Satiety Evaluation of Mature Okra Flour-incorporated Biscuit

Okra is one of the top ten most consumed vegetable crops by Malaysians, with 1.7kg per capita consumption as reported by the Department of Statistic Malaysia in 2023. Among the total 65.6 thousand tonnes of output available, around 7.7% of the okra is turned into waste and partially fed to the livestock. Mature okra becomes unmarketable and turns into waste because it develops a tougher, more fibrous texture and a less desirable taste, unlike tender, immature okra, which is typically preferred for consumption. Studies in the literature found that mature okra also possesses comparable nutrient content or is even better than immature okra in certain aspects. Incorporation of agricultural waste, for instance, mature okra, into food products can offer significant health advantages besides reducing waste and also promotes a sustainable future, aligning with the objectives of Sustainable Development Goals (SDGs) goal No.12. Besides, transforming mature okra and other agricultural by-products into functional or value-added foods is an urge to address environmental and unsustainable use of natural resources. This study aims to evaluate the impact of mature okra flour-incorporated biscuits (MOFB) on perceived satiety and metabolomic responses. Mature okra flour will be prepared by dehydrating and grinding okra pods, then incorporated into biscuits at 10% substitution of wheat flour. This ratio of mature okra flour incorporation is pre-determined based on the previous phase study, as this ratio of incorporation results in a better nutritional profile and an acceptable sensorial score. Subsequently, thirty healthy young adults will participate in a single-blind, randomised trial comparing MOFB and control biscuits. Satiety will be assessed using the SLIM scale and blood glucose measurements over 165 minutes post-consumption. Blood samples will be collected for hormone and metabolomic analyses using ELISA, RIA, and Liquid Chromatography-Mass Spectrometry (LC-MS). The findings will help determine whether MOFB enhances satiety and alters metabolic profiles, supporting its potential as a functional food for appetite regulation.

• Mature Okra Flour-incorporated Biscuit (MOFB) (Other): Standard biscuit formulation manufactured where 10% of the wheat flour is substituted with mature okra flour, designed to enhance the nutritional profile while maintaining acceptable sensory scores.
• Control Biscuit (Other): Standard biscuit formulation manufactured using 100% wheat flour, designed to match the sensory profile and appearance of the experimental biscuit without the inclusion of mature okra flour.

Primary outcomes

• Satiety Response [Baseline (0 minutes) and post-consumption at 15, 45, 75, 105, 135, and 165 minutes]: The satiety response after biscuit consumption will be determined by the Satiety Labeled Intensity Magnitude (SLIM) scale. The SLIM scale consists of a 100mm vertical line with labelled phrases to indicate satiety: "greatest imaginable fullness", "extremely full", "very full", "moderately full", "slightly full", "neither hungry nor full", "slightly hungry", "moderately hungry", "very hungry", and "extremely hungry", and "greatest imaginable hunger". The first satiety survey is filled before breakfast (time 0), and the remaining surveys at 15, 45, 75, 105, 135 and 165 min later.
• Change in Plasma Insulin Concentration [Fasting state (0 minutes), 75 minutes, and 165 minutes post-consumption]: Plasma concentration of insulin will be measured using an enzyme-linked immunosorbent assay (ELISA) kit. This metric evaluates metabolic responses to the biscuit interventions.
• Change in Active Glucagon-Like Peptide-1 (GLP-1) Concentration [Fasting state (0 minutes), 75 minutes, and 165 minutes post-consumption]: Plasma concentration of active GLP-1 will be measured using an enzyme-linked immunosorbent assay (ELISA) kit. This metric evaluates anorexigenic hormone responses to the biscuit interventions.
• Change in Plasma Peptide YY (PYY3-36) Concentration [Fasting state (0 minutes), 75 minutes, and 165 minutes post-consumption]: Plasma concentration of PYY3-36 will be measured using an enzyme-linked immunosorbent assay (ELISA) kit. This metric evaluates anorexigenic hormone responses to the biscuit interventions.
• Change in Plasma Cholecystokinin (CCK) Concentration [Fasting state (0 minutes), 75 minutes, and 165 minutes post-consumption]: Plasma concentration of CCK will be determined using a radioimmunoassay (RIA) kit. This metric evaluates anorexigenic hormone responses to the biscuit interventions.
• Change in Active Ghrelin Concentration [Fasting state (0 minutes), 75 minutes, and 165 minutes post-consumption]: Plasma concentration of active ghrelin will be measured using an enzyme-linked immunosorbent assay (ELISA) kit. This metric evaluates orexigenic hormone responses to the biscuit interventions.
• Changes in Metabolite Profiles [Fasting state (0 minutes) and 165 minutes post-consumption]: For metabolomic analysis, the blood samples will be taken in the fasting state (0 min) and after the last satiety survey (165 min). Metabolites will be quantified by Liquid Chromatography-Mass Spectrometry (LC-MS) analysis, and the serum samples will be prepared according to In Vitro Diagnostics Research (IVDr) standard operating procedures.

Secondary outcomes

• Blood Glucose Response [Baseline (0 minutes) and post-consumption at 15, 45, 75, 105, 135, and 165 minutes]: The capillary blood glucose will be measured using the finger-prick method during the fasting state (time 0), and at 15, 45, 75, 105, 135 and 165 min later.
• Ad Libitum Energy Intake at a Subsequent Meal [At the end of the satiety evaluation session (165 minutes post-biscuit consumption)]: Following the 165-minute satiety evaluation session, participants will be served an ad libitum meal consisting of fried rice and requested to consume as much as they want until comfortably full. The weight of the food consumed will be calculated by subtracting the remaining food weight from the initial served weight using a digital weighing scale (accurate to 1.0 g). Total energy intake will be calculated by multiplying the weight of the food consumed by the specific caloric content of the fried rice (expressed in kilocalories).

Sex: All, ages: 18 Years to 39 Years, accepts healthy volunteers

Inclusion Criteria:

* Young adults aged between 18 and 39 years.
* Normal Body Mass Index (BMI).

Exclusion Criteria:

* Smokers.
* Pregnant or lactating women.
* Individuals with daily medication prescriptions for chronic metabolic conditions, cardiovascular management, or hormonal regulation.
* Use of daily medications/supplements that influence appetite regulation, metabolic signalling, or systemic metabolomic profiles, including:

  1. Hormonal contraceptives, glucocorticoids, and oral/injectable anti-obesity medications (e.g., GLP-1 agonists).
  2. Antihypertensives and lipid-lowering agents (statins).
  3. Chronic Nonsteroidal Anti-Inflammatory Drugs (NSAIDs).
  4. Daily fibre supplements.

• Mohd Redzwan Sabran, PhD, Study chair, Universiti Putra Malaysia