EFFECTS of MORINGA OLEIFERA LEAF POWDERS on HEMATOLOGICAL PROFILES IN PREGNANT WOMEN WITH IRON DEFICIENCY ANAEMIA

Title: The Effect of Moringa Oleifera Leaf Micronized Powders on Hematological Profile, Hepcidin, and Cytokines (IL-1 and IL-6) in Pregnant Women with Iron Deficiency Anemia

Background and Rationale:Iron deficiency anemia (IDA) remains a significant public health challenge during pregnancy, especially in low-to-middle-income countries. The condition affects both maternal and fetal health, increasing the risk of preterm delivery, low birth weight, and maternal mortality. Despite the widespread use of conventional iron supplements, the prevalence of anemia in pregnant women in Indonesia consistently exceeds 40% over the last two decades (WHO, 2022). Conventional iron tablets often result in gastrointestinal side effects, poor adherence, and limited bioavailability, highlighting the urgent need for alternative therapies.

Moringa Oleifera leaves have been widely recognized for their high nutritional value, containing essential micronutrients such as iron, calcium, vitamin C, and antioxidants. Emerging studies suggest that Moringa leaves have hematopoietic, anti-inflammatory, and antioxidant properties, making them a promising adjunctive therapy for iron deficiency anemia. However, high-quality clinical trials evaluating their efficacy and safety in pregnant women are still limited.

This randomized controlled trial (RCT) will investigate whether Moringa Oleifera Leaf Micronized Powders (MOLMP) combined with standard iron supplementation improves hematological parameters and reduces inflammation compared to iron supplementation alone in pregnant women with IDA.

Study Design:

Type: Open-labeled, randomized controlled trial (RCT) Allocation: 1:1 Randomized Allocation Intervention Model: Parallel Groups Masking: None (Open Label) Study Duration: 60 Days Sample Size: 72 participants (36 in each group) Study Site: Community Health Centers (Puskesmas Muka, Cianjur Kota, Cijedil, Karang Tengah) in Cianjur, West Java, Indonesia

Intervention Groups:

Group A (Intervention): Moringa Oleifera Leaf Micronized Powders (MOLMP) capsules (3x650 mg/day) + standard iron tablets (Fe tablet 60 mg + Folic Acid 400 mcg/day) Group B (Control): Standard iron tablets (Fe tablet 60 mg + Folic Acid 400 mcg/day) only

The Moringa Oleifera powder will be processed through micronization technology to increase bioavailability and optimize nutrient absorption.

Primary Outcome Measures:

Hemoglobin (Hb) levels

Secondary Outcome Measures:

Hematocrit (Hct) levels Erythrocyte Indices (MCV, MCH, MCHC) Serum Hepcidin levels (as a biomarker of iron homeostasis) Serum IL-6 (Pro-inflammatory cytokines) Serum IL-1β (Pro-inflammatory cytokines) Peripheral Blood Smear Analysis Total Iron Binding Capacity (TIBC) Reticulocyte Hemoglobin Content (RET-He) Red Cell Distribution Width (RDW-SD, RDW-CV) Liver Function Tests (SGOT, SGPT) Kidney Function Tests (Creatinine levels)

Methodology:

Participants will undergo screening and baseline laboratory tests on Day 0, followed by random allocation into two groups. All participants will receive standard antenatal care. Intervention products will be administered daily under self-administration supervision for 60 days. Compliance will be monitored through weekly visits, phone reminders, and supplement consumption diaries.

Blood samples will be collected at baseline (Day 0), and Day 60 to assess hematological parameters, iron status markers, and inflammatory biomarkers. Safety assessments will be conducted throughout the study to monitor liver and kidney function.

Randomization and Allocation Concealment:

Random allocation will be conducted using computer-generated random numbers. Participants will be assigned to either Group A or Group B with a 1:1 ratio.

Statistical Analysis Plan:

Baseline characteristics will be compared using Chi-square tests for categorical variables and Independent t-tests or Mann-Whitney U tests for continuous variables.

Primary outcomes will be analyzed using Repeated Measures ANOVA or Mixed Effects Models to evaluate changes over time.

Secondary outcomes will be analyzed using ANCOVA adjusted for baseline values and potential confounders.

Statistical significance will be set at p < 0.05.

Ethical Considerations:

The study has received ethical approval from the Ethics Committee of Universitas Padjadjaran (No: 159/UN6.KEP/EC/2025). Written informed consent will be obtained from all participants prior to enrolment. All procedures will comply with ICH-GCP (Good Clinical Practice) guidelines and the Declaration of Helsinki.

Risk and Safety Monitoring:

Potential side effects of Moringa Oleifera supplements include hypotension and hypoglycemia. Adverse events will be recorded using standardized Adverse Event Report Forms. Liver and kidney function will be closely monitored.

Data Confidentiality:

Participant data will be anonymized using unique identification codes. All information will be stored in encrypted databases and retained for a minimum of 10 years.

Expected Outcomes:

It is anticipated that the addition of Moringa Oleifera Leaf Micronized Powders will result in:

A significant increase in hemoglobin levels Improved iron homeostasis through reduced hepcidin levels Lower levels of pro-inflammatory cytokines (IL-1 and IL-6) Minimal side effects compared to standard iron tablets

Significance of Study:

If proven effective, Moringa Oleifera leaf powder could serve as a safe, natural, and cost-effective alternative to improve iron deficiency anemia in pregnant women, especially in resource-limited settings.

Due to an extension of the participant recruitment period, the anticipated timelines of the study have been updated. Participant recruitment is expected to continue until February 2026. The primary completion date defined as the date when the final participant receives the intervention and primary outcome data are collected, is anticipated to be April 2026. Final data collection for all study outcomes is anticipated to be completed by May 2026.

Principal Investigator:

H. Awie Darwizar, M.D., Sp.OG., D.MAS., MMRS., FIHFAA Email: [email protected] Phone: +62 812 2323 132

Inclusion criteria

• Demographic Characteristics:

• Women aged 18-44 years
• Married, have a husband and/or family
• Have an ID card
• Clinical Characteristics:

• Pregnant with a gestational age of 28 to <32 weeks
• Live single fetus
• Hemoglobin (Hb) 7 - < 11 g/dl
• No congenital abnormalities
• No complication (DM, infection, genetic disease) based on history taking, clinical signs and symptoms and treatment history.
• Parity < 4
• Geographical/Administrative Characteristics:

• The study subjects (pregnant women) were located in the working area of the health facility where the study was conducted.

Health Facility where the study was conducted.

• Come to the health facility independently or by referral.

Exclusion criteria

• Pregnant women with high risk who dropped out or could not be followed up and pregnant women who gave birth prematurely.
• Allergy to Moringa Oleifera leaf micronized powders