented by a pediatrician. Tanner staging was assessed by a self-administered picture questionnaire and verified by a pediatrician.
Venous blood was obtained after a 12-h fast to evaluate biochemical parameters at the 1st and 6th visits of the study. Fasting plasma glucose (FPG) was measured by the hexokinase method (GLUCOSE, Architech; Abbott Laboratories, Irving, TX). Serum total cholesterol, HDL-C, and triglyceride were measured by enzymatic colorimetric assay (CHOLESTEROL, Architech; ULTRA HDL, Architech; and TRIGLYCERIDE, Architech; Abbott Laboratories). LDL-C was measured by homogeneous liquid selective detergent (DIRECT LDL, Architech; Abbott Laboratories). Serum alanine aminotransferase (ALT) was determined according to the standard of International Federation of Clinical Chemistry (ALANINE AMINOTRANSFERASE, Architech; Abbott Laboratories).
Sample size estimation: With power of 0.80 and alpha level of 0.05, sample size of 55 obese children per group was estimated using SD of 0.267 and minimal clinical difference for BMI z-score change was 0.17.
Baseline characteristics for participants in each group were described in mean and SD or frequency and percentages. One-way ANOVA was used to evaluate the difference in the change of variable outcomes between baseline and the 6th month. Generalized Estimating Equation (GEE) model was used to evaluate the changes in outcomes, which included body weight and adiposity, among three groups over three time points (baseline, three, and six months). To evaluate the effect of time on the difference in outcomes among the three groups, the GEE model with interaction term between time point and group was evaluated using likelihood ratio test. The alpha level of 0.05 was considered statistically significant for all analyses. All analyses were conducted using STATA version 16.1 (STATA Statistical Software: Release 16. College Station, TX: STATA Corp LLC. 2019).
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