Oyere Tanyi Ebob ·Smith B. Babiaka ·Fidele Ntie-Kang ,2

Abstract For the past 2 years, the coronavirus responsible for the COVID-19 infection has become a world pandemic, ruining the lives and economies of several nations in the world.This has scaled up research on the virus and the resulting infection with the goal of developing new vaccines and therapies.Natural products are known to be a rich source of lead compounds for drug discovery, including against infectious diseases caused by microbes (viruses, bacteria and fungi).In this review article, we conducted a literature survey aimed at identifying natural products with inhibitory concentrations against the coronaviruses or their target proteins, which lie below 10 μM.This led to the identifi cation of 42 compounds belonging to the alkaloid, flavonoid, terpenoid, phenolic, xanthone and saponin classes.The cut off concentration of 10 μM was to limit the study to the most potent chemical entities, which could be developed into therapies against the viral infection to make a contribution towards limiting the spread of the disease.

Keywords Antivirals·Drug discovery·Inhibitors·Lead compounds·Natural products·SARS-CoV-2

1 Introduction

The novel coronavirus disease 2019 (COVID-19) has led to a sudden change in the lifestyle of humans all over the whole world [1– 4].The World Health Organization (WHO) declared the disease resulting from the virus to be a pandemic, which has prompted an exponential increase in scientifi c research towards fi nding a drug or a vaccine to limit its spread and the number of casualties.At the time of writing this manuscript, the entire world had experienced about 150 million new infections, about 120 million recoveries and almost 3 million deaths [5].Two earlier coronaviruses were associated with severe acute respiratory syndrome (SARS) and Middle East respiratory syndrome (MERS).The symptoms associated with COVID-19 include (as in the earlier respiratory syndromes; SARS and MERS) fever, cough, dizziness, and shortness of breath, which might lead to pneumonia and acute respiratory distress, causing death [6,7].The virus associated with SARS, MERS and COVID-19 are air-borne and transmissible by contact with infected persons.However, COVID-19 has surpassed the earlier two syndromes in terms of the number of individuals infected and the number of deaths [6].

Coronaviruses (CoVs) belong to the family Coronaviridae, sub-family Coronaviridae and order Nidovirales.Viruses belonging to this family and sub-family are known to be large in terms of genome size (26−32 kb) [8], are enveloped, have a single-stranded ribonucleic acid (RNA) and can infect both animals and humans.CoVs can be subdivided into four genera, i.e., alpha (α), beta (β), gamma (γ) and delta (δ) coronaviruses, according to their genotype and serology [9].Presently all CoVs that can cause infection in humans belong to the fi rst two categories, i.e., the alpha-coronaviruses (α-CoVs) and the beta-coronaviruses (β-CoVs).Examples of α-CoVs include the human coronavirus Netherland63 (HCoV-NL63) and human coronavirus 229E (HCoV-229E), named after a student specimen coded 229E, while examples of β-CoVs include the human coronavirus Organ Culture43 (HCoV-OC43), the human coronavirus Hong Kong University 1 (HCoV-HKU1), SARS-CoV and MERSCoV [9].

Even though research on the structure and mode of action of the SARS-CoV-2 virus is ongoing, several known antiviral drugs (e.g.lopinavir/ritonavir, darunavir/umifenovir, oseltamivir, favipiravir, remdesivir, etc.) have been used to manage cases of SARS-CoV infection, together with some anti-malarials like chloroquine and hydroxychloroquine, the antibiotic azithromycin, etc.[10– 12].Besides, many local communities have made claims that some medicinal plants have been successfully used to treat patients who showed some signs of COVID-19 infection [13,14].A recent review revealed that 92% of 135 hospitalized patients in northeast Chongqing, China have used traditional medicine in combination with conventional medicine as treatment for COVID-19 [15].This is because medicinal plants contain natural products (NPs) which hold a huge potential to be used as lead compounds for the development of new drugs [16].This could include drug leads for treating diseases caused by CoV activity, with the goal of developing them into novel COVID-19 drugs.

Several excellent reviews have recently been published on NPs with the potential for treating SARS-CoV-2 infection [17– 19].Other reviews have focused on antiviral agents with potential for the drug development for the treatment of COVID-19 using crude extracts from plants [20,21].However, the focus of such reviews was on the activities of the crude extracts with little emphasis on the isolated bioactive compounds.In some other cases, only in silico hits, i.e.compounds that have been predicted computationally (via molecular docking, molecular dynamics, pharmacophore searches, etc.) have been described [19].The current study is intended to provide an update of the isolated NPs with the potential to be developed into drugs for the treatment of COVID-19, i.e.compounds showing activities at less than 10 μM against the coronaviruses and/or their target proteins in experimental assays that has been recently published in literature.In some cases, the drug target to which the compounds bind, have been presented in the discussion.The presentation of the compounds is according to the major compound classes, which have been arranged alphabetically.

2 Methods

All pertinent information about the botanical description, conventional uses, phytochemicals and the pharmacological activities of the isolated compounds were collected from available recently published literature.The electronic databases employed for the assortment of relevant information include Scopus, NISCAIR, Scifi nder, PubMed, Springer Link, Science Direct, Google Scholar, Web of Science, and an exhaustive library search.The chemical structures of the compounds were drawn using ChemDraw Ultra 8.0 software.PubChem and ChemSpider databases have been used to check the IUPAC names of the isolated phytoconstituents.The workflow for collecting the literature and writing the report has been provided in Fig.1.

3 Alkaloids

Alkaloids are nitrogen-containing compounds, mostly known to be bitter principles with diverse biological activities.A summary of alkaloids with potential for drug development against SARS-CoV-2 has been provided in Table 1.Berberine ( 1), Fig.2, is an isoquinoline alkaloid, known to be biosynthesized by many plant species, particularly those of the genusBerberisfrom which the compound derives its name.Some medicinal properties of 1 reported include anti-infl ammatory [40], antiviral [41], antidiabetic [42], antihypertensive [43], hepatoprotective [44] and anticancer [45], additionally is a widely used dietary supplement due to these medicinal properties.According to Warowicka et al.compound 1 could be a possible lead compound against SARS-COV-2, which might act by modulating the nuclear factor kappa of activated B cells (NF-κB) and mitogen-activated protein kinases (MAPKs) [46].NF-κB is a protein required for DNA transcription and many other cell processes including infl ammatory and immune responses.The modulation of NF-κB path is one of the ways by which the alkaloid biberine could inhibit virus infections, in addition to its anti-infl ammatory properties [47].Compound 1 has aninhibitory eff ect on the NF-κB signaling pathway and therefore might function as an antiviral agent against coronavirus infection.Moreover, during viral infection, the virus regulates expression of infl ammatory mediators such as tumor necrosis factor (TNF) [48,49].Moreover compound 1 is also known to inhibit herpes simplex virus (HSV) infection both the HSV-1 and HSV-2 virus with EC50values of 6.77 ± 1.13 μM and 5.04 ± 1.07 μM, respectively in a dose dependent manner [50].

Table 2 Diarylheptanoids

Table 4 Phenolics

Table 5 Saponin

Table 6 Terpenoids

Table 7 Xanthone

Cepharanthine ( 2), fangchinoline (3) and tetrandrine (7) are bis-benzylisoquinoline alkaloids.Stephania tetrandraand other species of the genusMenispermaceaeare major sources of bis-benzylisoquinoline alkaloids which are widely distributed in China and many East Asian countries [51].A recent review showed the use ofS.tetrandrain traditional Chinese medicine as antitumour, neuroprotective, anti-inflammatory, and antibacterial [52].Compound 7 exhibits a wide range of pharmacological activities which include anticancer, anti-infl ammatory [51] it is also reported to have inhibitory activity against Ebola virus; herein, tetrandrine acts as a closure of key calcium sensor called two pore channels which remain open during viral infection through endosomal or membrane bound route.[53].Compound 2 has antiviral activities against human immunodefi ciency virus type 1 (HIV-1) virus [54] and HSV-1 virus [55], while compound 3 replication of HIV-1[56].A time addition assay with MRC-5 cells infected with HCoV-OC43 was used to investigate the antiviral activity of compounds 2,3 and 7, the compounds strongly inhibited HCoV-OC43 virus with IC 50 values of 0.83, 1.01 and 0.33 μM, respectively.Additionally these compounds have no cytotoxicity eff ect on MRC-5 cell at concentrations up to 10 μM [22].

Fig.1 Workflow implemented in this project

Indigodole B (4) and tryptanthrin (8) have been isolated from methanol extract of the medicinal herbStrobilanthes cusia[57,58], which is commonly found in northeast India, Bangladesh, southern China, Myanmar, and Taiwan [59].The roots and leaf extracts of the plant have been used in traditional herbal medicine based on its anti-infl ammatory, antimicrobial, and antiviral activities [60,61].According to Tsai et al.the methanol extract ofS.cusiareduced HCoV-NL6 viral production with an IC50value of 0.12 μg/mL [24].Compounds 8 and 4 had high activity against HCoV-NL63, with compound 8 being reported to be the most potent compound, reducing HCoV-NL63 viral production with IC50= 0.06 μM.Meanwhile, compound 4 exhibited an eff ective virucidal activity against HCoV-NL63 with an IC50of 2.09 μM [24].Compound 8 specifi cally altered the antigenic structure of the viral spike protein and further inhibits the cleavage activity of the proteolipid protein 2 (PLP2) gene, associated with virucidal activity, and inhibits the post-entry stage of HCoVNL63 replication [24].

Cyclopeptide alkaloids are among the common compounds in plants belonging to the Rhamnaceae, especiallyZiziphusgenus, e.g.,Z.jujuba[62].The cyclopeptide alkaloids jubanine H (5) and nummularine B (6) have been isolated from the stem bark of this species [62,63].Compound 6 is also known for its moderate in vitro antimalarial activity (againstPlasmodium falciparum), withan IC50of 10.3 μM [64].Compounds 5 and 6 have been tested against porcine epidemic diarrhea virus (PEDV) in

Vero cells, with their EC50values established as 4.49 and 6.17 μM, respectively [17].

Table 1 Alkaloids

Fig.2 Chemical structures of promising alkaloids ( 1 to 8) with potential against SARS-CoV-2

Fig.3 Chemical structures of promising diarylheptanoids ( 9 to 13) with potential against SARS-CoV-2

4 Diarylheptanoids

Diarylheptanoids, also known as diphenylheptanoids, are plant secondary metabolites derived from many plant species.They are made up of phenolic aromatic rings fused together by linear seven carbon chains.They can be either open chain or macrocyclic diarylheptanoids [65].Diarylheptanoids possess numerous therapeutic benefi ts including anti-infl ammatory [66], antioxidant [67], anti-microbial [68] and anti-diabetic [69] activities.A summary of this class of compounds with potential for drug discovery against SARSCoV-2 is provided in Table 2, while the chemical structures of the compounds are shown in Fig.3.

In a study to discover new compounds that inhibit SARSCoV, Park et al.found the ethanol extract of the stem back ofAlnus japonicato exhibit PL pro inhibitory [25].A.japonicahas a lot of pharmacologic properties, including anti-infl ammatory [70], antioxidant [71] and anti-infl uenza activities [72].The isolated diarylheptanoids of interest include hirsutenone ( 9), hirsutanonol ( 10), rubranoside B ( 11) and rubranoside A ( 12) [25].The isolated diarylheptanoids were tested against SARS-CoV PLprousing a continuous fluorometric assay and showed a dose-dependent inhibitory eff ect against the PLpro[25].The compounds were found to be reversible inhibitors because an increase in concentration rapidly reduced enzyme activity.For compounds 9 and 10, it was found that α,β-unsaturated carbonyl and catechol groups may play a pivotal role in SARS-CoV PLproinhibition by interacting with the PL pro nucleophiles while the monohydroxy substitution led to drop in the inhibitory eff ect [73].When the compounds were tested against SARSCoV 3CLpro, the fi ndings showed that the diarylheptanoids displayed a signifi cant selectivity towards the 3CL pro proteases [25].The IC50values are 4.1, 7.8, 8.0 and 9.1 μM, respectively.The known viral protease inhibitor curcumin ( 13) was used as a reference inhibitor (with an IC50value of 5.7 μM) [74].

5 Flavonoids

The potential anti-COVID flavonoids (Figs.4,5 and Table 3) include tomentins A ( 22), B ( 23), E ( 24), 4′-O-methyl diplacol ( 14), and diplacone ( 16), which are geranyl flavonoids isolated from the fruits ofPaulownia tomentosa[26,27].P.tomentosais widely distributed in China and parts of this plant species (the bark, fruits, xylem, and leaves) have been used in traditional Chinese medicine (TCM) to treat several ailments, including tonsillitis, bronchitis, asthmatic attacks, and dysentery [75,76].The anti-infl ammatory property of the plant has been most exploited [77].Besides, some compounds isolated from the fruits of this species have demonstrated the ability to inhibit airway infl ammation [78], while other compounds are known to exhibit cytotoxic [79], antimicrobial and antioxidant activities [80].The antiviral properties of the fruit extract and the isolated compounds from this species have been investigated in vitro against the polyprotein target papain like protease (PL pro ), a protein involved in RNA replication [26].All the compounds from this species displayed a dose dependent inhibition of SARS-CoV PLpro, compound 22 being the most potent with an inhibitory constant,K i = 3.5 μM [26].The IC50values of the compounds are listed in Table 3.

Fig.4 Chemical structures of promising flavonoids ( 14 to 23) with potential against SARS-CoV-2

The ethanol extract ofAngelica keiskeihas shown eff ective inhibition against 3-chymotrypsin-like protease (3CLpro) and PLpro(with 75% and 88% inhibition at 30 μg/mL, respectively) [31].A.keiskeiis a large perennial plant widely distributed in Japan,Korean and China.Its traditional uses include as a tonic, diuretic, laxative, and analeptic [81].The plant extracts have exhibited several biological activities, including antitumor and antimetastatic eff ects [82].The compound xanthoangelol E ( 20) was isolated fromA.keiskei[31].Park et al.subjected this compound to fluorescence resonance energy transfer (FRET) and cell-based cis-cleavage inhibition assay to measure the SARS-CoV 3CLproand SARS-CoV PLproinhibition in vitro [30].The compound showed a dose-dependent inhibition both in the SARS-CoV PLproand SARS-CoV 3CLpro, with IC50values of 11.4 and 1.2 μM, respectively [30].These results show that compound 20 has specifi c inhibitory activity against the cysteine protease.

The crude ethanol extract ofTorreya nucifera(a plant found in a snowy area near the sea of Jeju island in Korea) was subjected to FRET assay to measure the inhibitory eff ectiveness against SARS-CoV 3CLpro[83].The results showed 62% inhibition at 100 μg/mL [27].The biofl avonoid amentofl avone ( 15) isolated from this species [27], has shown wide pharmacological importance in the treatment of Alzheimer [84] and in the treatment of bladder cancer [77].Compound 15 also showed potential antiviral activity against the syncytial virus (RSV), with IC 50 = 5.5 μg/mL [85].The compound also showed inhibitory eff ects against the hepatitis C virus (HCC) [86], as well as revealed antiviral activity against infl uenza-A and infl uenza-B viruses, although only showing moderate activity against the herpes viruses (HSV-1 and HSV-2) [87,88].Moreover, a computational study revealed compound 15 has a good binding affi nity against the NS2B–NS3 protease protein in docking simulation (binding affi nity of − 9.0 kcal/mol) and showed signifi cant inhibition of the Zika virus from the modelling studies [89].Ryu et al.subjected compound 10 to a fluorescence resonance energy transfer (FRET) assay and showed that this compound displayed a dose-dependent inhibitory activity of SARS-CoV 3CL pro [27].

Fig.5 More chemical structures of promising flavonoids ( 24 to 26) with potential against SARS-CoV-2

Myricetin ( 18) is polyhydroxy flavonoid fi rst isolated as a yellow-coloured crystal in the late eighteenth century from the back ofMyrica nagiharvested from India [90].The compound is an important element in a variety of human foods, including vegetables, tea and many fruits.Compound 18 is known for its iron-chelating, antioxidant, anti-infl ammatory, and anticancer properties [91].Yu et al.examined the inhibitory eff ect of compound 18 and scutellarein ( 21) against SARS-CoV helicase, Nsp13, hepatitis C virus (HCV) helicase by using the FRET- bases double stranded DNA unwinding assay as well as using a colorimetry-based ATP hydrolysis assay [29].Compound 21 was isolated fromScutellaria baicalensis(commonly known as Chinese skullcap), which is traditionally used for treating infl ammation and respiration [92].The results showed that compounds 18 and 21 potently inhibited SARS-CoV helicase protein in vitro by acting on ATPase with IC 50 values of 2.71 and 0.86 μM, respectively, although the compounds did not suppress the helicase activity of the HCV virus [29].Yu et al.modelling analysis revealed that compounds 18 and 21 could fi t in and directly interact with ATP/ADP binding pocket of SARS-CoV helicase protein inhibiting the direct binding of ATP/ADP [29].Moreover, it was observed that compounds 18 and 21 didn’t exhibit cytotoxicity against normal breast epithelial cell lines (MCF10A) [29].

Quercetin ( 20) is a polyphenolic flavonoid found in several vegetables and fruits such as berries, apples, and onions [93].The compound has been found to have many pharmacological properties, including anti-infl ammatory activity by cyclooxygenase inhibition [94– 96], lipoxygenase [96,97], expression of cyclooxygenase and production of prostaglandin E2(PGE2), as well as reducing production of interleukin (IL)-1α [98].The compound also showed antioxidant activity via it's radical scavenging ability [99] and possesses anti-hypertensive activity [100,101].Park et al.demonstrated the antiviral activity of compound 20 against SARS-CoV PL pro with an IC50value of 8.6 μM [30].No cellbased assay of antiviral activity was carried [30].Besides, compound 20 showed the capacity to block the entry of SARS-CoV into host cells and further antagonized HIVluc/SARS pseudo typed virus entry with an EC 50 of 83.4 μM [34].Additionally, compound 20 showed cytotoxicity, with CC50= 3.32 μM [34].

Hesperetin ( 17) is a flavone glycoside commonly found in citrus fruits [102].Recent studies have shown that antioxidant activity of compound 17 is not limited to its radical scavenging eff ects, but also boosts antioxidant cellular activity through extracellular signal regulated kinases/nuclear erythroid 2-related factor 2 (ERK/Nrf2) signaling pathway [28].According to Lin et al.compound 17 showed a dosedependent inhibited cleavage activity of SARS-CoV 3CL pro in cell-based assay [103].In addition, the compound is less cytotoxic in Vero cells [103].In silico studies have shown that 17 binds with high affi nity to helicase, spike protein and protease side on the ACE2 receptors used by SARS-CoV-2 to cause COVID-19 (Fig.6), suggesting that this compound could be a potential inhibitor of coronavirus cell growth [104].

Fig.6 A Chemical structure of hesperetin ( 17); B The docked pose of the compound towards the SARS-CoV-2 ACE2 receptor binding site for the identifi cation of its putative binding mode.This fi gure has been reproduced without copyright permission from MDPI because it was originally published under an open access license [104]

Papyriflavonol A ( 19) is a prenylated flavonol fromBroussonetia papyrifera[105].B.papyriferais a deciduous tree, whose extracts exhibit antifungal [106], antioxidant [107] and antihepatotoxic properties [108].The fruits of this plant have been used to treat ophthalmic disorder in China, with its effi cacy being proven by pharmacologic experiments [109].Compound 19 showed a dose-dependent inhibitory eff ect on SARS-CoV PL pro (IC 50 = 3.7 μM), when subjected to the fluorogenic peptide Z-RLRGG-AMC assay [30].The compound also shows a dose-dependent inhibitory eff ect on both α-glucosidase and cysteine proteases [30].It is known that PL pro exhibits deubiquitinating (DUB) activity and antagonizes the induction of type-1 interferon (IFN), the interferon-stimulated gene 15 (ISG15) is the most overexpressed gene upon IFN stimulation and it’s involved in marking newly synthesised protein during an antiviral response.Both ubiquitin and ISG15 are important for viral replication and pathogenesis, SARS-CoV PL pro can cleave ubiquitin and ISG15 from cellular conjugates, additionally compound 19 strongly inhibits both ubiquitin and ISG15 with an IC 50 values of 7.6 and 8.5 μM, respectively [30].

Du et al.reported the antiviral activity of leachianone G ( 26) against herpes simplex type 1 virus (HSV-1) [32].Compound 26 is a prenylated flavonoid present in many plant species particularlyMorus alba[32].This species is widely distributed in India, China, Japan, and Southern Europe [110].M.albais also a rich source of phenolic compounds, including flavonoids and anthocyanins which are of great pharmacological and biological importance because of their antioxidant properties [111].The plant has been used for the treatment of type 2 diabetes mellitus due to its hypoglycemic eff ects [112].Moreover,M.albais also known to possess other known antiviral activities, e.g., against rhinovirus [113], dengue virus [114] and hepatitis B virus [115].The compound showed potent antiviral activity against HSV-1 (IC50= 4.49 μM).Its cytotoxicity tested on Vero cells showed an IC50value of 250 μM.The known antiviral drug (acyclovir), which was used as a positive control in this experiment, also showed potent anti-HSV-1 activity (IC50= 3.65 μM).The antiviral activities and cytotoxic eff ects of compound 26 as well as for the control antiviral drug (acyclovir) were determined using the viral cytopathic eff ect assay [32].

6 Phenolics

Phenolics are known to be very important dietary components and are strong natural antioxidants [116].They also exhibit anti-infl ammatory, anti-cancer [117], antimicrobial [118] and other pharmacological properties.This class of secondary metabolites contains one or many hydroxyl groups attached directly to a benzene ring, including the flavonoid class earlier discussed [119].In the paragraphs beneath, we focus the discussion on non-fl avonoid-based and non-diarylheptanoid phenolic compounds.A summary has been provided in Table 4, with chemical structures shown in Fig.7.

Pentagalloylglucose ( 27) is a polyphenol isolated from the branches and leaves ofPhyllanthus emblica[120].It is an indigenous tree in Southeast Asia (also known as Indian Gooseberry or Amla).The extract ofP.emblicais known to possess great pharmacologic properties, such as anticancer, antitumor and antioxidant activities [121,122].Compound 27 has been reported to have several medicinal properties, e.g., as an anticancer agent, since the compound could elicit rapid and selective cytotoxicity in cancer cells [123].Computational studies reveal that compound 27 could inhibit viral entry by binding to Zika virus envelope protein [124].Additionally, this compound inhibits the early steps of hepatitis C virus [124], as well as reduces the growth of hepatitis B virus [125] and shows antiviral activities against respiratory syncytial virus [126].According to Pei et al., the EC50value of compound 27 was measured to be 4.12 μM, using the XTT and plaque reduction assay.Compound 27 shows strong inhibitory activity in early and late stages of HSV-1 virus, inhibiting gene replication, transcription, and related structural changes [33].

Fig.7 Chemical structures of promising phenolics ( 27 and 28) with potential against SARS-CoV-2

Fig.8 Ecsin ( 29), a promising saponin with potential against SARSCoV-2

Yi et al.investigated the antiviral activity of tetra-O-galloyl-beta- D-glucose ( 28) using a colorimetric assay for assessing cell metabolic activity (MTT) assay against SARSCoV [34].Compound 28 also expresses an eff ective SARSCoV inhibition with an EC 50 = 4.5 μM and a cytotoxicity value of 1.08 μM using Vero E6 cell following the MTT assay.The results suggest that 28 could be used at concentration to inhibit SARS-CoV without a considerable cytotoxic eff ect [34].

7 Saponin

Saponins are naturally occurring bioorganic compounds having at least one glycosidic linkage (C-O-sugar bond) between an aglycone and a sugar chain.Hydrolysis of a saponin molecule produces two portions, aglycone and a sugar moiety.Specifi cally, they are naturally occurring glycosides described by the soap-like foaming, and consequently, they produce foams when shaken in aqueous solutions.They are known to exhibit biological properties such as antibacterial, antifungal, antiviral [127] and anti-infl ammatory [127,128].

Escin ( 29), Fig.8 and Table 5, is a major principle from horse chestnutAesculus hippocastanum[129].The plant has been used in traditional medicine to treat several conditions, including hemorrhoids [130], postoperative edema [131], venomous congestion [132] and anti-infl ammatory action [133].Compound 29 was fi rst isolated in the year 1953 [129] and its pharmacologic and biological properties include anti-infl ammatory [134], anti-edematous [132] and preventing the hypoxic damage of the endothelium [135].According to Wu et al.this compound showed a potent antiviral activity against SARS-CoV-3CL pro , its measured EC50value being 6 μM and it had a cytotoxicity value of 15 μM [35].A cell-based assay with SARS-CoV and Vero E6 cells was used to measure this activity.

8 Terpenoids

Terpenoids constitute a diverse class of NPs biosynthesized from the condensation of isoprene units to yield terpenes.In plants, terpenoids can serve in communication and defense, for example they act as attractants for pollinators and seed dispersers (chemoattractants or chemorepellents) [115].Terpenoids are known to possess diverse biological properties, including anti-infl ammatory [136], analgesic [137], general antimicrobial [138], and specifi cally antiviral activities [139].Potential next generation lead compounds with a premise for SARS-CoV-2 have been summarized in Table 6.

8.1 Diterpenoids

Park et al.found the ethanol extract ofSalvia miltiorrhizato possess great inhibitory activity against both SARS-CoV 3CL pro and PL pro .The ethanol extract exhibited 60% and 88% inhibition of 3CL pro and PL pro , respectively, at 30 μg/mL).The plant speciesS.miltiorrhizais widely found in China, Korea, and Japan and has been greatly used to treat coronary heart disease, particularly angina pectoris and myocardial infarction [140].S.miltiorrhizais known to possess antioxidant, anti-infl ammatory and antiviral properties [39].Tashinones (Fig.9) isolated fromS.miltiorrhizainclude dihydrotanshinone I ( 30), tanshinone IIA ( 31), methyl tanshinonate ( 32), tanshinone I ( 33) and cryptotanshinone ( 34) [36].All the tanshinones are good inhibitors of the cysteine protease (3CL pro and PL pro ), for 3CL pro the inhibitory activity was assayed following the proteolysis of the fluorogenic substrate in the presence or absence of the test compounds, all compounds except compound 33 exhibited a dose dependent inhibitory eff ect on 3CL pro the activities ranged from 14.4 to 89.1 μM while PL pro inhibitory activity was assayed following continuous fluorometric, these compounds impressively show better activity against PL pro have a time dependent inhibitory profi le on PL pro with compound 33 having highest activity ( IC50value of 0.8 μM), all compounds have better activity against PL pro compared to 3CL pro [36].A detailed kinetic mechanism study showed that compounds 30,31,32,33 and 34 exhibited slow binding inhibitions with enzyme isomerisation and were shown to be non-competitive inhibitors.Moreover, compound 33 was reported to have a potent activity against cellular DUB with IC50= 0.7 μM [36].Recent studies have reported the use of compound 31 to treat myocardial infarction and delay of ventricular remodeling, in combination with puerarin.The compound is known to act by inhibiting the infl ammation in the early stage of myocardial infarction and plays an important role in inhibiting the ventricular remodeling in the later stage of myocardial infarction.The combination of compound 31 and puerarin can improve cardiac function, improve hemodynamics, reduce myocardial cells and reduce collagen synthesis in mice after myocardial infarction [141].

Fig.9 Chemical structures of tashinones, a promising sub-class of diterpenoids ( 30 to 34) with potential against SARS-CoV-2

8.2 Triterpenoids

Fig.10 Chemical structures of promising triterpenoids ( 35 to 41) with potential against SARS-CoV-2

Yang et al.isolated: 3β-hydroxy-28-norolean-12,1-dien-16-one 3-O-6′-methoxy-α- D-glucuronopyranoside ( 35), 3β-hydroxy-28-noroleana-12,17-dien-16-One ( 38), and 3β,16α-dihydroxy-olean-12-en-28-al 3-O-β- Dglucopyranoside ( 39), which are all oleanane triterpenes (Fig.10) from the flowers ofCamellia japonica.The compounds were tested for their inhibitory eff ectiveness against PEDV [135].By targeting the epithelial cells of the small intestine, the PEDV virus can cause severe mucosal atrophy and malabsorption resulting in acute and lethal diarrhea in piglets [127].The plant speciesC.japonicais found abundantly in Korea, Japan, and China [128].The ethanol extract (70%) ofC.japonicahas a potential inhibitory eff ect against PEDV replication [133].The anti-infl ammatory [33] and cytotoxic [116] properties of flowers ofC.japonicahave been exploited traditionally and used in treating hematemesis and internal and external bleeding injury [142].The compounds 35,38 and 39 had potent eff ects against the replication of PEDV.The known drug azauridine was used as a positive control.Additionally, the compounds gave higher selective indices (SI = 14.74, 32.72 and 6.68, respectively), with compound 38 being lower than azauridine (SI = 14.30).Moreover, compound 38 was shown to inhibit the virus replication in a time course study and it was further investigated detailly and found to inhibit PEDV RNA expression, encoding nucleocapsid, spike and membrane protein in a dose-dependent manner at concentrations of 2.0, 1.0, 0.5, and 0.25 μM, respectively [135].Therefore, the anti-PEDV molecules 35,38 and 39 could be investigated further as candidates against SARS-CoV-2.

Celastrol ( 36), tingenone ( 37) pristimerin ( 40) and iguesterin ( 41) are quinone methide triterpenoids isolated from the bark ofTripterygium regelii[143].T.regeliiis a vine found widely in China, Korea, Japan, and Taiwan [117].T.regeliiplant has a good cytotoxicity property against numerous cancer cell lines [118] and its effi cacy has been reported in rodent models of arthritis and other infl ammatory disease [119,120].Ryu et al.reported the SARS-CoV 3CL pro inhibitory properties of compounds from this plant species [143].All compounds displayed dose-dependent inhibitory activities, with the compounds 36, 37,40 and 41 having IC 50 values of 10.3, 9.9, 5.5 and 2.6 μM, respectively.Additionally, structure–activity relationship studies revealed that the quinone-methide moiety present in these compounds is important for SARS-CoV 3CL pro inhibition (Fig.11) [38].Molecular docking analysis of compounds from this class towards the 3CL pro protein with Protein Data Bank (PDB) code 1uk4 revealed that compound 41 could fi t well into the substrate-binding pocket of SARS-CoV 3CL pro , with the hydroxyl group of C3 of compound 41 forming a hydrogen bond with the oxygen atom of the carbonyl group of Cys44 and the OH of Thr25 located in domain I of the protein drug target (Fig.11) [38].

Fig.11 Molecular docking poses of the triterpenoids iguesterin ( A) and dihydrocelastrol ( B), isolated from the plant T.regelii showing binding to SARS-CoV 3CL pro (PDB code: 1uk4).This fi gure has been reproduced with copyright permission from Elsevier [38]

Fig.12 Blacoxanthone ( 42), a promising xanthone with potential against SARS-CoV-2

9 Xanthone

Xanthones are oxygen containing heterocyclic compounds commonly occurring in higher plants (especially those of the Guttiferae, Gentianaceae, Moraceae, Clusiaceae and Polygalaceae families), fungi, lichen, and bacteria [144,145], this class of secondary metabolites have been known to contain the following biological properties antimicrobial [146,147], anticancer [146,147], anti-infl ammatory [146,147], antimalaria [146,148] and antiviral [149].

Shen et al.studied the antiviral activity ofCalophyllum blancoiagainst SARS-CoV.The genusCalophyllumis a rich source of phenolics and xanthones, possess great antibacterial [143,150], antifungal [151], antiplatelet aggregation [150], immunomodulatory [152], anticancer [153] and anti-HIV-1 viral activities [154].Blancoxanthone ( 42 ), Table 7 and Fig.12, isolated from the rootsC.blancoi[42] showed the greatest in vitro inhibitory activity against HCoV-229E with an EC50value of 7.93 μM [42].The antiviral activity of the compound was evaluated using the Cell Proliferation Kit II (XTT) [42].

10 Discussion

Out of 42 NPs belonging to the alkaloid, flavonoid, terpenoid, phenolic, xanthone and saponin classes obtained from our literature survey.Our results showed that majority of the NPs with inhibitory concentrations against the virus or its target proteins below 10 μM were terpenoids, alkaloids, flavonoids, and diarylheptanoids, respectively.The terpenoids (diterpenoids and triterpenoids) were the most abundant among the isolated NPs, constituting 26.19% of the isolated compounds.This was followed by alkaloids (19.05%), flavonoids (11.90%), diarylheptanoids (9.52%), phenolics (4.76%), saponin (2.38%) and xanthones (2.38%), which showed a similar trend to previous studies [17– 19].

11 Conclusion

Natural products (NPs) have provided privileged scaff olds in drug design [16].The novel coronavirus disease-2019 (COVID-19) is caused by a positive-strand ribonucleic acid (RNA) virus, the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) [155].The virus has infected several million people and caused thousands of deaths worldwide since December 2019 [156].The pandemic is a signifi cant threat to public health and the global economy [155].To search for new bioactive compounds with anti-SARS-CoV-2 activity, 42 NPs with inhibitory concentrations against coronaviruses, their target proteins and other viruses, e.g., HIV.Infl uenza A virus, human simplex virus or their target proteins which lie below 10 μM were identifi ed from published data in literature.The SARS-CoV-2 virus was only discovered in 2019.References that show the testing of NPs against other SARS-CoV viruses (e.g.HCoV-NL63) and their targets (e.g.SARS-CoV PL pro ,3CL pro , SARS-CoV helicase, etc.) had earlier been published.We are presenting this review to encourage the testing of these compounds against the SARS-CoV-2 virus.In writing this manuscript, we must mention that the compounds were actually tested against the SARS-CoV-2 virus.Only ten of the 165 references date after 2020.It was the goal of the authors to summarize the results of compounds and plant species whose extracts have already been tested against earlier discovered coronaviruses so as to ease the task of discovery of NP antivirals that could be next generation drugs to treat infections caused by SARSCoV-2.The results of the current study reveal that promising NPs as potential inhibitors of SARS-CoV-2 were terpenoids, alkaloids, flavonoids, and diarylheptanoids, respectively.The mechanism of action of some of the NP hits and the plant species from which they were isolated has been included in the study.It is worth mentioning that some of the medicinal plants and the NPs have shown good safety in vitro studies.This review could serve as a starting point for further development of these NPs hits as potential leads for COVID-19.Thus, it would be interesting in future to evaluate the toxicities and binding mode of some of these NPs using in silico approaches.

Declarations

Conflict of interestThe authors declare no confl ict of interest.

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