**1. Introduction**

Cancer can be defined as a non-communicable disease with progressive uncontrolled cell growth/division with a probability to invade nearby and/or distant tissues and could appear in any part of the body [1–4]. Colorectal cancer, "CRC", is the second most common cancer in both genders [5]. Natural products with anticancer therapeutic potential have earned an increasing interest in recent years. Pharmacological research of plant extracts as a source of secondary metabolites is probably the most important step in the identification

**Citation:** Al-Abdallat, K.; Obeidat, M.; Ababneh, N.A.; Zalloum, S.; Al Hadidi, S.; Al-Abdallat, Y.; Zihlif, M.; Awidi, A. Phytochemical Analysis and Anticancer Properties of *Drimia maritima* Bulb Extracts on Colorectal Cancer Cells. *Molecules* **2023**, *28*, 1215. https://doi.org/10.3390/ molecules28031215

Academic Editors: Barbara De Filippis, Alessandra Ammazzalorso and Marialuigia Fantacuzzi

Received: 25 October 2022 Revised: 2 January 2023 Accepted: 4 January 2023 Published: 26 January 2023

**Copyright:** © 2023 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

of plant-derived compounds with therapeutic promise [6]. Plant extracts are a major source of phenolic compounds and other secondary metabolites, which have shown potential anticancer activities by regulating cell death pathways [7]. Furthermore, most of the anticancer drugs in the market are plant-derived such as taxanes (e.g., paclitaxel (Taxol) and docetaxel (Taxotere)) or synthetic compounds. Therefore, a need for basic and applied research to find novel cancer prevention and treatment options from plants as a source for safe and effective anticancer treatment [8].

To our knowledge, the cytotoxicity of the bulb extract of the medicinal plant *D. maritima* toward CRC cell lines was not previously investigated. Therefore, this research was initiated to study the therapeutic potential of *D*. *maritima* on CRC via assessment of the cytotoxic activity of *D. maritima* bulb extract on the colorectal Caco-2 and COLO-205 cancer cell lines. In this study, we prepared an aqueous extraction of the medicinal plant *D. maritima* bulb to assess the in vitro antiproliferative effects of *D*. *maritima* bulb extract against colorectal cancer cells. We also aimed to determine the mechanism of action of active *D*. *maritima* bulb extract against colorectal cancer cells. Finally, we aimed to examine the gene expression of some cell growth-related genes in bulb extract-treated colorectal cancer cells using quantitative RT-PCR.

#### **2. Results**

#### *2.1. Identification of Chemical Constituents of D. maritima Bulb Extract by GC-MS*

Analysis of the bioactive constituents of the bulb extract of *D. maritima* by GC-MS demonstrated the presence of 57 compounds and their chromatogram, as shown in Figure 1 and Table 1. Major peaks determined in the GC-MS chromatogram and their corresponding components were recognized according to NIST 20 and Willy 19 Libraries. The bulb extract was found to include a variety of essential phytochemicals. The results of GC-MS analysis showed that *D. maritima* bulb extract was rich in the phytosterol "Campesterol" containing ~14%, 2,3-Butanediol (glycol and secondary alcohol) and 5-Hydroxymethylfurfural (furans, an arenecarbaldehyde, and primary alcohol) (~10% each), and ~8% for each of the fatty acid derivatives "Hexadecanoic acid ethyl ester" and "9-Octadecenamide (Z)-". Interestingly, the cardiac glycoside "Proscillaridin" proved to be one of the predominant secondary metabolites in the bulb extract of *D. maritima* with about 5% content which may play an effective role as a pharmaceutical anticancer agent (Table 1).

#### *2.2. Drimia maritima Bulb Extract and ProA Can Selectively Inhibit the Proliferation of COLO-205 and Caco-2 Cancer Cells*

The MTT assay was performed to evaluate the cell viability and proliferation of COLO-205 and Caco-2 cells upon treatment with different concentrations of *D*. *maritima* bulb extract, ProA, and Doxorubicin (positive control). The treatment of COLO-205 and Caco-2 with different concentrations of *D*. *maritima* bulb extract showed cytotoxic effects proportional to the concentration of *D*. *maritima* bulb extract used. The IC50 of *D*. *maritima* bulb extract on COLO-205 cell lines was ~2.3 μg/mL and for Caco-2 cells was ~0.9 μg/mL (Figure 2). In order to determine whether *D*. *maritima* bulb extract has cytotoxicity effects on normal human cells, normal HGF cells were treated with the plant extract using the same concentrations used in colon cancer cell lines and incubated for 48 h. The results showed that *D*. *maritima* bulb extract has minimal cytotoxic effects on normal human cells at low concentrations, and the IC50 was achieved at a relatively high concentration of 13.1 μg/mL (Figure 2), indicating that *D*. *maritima* bulb extract potentially has selective cytotoxic effects on cancer cells but not on normal cells. From the reported IC50s, an estimation for the selectivity index can be found to be approximately 5.7 for the COLO-205 cells and 14.55 for Caco-2 cells.

**Retention Time (min)**

**Figure 1.** Gas chromatography and mass spectrometry chromatogram showing the major components of the aqueous bulb extract of *Drimia maritima*. \* a.u.: arbitrary unit.


