*4.3. Intervention*

In the brown seaweed extract group, each participant received 500 mg of polyphenolrich brown seaweed *Ascophyllum nodosum* and *Fucus vesiculosus* extract (2 capsules sold as a standardized extract under the registered trademark InSea2® of InnoVactiv Inc. (Rimouski, QC, Canada) to be taken daily 30 min before the main meal of the day for 12 weeks). The extract was prepared using an exclusive hot water extraction, followed by filtration and ultrafiltration processes and spray-drying. The extract used in the present study contained algae polyphenols (35%, as indicated in the certificate of analysis of the commercial product). The extraction process removed alginates and part of the salts, and the remaining constituents were mostly composed of algae polysaccharides, of which fucoidans were 11.5% fucose and minerals (iodine content = 67 mg/kg). The ability of InSea2® to inhibit αamylase was 97.8% at a concentration of 0.1 g/L algae extract, as indicated in the certificate of analysis of the commercial product. Heavy metal analysis showed that inorganic arsenic (<0.41 mg/kg), cadmium (0.31 mg/kg), lead (0.77 mg/kg), and mercury (0.03 mg/kg) were less than the limit of quantification. The chemical characterization of InSea2® has previously been described in detail [18], confirming that the extract is mainly composed of saccharide derivatives and contains ~35% algae polyphenols, of which phloroglucinol derivatives, measured by NMR analysis and revealing the presence of fatty acids by GC-MS.

Participants of the control group received the same dose of placebo capsule to be taken in the same conditions as described above. Treatment and placebo capsules were similar in form, color, and taste and were supplied by innoVactiv Cie (Rimouski, Canada). The treatment capsule contained extract of brown seaweed (250 mg), dibasic calcium phosphate (150 mg), microcrystalline cellulose (100 mg), croscarmellose (3 mg), magnesium stearate (5 mg), titanium dioxide (1.8 mg), and hypromellose (94.3 mg). Placebo capsule contained microcrystalline cellulose (191 mg), dibasic calcium phosphate (287 mg), magnesium stearate (5 mg), caramel (25 mg), titanium dioxide (1.8 mg), hypromellose (94.3 mg).

The nutritional intervention aimed for moderate calorie restriction of about 500 kcal based on basal metabolism corrected for physical activity, using the traditional Harris Benedict–formula [53]. The individualized counseling intervention was given in the form of a dietary plan based on the "Food Guide for Diabetic Patients" [54] and "Health Plate Approach" [55]. Protein powders, fish oil, omega-3 or any marine supplements, as well as natural health products known to have effects on glucose or insulin levels, weight, satiety, or appetite, were forbidden. Moreover, it was not allowed to use weight loss products available on the market or to follow another weight loss program. In addition, slimming or anti-cellulite cosmetic creams were not permitted. A validated online Food Frequency Questionnaire [56] was completed by participants. Short International Physical Activity Questionnaire [57,58] and Side Effect Reporting Form were completed. Compliance was assessed by capsule counts [59]. Any change in medication, temporary medication, natural health product intake, or consumption of any other food supplements were monitored according to the exclusion criteria during the entire study period. To document compliance, subjects were asked to return unused capsules at the end of the study. Capsule counts indicated a minimum of 80% compliance in both groups.
