EP_084. Senescence-related cytotoxicity and phytochemical profiling of Hibiscus sabdariffa

Wan Mohamad Adham Afiq Wan Zainuzzaman¹, Siti Khadijah Mustapha Kamal¹, Wan Nurul Nadia Wan Seman¹, Nik Aina Syazana Nik Zainuddin¹, Chin Long Poo¹, Mohd Rahimi Ashraf Abd Rahman¹, Nur Liana Md Nasir¹, Norizah Awang¹, Nor Syaidatul Akmal Mohd Yousof², Hussin Muhammad¹, Mei Siu Lau^1*

1. Unit Toxicology & Pharmacology Unit, Herbal Medicine Research Centre, Institute for Medical Research, National Institutes of Health, 40170 Shah Alam, Selangor, Malaysia

2. Unit Phytochemistry, Herbal Medicine Research Centre, Institute for Medical Research, National Institutes of Health, 40170 Shah Alam, Selangor, Malaysia

*Corresponding author: Mei Siu Lau. This email address is being protected from spambots. You need JavaScript enabled to view it.

ABSTRACT

INTRODUCTION: Hibiscus sabdariffa (Roselle) is rich in anthocyanins with antioxidant and anti-inflammatory properties, making it a promising candidate for senescence research in cardiovascular models. This study aimed to profile its phytochemical composition and establish a safe baseline concentration in human cardiac fibroblasts.

OBJECTIVE(S): This study characterised the anthocyanin content of H. sabdariffa extract using High-Performance Liquid Chromatography (HPLC) and High-Performance Thin-Layer Chromatography (HPTLC), and to assess its cytotoxicity on human ventricular cardiac fibroblasts (NHCF-V) to establish non-toxic working concentrations suitable for senescence studies.

MATERIALS & METHODS: An aqueous extract of H. sabdariffa calyces was prepared by hot water extraction, followed by centrifugation, filtration, and spray drying. Phytochemical profiling was conducted using HPTLC and HPLC to identify major anthocyanins, particularly delphinidin-3-O-sambubioside and cyanidin-3-O-sambubioside. Cytotoxicity was assessed in NHCF-V cells treated with extract concentrations ranging from 3.9 to 1500 μg/mL and hydrogen peroxide (100 to 500 µM)  for 72 hours. Cell viability was measured using an ATP-based luminescence assay with appropriate controls.

RESULTS: Chromatographic analysis confirmed the presence of major anthocyanins with distinct peaks and UV absorbance at ~516–524 nm. Delphinidin-3-O-sambubioside accounted for approximately 60% of total anthocyanins, indicating it as the dominant compound over cyanidin-3-O-sambubioside (40%). The extract showed a dose-dependent reduction in cell viability, with a half-maximal inhibitory concentration (IC₅₀) of 1132 µg/mL. Concentrations up to 250 µg/mL maintained viability above 90%. H₂O₂ at 100 µM minimally reduced viability (~100%), supporting its use as a senescence inducer without cytotoxicity.

CONCLUSION: H. sabdariffa extract exhibits a well-defined anthocyanin profile and is non-toxic to NHCF-V cells up to 250 µg/mL. H₂O₂ at 100 µM effectively induces senescence without compromising viability, supporting its application in further senescence-modulating studies.

KEYWORDS: Hibiscus sabdariffa, Phytochemicals, Cytotoxicity, Cardiac Fibroblast, Senescence