Plants have been widely used in healthcare to treat many diseases since prehistorical times. The increase in population size, lack of some medications and often high cost, as well as the unwanted side effects observed with many synthetic drugs, has led to increased scientific and commercial attention to medicinal plants. In fact, many effective drugs used nowadays are derived from plant sources. For example, morphine, which is used to relieve severe pain, is derived from Papaver somniferum plant or the opium poppy ^[1]. Other examples include digoxin, which is used in treating heart failure, is derived from Digitalis purpurea or foxglove plant ^[2] and quinine from Cinchona bark, which is used in treating malaria ^[3]. With the huge medicinal success of herbs, the development of plant-based drugs will continue.

The genus Ziziphus of the Rhamnaceae family contains over 58 accepted species of thorny shrubs and small trees growing mostly in arid and semi-arid regions. Species of this genus are traditionally known for their health benefits, nutritional values, and therapeutic properties, as described by various cultures around the world, specifically in India, Pakistan, China and the Middle East ^[4][5][6].

Zizyphus nummularia is rich in several phytochemical compounds, particularly cyclopeptide alkaloids. One of the earliest studies performed in 1983 reported the isolation of two new cyclopeptide alkaloids, nummularine-M and nummularine-N, and the presence of a dozen cyclopeptide alkaloids in the root bark of the plant ^[7]. Cyclopeptide alkaloids are natural macrocyclic compounds with fascinating chemical and biological characteristics, widely present in the Rhamnaceae family, and particularly the Ziziphus genus ^[8][9]. In fact, these compounds have been described as playing a role in the central nervous system due to their sedative, analgesic, and anti-nociceptive effects, in addition to having antimicrobial properties and an antidiabetic potential ^[8], This has been drawing the attention of chemists and biologist to elucidate the structure-activity relationships of these compounds, being a promising source of therapeutic agents. The use of different solvents in the process of extraction and plant part can lead to the identification of different phytochemicals. For instance, in a study investigating the composition of Ziziphus nummularia leaves showed that 105 constituents were identified in the n-hexane extract consisting mainly of terpenoids, and 56 compounds were isolated from the ethanolic extract, mostly alkaloids and flavonoids with ethyl alpha-d-glucopyranoside as the main constituent ^[10]. Moreover, a comparison of the phytochemical composition between the leaves and fruits of various Ziziphus nummularia populations revealed that leaves were richer in phenols saponins and flavonoids ^[11]. And when it comes to the essential oils of this plant, these were shown to be rich in monoterpenes, aliphatic hydrocarbons, alkane hydrocarbons, primary terpene compounds, and decarbonated alcohols ^[12].

3.1. Anti-Drought and Anti-Thermal Characteristics of Ziziphus nummularia

Ziziphus trees and shrubs are able to thrive in extreme habitats, particularly in arid regions ^[13]. In the wake of climate change, drought has become a calamitous abiotic stress, affecting plant growth and hampering agricultural production and yield. The economic impact of this stress is likely to increase further; as a result, there has been growing interest in studying drought stress responses and tolerance mechanisms of natural drought-tolerant plants ^[14], such as Ziziphus nummularia. The purpose will be to develop crops with increased tolerance and resilience to drought. In a study to explore the physiological basis of drought, Ziziphus nummularia plants were subjected to moisture stress and results showed that chlorophyll a and b content in leaves decreased, and electrolyte leakage, which normally accompanies the plant’s response to stress, was increased ^[15]. Furthermore, in another effort to understand the molecular basis of abiotic stress tolerance mechanisms, isolate drought-responsive genes, and eventually develop drought resistant crop varieties, transcriptome profiling and gene expression analysis were carried out on Ziziphus nummularia seedlings subjected to drought stress caused by treatment with 30% polyethylene glycol (PEG 6000) ^[16][17]. In line, Padaria et al. ^[18] characterized the Ziziphus nummularia abscisic acid-stress-ripening gene 1 (ZnAsr1), and validated its transcriptional pattern in response to drought stress. This gene was upregulated in response to drought stress, and when recombinantly expressed in Escherichia coli, it allowed the bacteria to survive better in PEG-containing media, suggesting that the ZnASR1 may be used to develop drought tolerant transgenic crops. Sivalingam et al. ^[19] screened the parameters contributing to drought tolerance by comparing three Ziziphus nummularia ecotypes from semi-arid, arid and hyper-arid regions. Results showed that the latter was the most drought-tolerant ecotype and was associated with an increase in root length and number, an increase in leaf hair growth, and in elongation of thorns. The leaves were reduced in size and curled, while older leaves fell and shoot growth was sustained. In order to maintain a high relative water content for longer periods of time, membrane permeability was increased, and proline, catalase and sugar content was higher. Transcriptome profiling showed that genes associated with drought response mechanisms were constitutively expressed at higher levels ^[19]. Ziziphus nummularia also show tolerance to heat stress. Here, the chaperone protein ClpB1, which plays an important role in thermotolerance, was investigated ^[20]. It was found that the transcript levels of the JClpB1-C isoform from a heat-tolerant Ziziphus nummularia ecotype was highly up-regulated under heat stress conditions and that the overexpression of ZnJCIpB1-C in tobacco showed a significant increase in its heat tolerance by exhibiting higher photosynthetic rates, relative water content, chlorophyll content, and membrane stability index, suggesting that Ziziphus nummularia could provide an important source for developing heat-stress tolerant germplasms. Therefore, Z. nummularia can have an industrial potential as it enhances drought resistance in certain areas as well as having heat stress tolerance.

3.2. Pharmacological Properties of Ziziphus nummularia

3.2.1. Antimicrobial Activity

Antimicrobial resistance is nowadays a major global challenge. It occurs when infectious agents such as bacteria, viruses, fungi, and parasites become resistant to standard treatment, instigating a public health threat by spreading infectious diseases and causing huge global economic losses through food spoilage and damages to crops ^[21][22]. Interestingly, it was found that the fruit part of the plant displayed substantial antibacterial activity against various gram-positive strains tested ^[23]. Another study demonstrated that the chloroform and ethyl acetate fractions of Ziziphus nummularia were effective against both gram-negative and gram-positive bacteria, but the methanol and aqueous fractions did not exhibit any activity towards any of the tested microorganisms ^[24].  With regard to the antifungal activity of Zizuphus nummularia, it was shown that the ethanolic extract had strong activity against Aspergillus niger, Aspergillus flavus, Candida albicans, and particularly against Trichophyton rubrum ^[26]. All these data show that Ziziphus nummularia could be a potential source of novel antimicrobial agents that would support botanical screening efforts in the search for new drugs to overcome antimicrobial resistance and ensure antibiotic-free control of microbial growth in the future.

3.2.2. Anthelmintic Activity

Anthelmintic synthetic drugs are often associated with many side effects, toxic when inappropriately administered, and worrisome regarding the accumulation of drug residues in animals. Moreover, nematodes have developed resistance against many of these drugs, further fuelling interest in plants as sources for anthelmintic bioactive compounds ^[27]. Ziziphus nummularia has been traditionally used in veterinary medicine in an effort to control parasitic helminths in small ruminants and livestock. In line, Bachaya et al. have shown that Ziziphus nummularia crude methanolic extract is a rich source of chemicals effective against Haemonchus contortus, one of the most pathogenic nematodes of ruminants. The extract increased the mortality of worms, and inhibited egg hatching and larval development ^[28]. Its activity was also tested in vivo, where the same extract exhibited an 84.7% reduction in faecal egg count 13 days after treatment in sheep, a reduction comparable to that obtained with the synthetic drug levamisole ^[27]. This suggests that Ziziphus nummularia provides an economical and safe approach to search for new anthelmintics. It would be interesting to identify the constituents that are active against pathogenic nematodes, study the structure-activity relationship, and explore the underlying molecular mechanisms in order to assess their potential for drug discovery. Aging and many human diseases, such as cancer, inflammatory disorder, and neurodegenerative and digestive diseases, are associated with the overproduction of reactive oxygen species (ROS) or other free radicals ^[29][30][31]. These molecules are very reactive because of their unpaired electrons and, as a result, can cause cellular damage. Excessive amounts of ROS are mainly regulated by either endogenous antioxidant cellular mechanisms or antioxidants supplemented from exogenous sources. Hence, there has been an increasing interest in natural exogenous sources of antioxidants including plant sources. Plant antioxidant sources are regarded as a safer alternative to synthetic antioxidants such as butyl hydroxy anisole (BHA) and butylated hydroxytoluene (BHT) ^[32][33]. The methanolic and acetone extracts from the leaves of Ziziphus nummularia showed significant scavenging activities of the free radicals 1,1-diphenyl-2-picryl-hydrazyl (DDPH) and superoxide anion radicals. In addition, a high reducing power was recorded, supporting the strong antioxidant potential of this plant ^[34]. In another study, the methanolic extract from the fruit part of Ziziphus nummularia, which contains a high phenolic and flavonoid content, was shown to have a significant antioxidant activity against DDPH and hydrogen peroxide (H₂O₂) ^[35]. Similar results were obtained with the hydro-alcoholic extract from the stem bark, rich in phenolics ^[36], and with the methanolic extract of roots ^[37]. Finally, an investigation of the antioxidant capacity of methanolic extracts from different Ziziphus nummularia genotypes showed that the antioxidant potential was directly correlated with the extract’s phenolic content ^[38]. Hence, Ziziphus nummularia exhibits potent antioxidant capacities that warrant further investigation.  Inflammation is part of the body’s homeostatic and self-defence mechanism, and is always kept under homeostatic regulation. However, when inflammation becomes excessive or chronic, it may contribute to the development of several diseases such as diabetes, cardiovascular disease, and cancer ^[39][40]. Using the carrageenan induced paw edema test, the topical application of a gel ethanolic extract from Ziziphus nummularia leaves to Wistar albino rats caused a significant reduction in paw edema in a dose-dependent manner, confirming the anti-inflammatory effect of the extract ^[41]. It also showed faster wound repair abilities when compared to the marketed formulation Betadine^{® [41]}. Dey Ray et al. orally administered the ethanolic extract from the root bark into mice and found it to cause a significant inhibition of carrageenan and arachidonic acid induced edema, and reduce the formation of granuloma tissue, further confirming the anti-inflammatory properties of the plant ^[42]. Moreover, octadecahydro-picene-2,3,14,15-tetranone, a compound isolated from the root bark of Ziziphus nummularia, had a more pronounced anti-inflammatory than the extract, nearly comparable to that of aspirin. It was suggested that this compound controls inflammation through the inhibition of TNF-α and nitric oxide production ^[42]. Goyal et al. found that the anti-inflammatory effect of the ethanolic extract of Ziziphus nummularia was due to the inhibition of histamine release ^[43], which initially causes vasodilation and increases vascular permeability. This extract also inhibited peritoneal leukocyte migration ^[43]. Finally, a recent study demonstrated that the ethanolic extract of the leaves of Ziziphus nummularia exhibited an anti-inflammatory effect in human aortic smooth muscle cells by decreasing their proliferation, and invasion and migration potentials. Importantly, the extract caused a concentration- and time-dependent decrease in the pro-inflammatory effect of tumor necrosis factor α (TNFα). The extract reduced the TNFα-induced expression of matrix metalloproteases, NF-κB, and cell adhesion molecules ^[44]. In fact, upon inflammatory stimuli, vascular smooth muscle cells switch from a quiescent phenotype to a synthetic one, entailing detachment of these cells from the extracellular matrix and their migration to the tunica intima layer of the vessel, contributing to the formation and growth of atherosclerotic lesions. By this, the study suggests that Ziziphus nummularia extracts reduce inflammation and thereby may reduce inflammation-induced atherogenic atherosclerosis ^[44]. Cancer continues to be one of the leading causes of mortality worldwide despite great advancements in cancer therapy. Moreover, conventional treatment regimens are often associated with adverse effects and multidrug resistance that have resulted in increasing interest in the search for new bioactive compounds from plant sources ^[45]. In this regard, several studies have assessed the anticancer properties of Ziziphus nummularia. Lapachol (2-hydroxy3-(3-methy1-2-buteny1)-1,4-naphthoquinone), a naphthoquinone compound isolated from Ziziphus nummularia, was shown to have a strong antitumor activity in female Swiss albino mice engrafted with sarcoma ascetic tumor cells (S-180 cells). Lapachol also enhanced the response of the engrafted tumors to radiation therapy, suggesting that it can have a potential use as an adjuvant in radiation therapy ^[46]. Another phytoconstituent isolated from the root bark of Ziziphus nummularia is a new triterpene derivative referred to as Identified Compound (IC) ^[47]. IC showed high cytotoxicity in several human cancer cell lines in vitro, including breast cancer, leukaemia, ovarian cancer, colon adenocarcinoma, and kidney carcinoma. This anticancer activity was higher compared to the plant’s crude ethanolic extract. In vivo, both IC and the ethanolic extract were administered to mice, engrafted with Ehrlich ascites carcinoma, and were found to decrease tumor parameters including decreasing the viable cancer cell count, restoring serum biochemical parameters, and decreasing ascitic fluid volume, thereby inhibiting cancer growth and increasing the life span of mice. The treatment also decreased the count of white blood cells, increased the count of red blood cells, and elevated haemoglobin content ^[47]. In another study, the methanolic fruit extract of Ziziphus nummularia showed appreciable cytotoxic activity against cervical carcinoma cells, HeLa cells ^[48]. Furthermore, the ethanolic extract from Ziziphus nummularia leaves was shown to inhibit the hallmarks of pancreatic cancer by interfering with tumorigenic and metastatic events ^[49]. Indeed, treatment of capan-2 pancreatic cancer cells resulted in an attenuation of cell proliferation and a significant decrease in their migration and invasion potentials. The same extract was also able to attenuate angiogenesis by reducing the production of secreted and intracellular vascular endothelial growth factor (VEGF), a pro-angiogenic protein, as well as the levels of nitric oxide. This anti-angiogenic effect was further confirmed by in ovo experiments using the chick embryo chorioallantoic membrane assay ^[49]. These studies have shown that Ziziphus nummularia extracts and the isolated compounds described above exhibit strong anticancer activities against various types cancers and target the hallmarks of cancer. As such, Ziziphus nummularia proves to be an effective and safe source of bioactive compounds with anticancer properties. Diabetes, a very common chronic disease associated with a deregulation of blood sugar, has been on the rise, especially in low- and middle-income countries, and is associated with significant mortality and morbidity rates. Over time, diabetes, if left uncontrolled, can cause serious damage to the blood vessels and nerves, leading to blindness, kidney failure, and cardiovascular complications among others ^[50]. Before the discovery of insulin, medicinal plants were widely used in the treatment of diabetes; today diabetic patients are showing more interest in herbal remedies and natural compounds due to their cost-effectiveness, availability, and being regarded as safe with limited side effects ^[51]. As prolonged hyperglycaemia during diabetes leads to abnormalities in lipid profiles and is associated with atherosclerosis, Rajasekaran et al. investigated the anti-hyperglycaemic and hypolipidemic effects of Ziziphus nummularia ethanolic and aqueous leaf extracts in alloxan-induced diabetes in rats ^[52]. Results showed that both extracts, the ethanolic extract in particular, reversed pancreatic damage, significantly decreased blood sugar levels, and reduced the levels of triglycerides (TGL), low density lipoprotein (LDL), very low density lipoprotein (VLDL), and total cholesterol (TC). Levels of cardioprotective high density lipoprotein (HDL) increased ^[52]. Similar results were obtained in another study ^[53] and in a dexamethasone-induced diabetic rat model ^[54], further supporting the importance of Ziziphus nummularia as a source of anti-hyperglycaemic and hypolipidemic agents in the treatment of diabetes. Dubey et al. tested the anti-diabetic potential of the aqueous, methanolic, and saponin extracts of the leaves and found that all the extracts, particularly the saponin extract, had a strong anti-diabetic activity that inhibited α-amylase. They proposed that Ziziphus nummularia may be a good source for pharmacologically active agents of the metabolism of carbohydrates, and hence the management of diabetes ^[55]. Furthermore, a comparison of the anti-diabetic activity of hydro-alcoholic extracts from the leaves and fruits of Ziziphus nummularia using the Soxhlet and maceration methods was carried out. All extracts showed significant inhibitory activity, but leaf extracts from the maceration process showed the strongest activity and were associated with the highest phenolic content ^[56]. Alzheimer’s disease is a neurodegenerative disorder of the brain, clinically characterized by a progressive decline in cognitive abilities, and is the most common cause of dementia. It is associated with a significant reduction in central cholinergic neurotransmission due to a loss or deficiency of the neurotransmitter acetylcholine. Restoring acetylcholine levels through the inhibition of the enzymes which hydrolyze acetylcholine, acetyl cholinesterase (AChE) and butyrylcholinesterase (BChE), is used as the main treatment strategy. However, these treatments are often associated with side effects such as nausea and vomiting ^[57]. As a rich source of phenolic compounds, Uddin et al. investigated the anticholinesterase activity of Ziziphus nummularia using methanolic extracts of the fruits of different genotypes ^[38]. The extracts showed significant activity against both AChE and BChE, suggesting that Ziziphus nummularia could be used as a source for potential therapeutic agents in the treatment of neurodegenerative diseases. Differences observed among the different genotypes were attributed to variations in the composition and concentration of their bioactive constituents and secondary metabolites ^[38]. Ziziphus nummularia has been widely prescribed in traditional medicine for relieving pain. Hence, Goyal et al. investigated its analgesic effects and showed that ethanolic extract from leaves of Ziziphus nummularia significantly reduced the number of writhes of mice in a visceral pain model that involved acetic-induced abdominal writhing in mice. The pain relief was attributed to the anti-inflammatory properties of Ziziphus nummularia ^[43][58]. This extract also increased tail flick latency in mice, indicating that its analgesic activity is mediated by central mechanisms ^[43][58]. Administration of the cyclopeptide alkaloid fraction of the ethanolic extract of the leaves of Ziziphus nummularia in mice subjected to acetic acid-induced pain, confirmed the analgesic properties of this plant in addition to possessing significant anti-nociceptive effects ^[58].  Ziziphus nummularia has been traditionally used in the treatment of gut disorders such as diarrhea, gastrointestinal spasms, and ulcer. A study by Sayed et al. was carried out to evaluate these ethnopharmacological properties of the methanolic crude extract of Ziziphus nummularia leaves ^[59]. The extract also caused a relaxation of spontaneous KCl-induced contractions in an isolated rabbit jejunum model by blocking Ca²⁺ influx, in a manner similar to that of the drug verapamil ^[59]. Finally, Ziziphus nummularia was shown to also have anti-ulcer properties by decreasing lesions and protecting the gastric mucosa in an ethanol-induced gastric ulceration assay comparable to that of the standard drug omeprazole, which is widely used in the treatment of stomach ulcers as well as indigestion, heart burn and acid reflux ^[59]. Nanotechnology, which involves the development and controlled manipulation of molecules with length scales in the 1- to 100-nanometers range, is an emerging field that is rapidly gaining impetus with applications in drug delivery, molecular imaging, catalysis, and for diagnostic purposes, among others ^[60]. Recently there has been growing interest in the “green synthesis” of nanoparticles through the use of microbes, plants, or plant parts for the bioreduction of metal ions into their elemental nanoparticle form ^[61][62][63]. Not only has green synthesis given more stable nanoparticles compared to other methods, but also it has been scalable and eco-friendly without any requirement for high temperatures and pressure or the use of toxic chemicals. Silver nanoparticles are mostly used as anti-microbial agents in the medical field mainly due to their strong growth inhibitory effects against fungi, bacteria and viruses, while having low toxicity to human cells. These nanoparticles have been utilized in food storage, textile coatings, as well as water treatment ^[64]. Khan et al. were the first to synthesize silver nanoparticles using Ziziphus nummularia aqueous leaf extract as the reducing and capping agent ^[65]. Results showed that the prepared nanoparticles exhibited better antioxidant activity compared to the leaf extract. Antibacterial activity against Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Salmonella typhi, and Bacillus cereus, as well as antifungal activity against Aspergillus niger, Aspergillus flavus and Candida albicans were also enhanced. Similar enhancements were reported in another study, also using Ziziphus nummularia aqueous leaf extracts, where increased antioxidant and antibacterial activity against the cariogenic Staphylococcus aureus and Streptococcus mutans ^[66], Bacillus subtitles, Pseudomonas aerugionsa, Klebsiella pneumonia, Escherichia coli and Streptococcus aureus were reported ^[6]. These nanoparticles also showed good fungicidal activity against Aspergillus niger and Aspergillus flavus ^[6]. Gold nanoparticles, which are also known to possess antimicrobial activities ^[67], were also synthesized using Ziziphus nummularia aqueous leaf extracts. These showed enhanced antioxidant potential and anti-microbial activities, similar to the results obtained with silver nanoparticles ^[66]. Zinc oxide (ZnO) nanoparticles are widely used in various medical applications such as drug delivery and bio-imaging ^[68] and are known to exhibit anti-microbial activities ^[69]. The anti-microbial properties of ZnO nanoparticles synthesized with the aid of Ziziphus nummularia leaf extracts were tested against clinical isolates of Candida species (Candida albicans, Candida glabrata) and Cryptococcus neoformans ^[70]. Results showed strong fungicidal activity, better than the leaf extract and the standard antibiotic amphotericin B. The prepared ZnO nanoparticles also exhibited a dose-dependent cytotoxic effect against HeLa cancer cells, higher than the Ziziphus nummularia leaf extract ^[70].