Objectives:

The study aimed to formulate a novel herbal mixture of Hypericum perforatum (H) and Melissa officinalis (M), and evaluate its toxicity, microbial load, and phytochemical content.

Materials and Methods:

Total flavonoids were measured by AlCl3/NaNO2 complex formation method and colorimetric assay. Quercetin content of the herbal mixture was determined by reverse-phase HPLC. The in vitro and in vivo safety of the herbal formulation was analyzed using the MTT assay, and acute oral toxicity analysis in the rat model respectively.

Results:

The formulated extract (HM), in comparison with rutin used as standard, had a total flavonoid content of 15.29±0.64 mg rutin per ml sample. Reverse-phase HPLC revealed a quercetin content of 0.187 mg/ml. Microbial tests for Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus and Salmonella spp. were negative. The colony counts for the total aerobic microbial count, and total yeast and mold counts were less than 10 in each case. The MTT assay (with up to about 5 %v/v HM extract) on NIH/3T3 cell line revealed no cell toxicity in the range of concentrations tested. Acute oral toxicity was tested in the Wistar rat model, and the LD50 was 695.2 ± 7.5 mg/Kg. HM extract had a dry weight of 38.1 mg/ml.

Conclusion:

The preliminary results proved the safety of the herbal mixture HM, with toxicity and microbial load well within the limits of accepted guidelines allowable for use in clinical trials.

Keywords: Melissa officinalis, Hypericum perforatum, combined hydroethanolic extract, cell toxicity, animal toxicity