Chemical profiling and cytotoxic potential of the n-butanol fraction of Tamarix nilotica flowers

Abstract

Background: Cancer represents one of the biggest healthcare issues confronting humans and one of the big challenges for scientists in trials to dig into our nature for new remedies or to develop old ones with fewer side effects. Halophytes are widely distributed worldwide in areas of harsh conditions in dunes, and inland deserts, where, to cope with those conditions they synthesize important secondary metabolites highly valued in the medical field. Several Tamarix species are halophytic including T.nilotica which is native to Egypt, with a long history in its tradition, found in its papyri and in folk medicine to treat various ailments.

Methods: LC-LTQ-MS-MS analysis and ¹H-NMR were used to identify the main phytoconstituents in the n- butanol fraction of T.nilotica flowers. The extract was tested in vitro for its cytotoxic effect against breast (MCF-7) and liver cell carcinoma (Huh-7) using SRB assay.

Results: T.nilotica n-butanol fraction of the flowers was found to be rich in phenolic content, where, LC-LTQ-MS-MS allowed the tentative identification of thirty-nine metabolites, based on the exact mass, the observed spectra fragmentation patterns, and the literature data, varying between tannins, phenolic acids, and flavonoids. ¹H-NMR confirmed the classes tentatively identified. The in-vitro evaluation of the n-butanol fraction showed lower activity on MCF-7 cell lines with IC₅₀ > 100 µg/mL, while the higher promising effect was against Huh-7 cell lines with an IC₅₀= 37 µg/mL.

Conclusion: Our study suggested that T.nilotica flowers' n-butanol fraction is representing a promising cytotoxic candidate against liver cell carcinoma having potential phytoconstituents with variable targets and signaling pathways.

Keywords: 1H-NMR; Cytotoxicity; Huh-7; LC–LTQ–MS–MS; MCF-7; Tamarix nilotica flowers.

Fig. 1

Chemical structures of the tentatively identified compounds in the n-butanol fraction of T. nilotica flowers numbered according to compounds listed in Table 1

Fig. 2

LTQ-LC-MS-MS chromatogram of the n- butanol fraction of T. nilotica flowers

Fig. 3

Molecular network (showing clusters of metabolites of interest) based on tandem mass spectrometry data in the positive ionization mode of the n-butanol fraction of T. nilotica flowers. Twenty metabolites have been identified as labeled in Fig. 3, green color indicating the number of compounds in Table 2, light blue nodes are compounds identified using GNPS databases, while the identified compounds using fragmentation matching have the pink color

Fig. 4

¹H-NMR spectrum exhibiting the identified metabolites in the n-butanol fraction of T. nilotica flowers; primary metabolites i.e., fatty acids and sugars (M1-M4) as well as organic acid (M5) at the aliphatic region δ_H 0.5—5.5 ppm as mentioned in Table 3

Fig. 5

¹H-NMR spectrum exhibiting the identified metabolites in the n-butanol fraction of T. nilotica flowers; in aromatic region δ_H 5.5—8.2 ppm prescribing coumarins and flavonoids

Fig. 6

In-vitro SRB cytotoxicity assay of the n-butanol fraction of T. nilotica flowers against A: Huh-7 and B MCF-7 cell lines in increasing concentrations (0.01–100 µg/mL). Data points are expressed as mean ± SD (n = 3)

Fig. 7

Optical microscope-stained images of quick screening SRB cytotoxicity assay of the n- butanol fraction of T. nilotica flowers against MCF-7; A: negative control, B: 10 µg/mL, C: 100 µg/mL, and Huh-7; D: negative control, E: 10 µg/mL, F: 100 µg/mL