In-vitro Cytotoxicity and In-silico Insights of the Multi-target Anticancer Candidates from Haplophyllum tuberculatum

This study aimed to investigate the anticancer activity of Haplophyllum tuberculatum(Forsk.) aerial parts ethanol extract and fractions and reveal the potential anticancer targets, binding modes, pharmacokinetics, and toxicity properties of its phytoconstituents. MTT assay was used to investigate the anticancer activity. TargetNet, ChemProt version 2.0, and CLC-Pred web servers were used for virtual screening, and Cresset Flare software was used for molecular docking with the 26 predicted targets. Moreover, pkCSM, swiss ADME, and eMolTox web servers were used to predict pharmacokinetics and safety. Ethanolic extracts of H. tuberculatum on HepG2 and HeLa cell lines showed promising activities with IC50 values 54.12 and 48.1 µg/mL, respectively. Further, ethyl acetate fraction showed the highest cytotoxicity on HepG2 and HeLa cell lines with IC50 values 41.7 and 52.31 µg/mL. Of 70 compounds screened virtually, polygamain, justicidin A, justicidin B, haplotubine, kusunokinin, and flindersine were predicted as safe anticancer drugs candidates. They showed the highest binding scores with targets involved in cell growth, proliferation, survival, migration, tumor suppression, induction of apoptosis, metastasis, and drug resistance. Our findings revealed the potency of H. tuberculatum as a source of anticancer candidates that further studies should support.

Cytotoxicity and Antiviral Activities of Haplophyllum tuberculatum Essential Oils, Pure Compounds, and Their Combinations against Coxsackievirus B3 and B4

Haplophyllum tuberculatum is a plant commonly used in folk medicine to treat several diseases including vomiting, nausea, infections, rheumatism, and gastric pains. In the current study, H. tuberculatum essential oils, hydrosols, the pure compounds R-(+)-limonene, S-(−)-limonene, and 1-octanol, as well as their combinations R-(+)-limonene/1-octanol and S-(−)-limonene/1-octanol, were screened for their cytotoxicity on HEp-2 cells after 24, 48, and 72 h, and then tested for their activity against Coxsackievirus B3 and B4 (CV-B3 and CV-B4) at 3 different moments: addition of the plant compounds before, after, or together with virus inoculation. Results showed that the samples were more cytotoxic after 72 h than after 24 h or 48 h cell contact. However, the combinations R-(+)-limonene/1-octanol and S-(−)-limonene/1-octanol showed less effect on HEp-2 cells than pure R-(+)-limonene and S-(−)-limonene after 24 h, 48 h, and 72 h. 1-octanol exhibited the highest concentration causing 50% cytotoxicity (CC50) on HEp-2 cells after 24 h (CC50 = 93 µg/mL) and 48 h (CC50 = 83 µg/mL). The antiviral assays showed that the tested samples exhibited potent inhibition of CV-B. IC50 values ranged from 0.66 µg/mL to 28.4 µg/mL. In addition, CV-B3 was more sensitive than CV-B4. Both CV-B strains are more inhibited when cells were pretreated with the plant compounds. The hydrosols have no effect, neither on HEp-2 cells nor on the virus. 1-octanol, S-(−), and R-(+)-limonene/1-octanol had important selectivity indexes over time. Although essential oils had potent antiviral activity, they can be considered for application in the pretreatment of cells. However, 1-octanol and the combinations are within the safety limits, and thus, they can be used as an active natural antiviral agent for CV-B3 and CV-B4 inhibition.

The Phytochemical, Antifungal, and First Report of the Antiviral Properties of Egyptian Haplophyllum tuberculatum Extract

In this study, ethanol whole plant extract (WPE) of Haplophyllum tuberculatum was characterized and tested for its antifungal and antiviral activities against Fusarium culmorum, Rhizoctonia solani and tobacco mosaic virus (TMV). High Performance Liquid Chromatography (HPLC) analysis showed that the main phytochemical constituents of H. tuberculatum WPE were resveratrol (5178.58 mg/kg), kaempferol (1735.23 mg/kg), myricetin (561.18 mg/kg), rutin (487.04 mg/kg), quercetin (401.04 mg/kg), and rosmarinic acid (387.33 mg/kg). By increasing H. tuberculatum WPE at concentrations of 1%, 2%, and 3%, all of the fungal isolates were suppressed compared to the two positive and negative controls. Under greenhouse conditions, WPE-treated Chenopodium amaranticolor plants strongly inhibited TMV infection and significantly reduced TMV accumulation levels when compared to non-treated plants. Moreover, the induction of systemic resistance with significant increases in the transcriptional levels of the pathogenesis-related protein-1 (PR-1), chalcone synthase (CHS), and hydroxycinnamoyl-CoA quinate transferase (HQT) genes for treated plants were noticed at 3 and 5 days post-inoculation (dpi) for both assays. To the best of our knowledge, this is the first reported observation of the antiviral activity of H. tuberculatum extract against plant viral infections. Finally, the results obtained suggest that H. tuberculatum WPE can be considered a promising source of both antifungal and antiviral substances for practical use and for developing plant-derived compounds for the effective management of plant diseases.

Lignans, Amides, and Saponins from Haplophyllum tuberculatum and Their Antiprotozoal Activity

A screening of Sudanese medicinal plants for antiprotozoal activities revealed that the chloroform and water fractions of the ethanolic root extract of Haplophyllum tuberculatum exhibited appreciable bioactivity against Leishmania donovani. The antileishmanial activity was tracked by HPLC-based activity profiling, and eight compounds were isolated from the chloroform fraction. These included lignans tetrahydrofuroguaiacin B (1), nectandrin B (2), furoguaiaoxidin (7), and 3,3′-dimethoxy-4,4′-dihydroxylignan-9-ol (10), and four cinnamoylphenethyl amides, namely dihydro-feruloyltyramine (5), (E)-N-feruloyltyramine (6), N,N′-diferuloylputrescine (8), and 7′-ethoxy-feruloyltyramine (9). The water fraction yielded steroid saponins 11–13. Compounds 1, 2, and 5–13 are reported for the first time from Haplophyllum species and the family Rutaceae. The antiprotozoal activity of the compounds plus two stereoisomeric tetrahydrofuran lignans—fragransin B2 (3) and fragransin B1 (4)—was determined against Leishmania donovani amastigotes, Plasmodium falciparum, and Trypanosoma brucei rhodesiense bloodstream forms, along with their cytotoxicity to rat myoblast L6 cells. Nectandrin B (2) exhibited the highest activity against L. donovani (IC50 4.5 µM) and the highest selectivity index (25.5).

A Potent In Vitro α-Amylase Inhibitory Action of Haplophyllum tuberculatum Extracts

Background: Natural plant active compounds were found to inhibit the activity of several enzymes that may be related to several diseases. Objective: This study aimed at testing the antidiabetic activity related to the phenol content by in vitro α-Amylase inhibitory action effect of aqueous, organic and essential oil extracts of Haplophyllum tuberculatum, collected in the town of Laghouat in the steppe region of Algeria. Methods: Two types of aqueous extracts were prepared: Decoction and Diffusion extracts. The organic extracts were prepared with successful maceration in hexane, dichloromethane, ethyl acetate, ethanol and methanol. Also, essential oils were obtained by hydrodistillation. The analysis of the total Phenol content of our extracts was done with Folin-Ciocalteu reagent, as the flavonoid content was obtained in mixture with aluminum trichloride. The effects of the plant extracts on the catalytic efficiency of α-amylase enzyme were represented by the enzymatic inhibitory percentage of each extract in which the inhibitory activity was expressed as IC50. Results: The total phenol content showed values ranging between 0.27 and 11.97 mg gallic acid equivalent / g dry matter. The flavonoid contents vary from 0.05 to 1.50 mg equivalent of rutin /g of dry matter. All the extracts showed good inhibitory activity against α- amylase of IC50, values ranged from 0.05 to 50.03 mg/ml. Conclusion : This study reports for the first time the inhibitory capacity of Algerian Haplophyllum tuberculatum species against α-amylase which could provide natural biologically active agents to be used in the management of diabetes.