Recent advances in the discovery of bioactive metabolitesfrom Pestalotiopsis

Sunil Kumar Deshmukh . Ved Prakash . Nihar Ranjan

Received: 19 November 2016 / Accepted: 14 February 2017

� Springer Science+Business Media Dordrecht 2017

Abstract Fungal endophytes have marked a signif-

icant impact on drug discovery reducing the burden

and dependency on plants. The vast diversity of

Pestalotiopsis sp. has emerged as promising source of

wide range of bioactive natural compounds. Recently

a series of numerous novel secondary metabolites

have been discovered of which taxol has drawn

attention of scientific community towards its medic-

inal potential. A wide variety of compounds like

alkaloids, polyketides, terpenoids, flavonoids, cou-

marins, xanthones, quinones, semiquinones, peptides,

phenols, phenolic acids, and lactones have been

identified which have usage as antimicrobial, antifun-

gal, antiviral antoneoplastic, and antioxidant activi-

ties. This review aims to highlight recent discoveries

of different strains of Pestalotiopsis identified for

producing natural bioactive compounds along with

insights of their source of origin and potential in

biotechnological applications.

Keywords Pestalotiopsis � Endophytes � Drugdiscovery � Natural products � Anticancer agents �Taxol

Introduction

Endophytic fungi have been area of wide research for

their unexplored potential in the discovery of bioactive

compounds. The genus Pestalotiopsis was established

by Steyaert in 1949, following a taxonomic amend-

ment to the genus Pestalotia (Steyaert 1949, 1953a, b).

To date, 234 described species of Pestalotiopsis that

are differentiated on conidial characteristics are listed

in Index Fungorum (http://www.indexfungorum.org/

Names/Names.asp). Pestalotiopsis species (Amphis-

phaeriaceae) are widely distributed in nature of which

many are saprobes, while others are either pathogenic

or endophytic to living plants (Jeewon et al. 2003).

Pestalotiopsis is a widespread genus, usually endo-

phytic, occurring in a wide range of substrata which

have attracted attention for their ability to produce a

variety of bioactive secondary metabolites. Chemical

studies of the fungal genus Pestalotiopsis have affor-

ded a variety of bioactive natural products (Yang et al.

2012). To underline its potential, over seventy new

bioactive secondary metabolites have been isolated

from Pestalotiopsis fici which is now one of the pro-

lific producers of novel natural products (Liu 2011). In

recent years, Pestalotiopsis, has gained considerable

attention (Xu et al. 2010). Since the discovery of taxol,

S. K. Deshmukh (&) � N. Ranjan (&)

TERI–Deakin Nano Biotechnology Centre, The Energy

and Resources Institute, Darbari Seth Block, IHC

Complex, Lodhi Road, New Delhi 110003, India

e-mail: sunil.deshmukh@teri.res.in

N. Ranjan

e-mail: nihar.ranjan@teri.res.in

V. Prakash

Department of Biotechnology, College of Engineering

and Technology, IILM-Academy of Higher Learning,

Greater Noida 201306, India

123

Phytochem Rev

DOI 10.1007/s11101-017-9495-3

an anticancer agent, from an endophytic fungal strain

Pestalotiopsis microspora (Strobel et al. 1996), sig-

nificant interest has been generated in the search of

bioactive compounds from this genus. This review

aims to highlight the different strains of Pestalotiopsis

identified for their various bioactive roles. Special

emphasis has been put to detail their source of origin,

functions and their biological roles.

Taxol producing Pestalotiopsis species

Paclitaxel (taxol) (1) (Fig. 1), a well-known and

highly functionalized tetracyclic diterpenoid bioactive

compound, was isolated from the bark of Taxus

brevifolia (Wani et al. 1971). Taxol is found in

extremely low amounts in the needles, bark, and roots

of yews (Taxus sp.). It is specifically targeted to treat

prostate, ovarian, breast, and lung cancers (Rowinsky

1997). Taxol stabilizes plus end dynamic instability of

microtubules both in vitro and in vivo (Jordan et al.

1993; Yvon 1999). The discovery of a paclitaxel-

producing endophytic fungus Taxomyces andreanae

from the Pacific yew (T. brevifolia) by Stierle et al.

(1993) generated immense interest in the scientific

community for the isolation of endophytic fungi,

which produce several other active metabolites.

Paclitaxel and its analogues such as baccatin III (2)

and 10-deacetylbaccatin III (3) (Fig. 1) have been

reported from a large number of endophytic fungi

(Zhao et al. 2011). Table 1 provides a comprehensive

list of taxol-producing species from the genus

Pestalotiopsis.As shown in Table 1, taxol is produced

by a large number of species of the same genus

(Pestalotiopsis) suggesting similarity of pathways in

their production.

Bioactive metabolites from Pestalotiopsis fici

Endophytic fungus Pestalotiopsis fici from the

branches of an unidentified tree in the suburb of

Hangzhou (China) was found to be highly favorable

for producing novel natural products. Liu (2011a) has

reviewed the work on compounds reported from this

particular fungus. Seventy new bioactive secondary

metabolites have been reported by in-depth chemical

studies. Some representative metabolites identified

from the Pestalotiopsis fici are reported here.

Pestalofones A, B, C and E (4–7) (Fig. 1) were

isolated from this fungus. The chemical structure

determination was done using standard one and two-

dimensional spectroscopic techniques. Compounds (4,

5 and 7) showed inhibitory effects on HIV-1 replica-

tion in C8166 cells, with EC50 values of 90.4, 64.0, and

93.7 lM, respectively (all three compounds showed

CC50 values of greater than 200 lM; the positive

control indinavir sulfate showed an EC50 value of

8.81 nM). Pestalofone C (6) and E (7) also showed

significant antifungal activity against Aspergillus

fumigatus, with IC50/MIC values of 1.10/35.3, 0.90/

31.2 lM, respectively (the positive control flucona-

zole showed IC50/MIC values of 7.35/163.4 lM) (Liu

et al. 2009a).

Liu et al. (2011a) isolated Pestalofone F (8) and

Pestalodiol C (9) (Fig. 1) from P. fici isolated from the

branches of C. sinensis. Compound (8) displayed

cytotoxicity against HeLa and MCF-7 cells with IC50

values of 14.4 and 11.9 lM, respectively and Pestalo-

diol C displayed cytotoxicity against the HeLa and

MCF-7 cells, with IC50 values of 16.7 and 57.5 lM,

respectively. Pestalofone J (10), and K (11) (Fig. 1)

were obtained from P. fici isolated from the branches

of C. sinensis. Using a combination of 1D, 2D NMR

(1H-1H COSY, HMBC, and NOESY) and mass

spectroscopic techniques, the chemical structure was

determined. Compound (10) showed weak cytotoxic

activities against HeLa, T24, A549, and MCF-7 cell

lines with IC50 values of 44.3, 39.3, 35.3, and

38.3 lM, respectively. Compound (11) was also found

to have weak cytotoxic activities against HeLa, T24,

A549, and MCF-7 cell lines, with IC50 values 65.5,

45.7, 58.9, and 29.2 lM, respectively (the positive

control cisplatin showed the IC50 values of 7.4, 3.9,

8.4, and 6.4 lM, respectively) (Wang et al. 2016).

Chloropupukeananin (12) (Fig. 1) was another

bioactive compound isolated from the same fungus.

Proton COSY analysis of this compound revealed the

presence of two different sets of proton spin systems

and further analysis using HMBC indicated the

presence of methyl benzoate unit in this isolate. Using

HMBC correlation and single crystal X-ray diffrac-

tion, the structure of compound 12 was unambigu-

ously assigned with defined stereochemistry at all the

chiral centers. Compound (12) showed an inhibitory

effect on HIV-1 replication in C8166 cells, with an

IC50 value of 14.6 lM, and it also displayed cytotoxic

effect against HeLa and HT29 cells, showing IC50

Phytochem Rev

123

Fig. 1 Chemical structures of Taxol and its analogs (1–3) and metabolites isolated from the genus Pestalotiopsis fici

Phytochem Rev

123

values of 1.4 and 6.7 lM, respectively. In addition,

modest antimicrobial activity was also observed for

this metabolite against the Gram-positive bacterium,

Staphylococcus aureus (ATCC 6538) with IC50 and

MIC values of 21.8 and 97.3 lM, respectively (the

positive control ampicillin showed IC50 and MIC

values of 1.2 and 3.9 lM) (Liu et al. 2008a).

In other studies, Chloropestolide A (13) (Fig. 1)

was isolated from the same fungus. Chloropestolide A

(13) showed significant inhibitory effects on the

growth of HeLa and HT29 cancer cell lines with

GI50 values of 0.7 and 4.2 lM, respectively (Liu et al.

2009b). Chloropestolide B (14) (Fig. 1) was cytotoxic

to three tested cell lines CNE1-LMP1, A375 and

MCF-7 showing IC50 values of 16.4, 9.9, and

23.6 lM, respectively while the positive control

paclitaxel showed IC50 values of 4.2, 8.9, and

0.14 nM, respectively (Liu et al. 2013a).

Chloropupukeanolide A (15) (Fig. 1) was another

metabolite reported from P. fici. The structure was

determined using homo and heteronuclear NMR

techniques which established the presence of

fragments of a tricyclo[4.3.1.03,7]decane, an aniso-

prenylated 2,3-epoxycyclohex-5-en-1,4-diol (ECH),

and a 2,6-dihydroxy-4-methylbenzoate (DMB) unit.

Compound (15) showed an inhibitory effect on HIV-1

replication in C8166 cells, with an EC50 value of 6.9

lM (the positive control indinavir sulfate showed an

EC50 value of 8.81 nM) and also showed significant

cytotoxicity against HeLa, MCF-7 and MDA-MB-231

cell lines with IC50 values of 16.9, 15.5 and 15.9 lM,

respectively (Liu et al. 2010).

Liu and coworkers have used large scale P. fici re-

fermentation to isolate minor fragments observed in

their previous work. Extending their work to isolate

Chloropupukeananin (12) (Fig. 1), large-scale re-

fermenatation at 1 kg and subsequently at 3 kg on

rice led to the isolation of Chloropupukeanolides C–D

(16, 17) (Fig. 1), a novel spiroketal skeleton. The

solution structure through-bond and spatial connec-

tivities deduced using NMR were supported by the

single crystal X-Ray diffraction data which aided in

unambiguous stereochemistry determinations at the

chiral centers. Chloropupukeanolide C (16) and D (17)

Fig. 1 continued

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123

Table 1 Taxol-producing Pestalotiopsis species

Sr. No. Fungus Plant source Reference

1 Pestalotiopsis microspora Ja-73 Taxus cuspidata Strobel et al. (1996)

2 Pestalotiopsis guepinii W-1f-2 Wollemia nobilis Strobel et al. (1996)

3 P. microspora Ne 32 Taxus wallachiana Strobel et al. (1996)

4 P. microspora No. 1040 Taxus wallachiana Strobel et al. (1996)

5 P. microspora Cp-4 Taxodium distichum Li et al. (1996)

6 Pestalotiopsis sp.W-x-3 Wollemia nobilis Strobel et al. (1996)

7 Pestalotiopsis sp.W-1f-1 Wollemia nobilis Strobel et al. (1996)

8 P. guepinii Wollemia nobilis Strobel et al. (1997)

9 Pestalotiopsis menezesiana and

Pestalotiopsis uvicola

Not reported Muthumary and Sashirekha (2007)

10 Pestalotiopsis neglecta Not reported Gangadevi and Muthumary (2008)

11 P. terminaliae Not reported Gangadevi and Muthumary (2008)

12 Pestalotiopsis pauciseta (strain CHP-11) Cardiospermum helicacabum Gangadevi and Muthumary (2008)

13 P. Paucisrta Not reported Gangadevi et al. (2008)

14 Pestalotiopsis mangiferae Not reported Kathiravan and Sureban (2009)

15 Pestalotiopsis sp. Catharanthus roseus Srinivasan and Muthumary (2009)

16 Pestalotiopsis terminaliae, Terminalia arjuna Gangadevi and Muthumary (2009)

17 P. microspora T. cuspidata, Kumaran et al. (2010)

18 P. neglecta T. cuspidata Kumaran et al. (2010)

19 Pestalotiopsis breviseta Infected leaf of Ervatamia divaricata Kathiravan and Sri Raman (2010)

20 P. pauciseta Tabebuia pentaphylla Vennila and Muthumary (2010)

21 Pestalotiopsis versicolor T. cuspidata Kumaran et al. (2010)

22 Pestalotiopsis malicola NK101 Plant debris in the soil. Bi et al. (2011)

23 P. breviseta CR01 Catharanceus roceus Karthik et al. (2012)

24 Pestalotiopsis acaciae Not reported Kathiravan et al. (2014)

25 Pestalotiopsis adusta Not reported Kathiravan et al. (2014)

28 P. breviseta Not reported Kathiravan et al. (2014)

29 Pestalotiopsis calabae Not reported Kathiravan et al. (2014)

30 Pestalotiopsis coangae Not reported Kathiravan et al. (2014)

31 Pestalotiopsis coffeae Not reported Kathiravan et al. (2014)

32 Pestalotiopsis conigena Not reported Kathiravan et al. (2014)

32 Pestalotiopsiseriobotryofolia Not reported Kathiravan et al. (2014)

33 Pestalotiopsis foedaris Not reported Kathiravan et al. (2014)

34 Pestalotiopsis fibricola Not reported Kathiravan et al. (2014)

35 Pestalotiopsis glandicola Not reported Kathiravan et al. (2014)

36 Pestalotiopsis japonica Not reported Kathiravan et al. (2014)

37 Pestalotiopsis matildae Not reported Kathiravan et al. (2014)

38 Pestalotiopsis oleandri Not reported Kathiravan et al. (2014)

39 Pestalotiopsis paeoniae Not reported Kathiravan et al. (2014)

40 Pestalotiopsis paciseta Not reported Kathiravan et al. (2014)

41 Pestalotiopsis taxicab Not reported Kathiravan et al. (2014)

42 Pestalotiopsis torulosa Not reported Kathiravan et al. (2014)

43 Pestalotiopsis zalbrukneriana Not reported Kathiravan et al. (2014)

44 Pestalotiopsis hainanensis Dermatitic scurf of the giant panda

(Ailuropoda melanoleuca)

Gu et al. (2015)

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123

showed significant cytotoxicity against HeLa and

HT29 cell lines, with IC50 values ranging from 1.2 to

7.9 lM which was lower than the positive control

5-fluorouracil, that gave IC50 values of 10.0 and

15.0 lM (Liu et al. 2011b).

Siccayne (18) (Fig. 1) was a known metabolite

isolated from endophytic fungus P. fici which showed

cytotoxic activity against the human cancer cell lines

HeLa and HT29 with IC50 values 48.2 and 33.9 lM,

respectively (5-fluorouracil, the positive control, gave

IC50 values of 8.0 and 12.0 lM, respectively) (Liu

et al. 2013b). Using further large-scale fermentation,

Liu and coworkers isolated Pestaloficiol J (19) and L

(20) (Fig. 1), two new isoprenylated chromone

derivatives with compound (20) being a heterodimer,

from endophytic fungus P. fici using two-dimensional

NMR techniques. Compound (19) showed an inhibi-

tory effect on HIV-1 replication in C8166 cells, with

an EC50 value of 8.0 lM (the CC50 value was greater

than 100 lM; the positive control indinavir sulfate

showed an EC50 value of 8.2 nM). Compound (20)

displayed cytotoxic activity against the HeLa and

MCF-7 cells, with IC50 values of 8.7 and 17.4 lM,

respectively (the positive control 5-fluorouracil

showed IC50 values of 10.0 and 15.0 lM, respec-

tively) (Liu et al. 2009c). Pestaloficiols A, B, D (21,

22, 23) (Fig. 1), a new cyclopropane derivative,

showed inhibitory effects on HIV-1 replication in

C8166 cells, with EC50 values 26.0, 98.1, and 64.1

lM, respectively (all three compounds showed CC50

values of greater than 200 lM; the positive control

indinavir sulfate showed an EC50 value of 8.81 nM)

(Liu et al. 2008b). Pestaloficiols N-P (24, 25, 26)

(Fig. 1), displayed inhibitory effects on HIV-1 repli-

cation in C8166 cells, whereas compounds (25) and

(26) showed cytotoxic activity against the human

tumor cell line HeLa. Compound (25) also showed

antifungal activity against A. fumigatus (Liu and Liu

2010).

A new a-pyrone derivative ficipyrone A (27)

(Fig. 1) was isolated from P. fici. 1D and 2D NMR

data of ficipyrone A (27) revealed the presence of a

single ring and homonuclear COSY showed the

presence of one isolated spin system. Further HMBC

correlations indicated the presence of an a-pyronemoiety. The absolute configuration of the chiral center

at C-7 was determined using circular dichroism

spectroscopy which displyed negative cotton effect

leading to ‘7S’ assignment. Compound (27) showed

antifungal activity against the plant pathogen Gib-

berella zeae (CGMCC 3.2873) with an IC50 value of

15.9 lM (the positive control ketoconazole showed an

IC50 value of 6.02 lM) (Liu et al. 2013c). Using a

similar approach as in ficipyrone A (27) which utilized

one and two dimensional NMR as well as circular

dichroism techniques to determine the absolute struc-

ture, the chemical structure of Pestalotriols B (28) was

identified. Pestalotriol B (28) (Fig. 1) displayed weak

cytotoxicity against the HeLa cells showing an IC50

value of 87.0 lM (the positive control cisplatin

showed an IC50 value of 7.4 lM) (Liu et al. 2015).

Detailed genomic analysis has been done to trace

the biosynthetic origin of these metabolites. In the case

of diphenyl ether pestheic acid, the precursor of the

unique chloropupukeananes, biosynthesis proceeded

through the formation of the polyketide backbone,

cyclization of a polyketo acid to a benzophenone,

chlorination, and formation of the dipehnyl ether

skeleton through oxidation and hydrolyzation (Xu

et al. 2014). Wang et al. (2015a) reported a rich set of

secondary metabolite synthesis genes, including

twenty-seven polyketide synthases (PKSs), twelve

non-ribosomal peptide synthases (NRPSs), five

dimethylallyl tryptophan synthases, four putative

PKS-like enzymes, fifteen putative NRPS-like

enzymes, fifteen terpenoid synthases, seven terpenoid

cyclases, seven fatty-acid synthases, and five hybrids

of PKS-NRPS.Wang et al. (2015b) also uncovered the

role of oxidative stress in the biosynthesis of metabo-

lites. These studies reveal that biosynthesis in P. fici

uses a different set of genes for constructing different

class of metabolites.

Bioactive metabolites from Pestalotiopsis

microspora

Strobel and Daisy (2003) commented that endophytes

could be a goldmine of secondary metabolites.

Pestalotiopsis spp. can be considered as ‘‘the E. coli

of the rain forests’’ and P. microspora a ‘‘microbial

factory’’ of bioactive secondary metabolites. Accord-

ing to them a variety of chemical structures are seen

such as taxol, torreyanic acid, ambuic acid, crypto-

candin, subglutinol A and B etc.

The phytotoxins pestalopyrone (29), hydrox-

ypestalopyrone (30) and pestaloside (31) (Fig. 2)

were produced by P. microspora which was isolated

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123

from artificially infected Florida torreya (Torreya

taxifolia), a rare tree in North America. The structural

assignments were done using mass spectra and NMR

techniques. The fungus resides in the inner bark of

symptomless trees where physiological or environ-

mental factors could trigger its pathological activity

which could give rise to diseases. Pestaloside (31) has

antifungal activity and could reduce competition from

other fungal endophytes within the host. Pestaloside

caused distinct zones of inhibition against a Cla-

dosporium sp. and a sterile hyphomycete isolated from

T. taxifolia. Pestaloside (31) was also active against

Rhizoctonia solani, Geotrichum candidurn and Agar-

icus compestris, but not against Trichoderma species

(Lee et al. 1995).

Ambuic acid (32) (Fig. 2), a highly functionalized

cyclohexenone, was isolated and characterized from P.

microspora andMonochaetia sp using two-dimensional

NMR techniques such as COSY, HBMC, INADE-

QUATE, DEPT and NOESY as well as mass spectral

techniques. These are biologically related endophytic

fungi associated with many tropical plant species. In

plate well dilution assay, ambuic acidwas active against

Pythium ultimum with MIC of 7.5 lg/mL (Li et al.

2001).LL-P880c (33) (Fig. 2), a knowncompound,was

isolated fromendophytic fungusP.microspora from the

branch of Taxus chinensis in Yichang, Hubei Province,

China. Compound 33, as pestalotin analog, showed

significant gibberellin synergistic activity towards

Distylium chinense seeds with the substrate LL-P880cconcentration of 0.6 mg/L. The germination rate of the

seeds was 85.5% (Li et al. 2015a).

Pestalotiopsis microspora, native to the rainforest

of Papua New Guinea, produces pestacin (34) (Fig. 2)

which exhibits antioxidant activity eleven times

greater than vitamin E derivative trolox. Antioxidant

Fig. 2 Chemical structures of metabolites isolated from the genus Pestalotiopsis microspora

Phytochem Rev

123

activity is proposed to arise primarily via cleavage of

an unusually reactive C–H bond and to a lesser extent

through O–H abstraction. It has moderate antifungal

property with minimum inhibitory concentration

(MIC) of approximately 10 lg/mL against P. ultimum

an important root-invading pathogen (Harper et al.

2003).

Isopestacin (35) (Fig. 2) is an isobenzofuranone,

obtained from the endophytic fungus P. microspora

associated with Terminalia morobensis located in the

Sepik river drainage of Papua New Guinea. Using a

combination of 2D INADEQUATE, HMQC and

X-ray crystallography data, the structure was assigned

which contained the isobenzofuranone framework.

Isopestacin (35) is moderately antimycotic, with total

inhibition of P. ultimum, a plant pathogenic oomycete,

at 40 lg/mL at 48 h. It also behaves as an antioxidant

scavenger to both superoxide and hydroxyl free

radicals (Strobel et al. 2002).

Torreyanic acid (36) (Fig. 2) was isolated from P.

microspora associated with Florida torreya (Torreya

taxifolia), an endangered species, closely related to the

taxol-producing Pacific yew (Taxus brevifolia). The

complete structural characterization was achieved by

using two-dimensional NMR and single crystal X-ray

analysis which showed the presence of a seven ring

system that could be synthesized by Diels–Alder

cyclization of two identical units. Torreyanic acid (36)

is fifteen times more potent in cell lines that are

sensitive to protein kinase C (PKC) agonists and

causes cell death by apoptosis. IC50 values of

compound (36) range from 3.5 (NEC) to 45 (A549)

lg/mL with a mean value of 9.4 lg/mL for 25

different cell lines. Torreyanic acid also shows G1

arrest of G0 synchronized cells at 1–5 lg/mL level

depending on the cell line used (Lee et al. 1996).

Bioactive metabolites from Pestalotiopsis foedan

(-)-(4S,8S)-Foedanolide (37) and (?)-(4R,8R)-

Foedanolide (38) (Fig. 3), a pair of new spiro-c-lactone enantiomers, were obtained from Pestalotiop-

sis foedan from the branch of Bruguiera sexangula. A

combined NMR, circular dichroism, mass spec-

troscopy and computation analysis was used to assign

the absolute configuration. Both compounds exhibited

inhibitory activities against HeLa, HepG2 and MCF-7

where the cytotoxic activity of compound (38) was

higher than compound (37). Compound (37) exhibited

cytotoxic activity against HeLa, HepG2 and MCF-7

cell lines with IC50 values 15.8, 22.8 and 70.2 lg/mL,

respectively while the corresponding IC50 values of

cytotoxic activity of (38) were 5.4, 19.0 and 20.8 lg/mL, respectively (Yang and Li 2013).

Fig. 3 Chemical structures of metabolites isolated from the genus Pestalotiopsis foedan

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123

Monoterpene lactones, (3R,4R,6R,7S)-7-hydroxyl-

3,7-dimethyl-oxabicyclo[3.3.1]nonan-2-one (39), and

(3R, 4R)-3-(7-methylcyclohexenyl)-propanoic acid

(40) (Fig. 3) were obtained from P. foedan from the

branch of Bruguiera sexangula. ESI–MS, NMR and

computation analysis were used to determine their

complete structures. Both compounds exhibited strong

antifungal activities against Botrytis cinerea and

Phytophthora nicotianae with MIC values of 3.1 and

6.3 lg/mL, respectively which are comparable to that

of the known antifungal drug ketoconazole. Com-

pound (40) also showed modest antifungal activity

against Candida albicans with a MIC value of 50 lg/mL (Xu et al. 2016a).

Pestafolide A (41), a new reduced spiro aza-

philone derivative and pestaphthalides A (42) and B

(43) (Fig. 3), two new isobenzofuranones, were

isolated from solid cultures of an isolate of P. foedan

using bio-assay guided fractionation of the organic

extract. Pestafolide A (41), displayed antifungal

activity against A. fumigatus (ATCC 10894) afford-

ing a zone of inhibition of 10 mm at 100 lg/disk.Pestaphthalide A (42) showed activity against Can-

dida albicans (ATCC 10231) causing a zone of

inhibition of 13 mm, and pestaphthalide B (43)

showed activity against G. candidum (AS2.498) with

11 mm zone of inhibition when tested at the same

level (fluconazole: 18–28 mm zones of inhibition for

C. albicans, A. fumigatus, and G. candidum at

100 lg/disk) (Ding et al. 2008a).

Bioactive metabolites from Pestalotiopsis adusta

Bioassay-guided fractionation of culture extract of P.

adusta, an endophytic fungus isolated from the

medicinal plant Clerodendrum canescens led to the

isolation of (10S)-12,16-epoxy-17(15 ? 16)-abeo-

3,5,8,12,15-abietapentaen-2,7,11,14-tetraone (44),

teuvincenone F (45), uncinatone (46), coleon U (47)

and coleon U-12-Me ether (48) (Fig. 4). The structural

characterization of novel metoabolite (44) discovered

in this study was done using a combination of

spectroscopic techniques which revealed the presence

of a benzoquinone core. Compounds (44–48) dis-

played cytotoxicity agaisnst HL-60 tumor cell line

with IC50 values of 12.54, 25.06, 15.66, 57.60, and

66.41 lM, respectively in comparision to cisplatin

with IC50 of 9.20 lM (Xu et al. 2016b).

The organic soluble extract of plant endophytic

fungus P. adusta (L416), obtained from the stem of an

unidentified tree in Hainan Province (China), was

subjected to bio-assay guided fractionation. Structural

analysis using a combination of NMR and X-ray

Fig. 4 Chemical structures of metabolites isolated from the genus Pestalotiopsis adusta

Phytochem Rev

123

crystallography techniques resulted in the identifica-

tion of three new metabolites Pestalachlorides A–C

(49–51) (Fig. 4) containing chlorinated benzophe-

nones. Pestalachloride A was obtained as a mixture of

two atropisomers in a 5:4 ratio. Exhaustive efforts to

isolate this mixture using column chromatography and

HPLC were unfruitful leading to difficulties in their

structural characterization. The final structure was

assigned on the basis of X-ray crystal structure which

was found to contain two enantiomers (R, S) of

compound (49). Pestalachloride A (49) displayed

potent antifungal activity against Fusarium culmorum

with an IC50 value of 0.89 lMwhile pestalachloride B

(50) exhibited remarkable activity against Gibberella

zeae with an IC50 value of 1.1 lM. Pestalachloride C

(51) did not show noticeable in vitro antifungal

activities against plant pathogens F. culmorum, G.

zeae, and Verticillium aibo-atrum (IC50[ 100 lM)

(Li et al. 2008a).

Bioactive metabolites from Pestalotiopsis photiniae

Photinides A–F (52–57) (Fig. 5), six unique benzofu-

ranone-derived c-lactones were isolated from the

crude extract of P. photiniae endophtes of Roystonea

regia (H.B.K.) Cook (Arecoideae) collected from

JianfengMountain, Hainan Province, People’s Repub-

lic of China. Their structure elucidation was done

using NMR and mass spectroscopy which revealed the

presence of three cyclic ring systems of which the

aromatic unit was tri-substituted. NOESY and circular

dichroism were used to determine absolute configura-

tions of the stereogenic centers. Photinides A–F

(52–57) showed modest but selective cytotoxicities

against MDA-MB-2311 with inhibitory rates of 24.4,

24.2, 23.1, 24.4, and 24.6%, respectively when tested

at 10 lg/mL (Ding et al. 2009a). Photipyrone B (58)

(Fig. 5), was isolated from the same culture ‘using one

strain many compounds’ (OSMAC) approach.

Photipyrone B (58) showed modest inhibitory effect

on the growth of MDA-MB-231 with the inhibitory

rate at 25.0 and 23.0% respectively when tested at

10 lg/mL (Ding et al. 2012).

DMMP [4-(30,30-dimethylallyloxy)-5-methyl-6-

methoxyphthalide] (59) (Fig. 5) was obtained from

P. photiniae from the Chinese podocarpaceae plant

Podocarpus macrophyllus. DMMP (59) significantly

inhibited the proliferation of HeLa tumor cell lines.

After treatment with DMMP (59), characteristic

apoptotic features such as DNA fragmentation and

chromatin condensation were observed in DAPI-

stained HeLa cells. Flow cytometry showed that

DMMP (59) induced G1 cell cycle arrest and apop-

tosis in a dose-dependent manner. Western blotting

and real-time reverse transcription-polymerase chain

reaction were used to investigate protein and mRNA

expression. DMMP (59) caused significant cell cycle

arrest by upregulating the cyclin-dependent kinase

inhibitor p27KIP1 protein and p21CIP1 mRNA levels

in HeLa cells. The expression of p73 protein was

increased after treatment with various DMMP (59)

concentrations. mRNA expression of the cell cycle-

related genes, p21CIP1, p16INK4a and Gadd45a, wassignificantly upregulated and mRNA levels demon-

strated significantly increased translation of p73,

JunB, FKHR, and Bim (Chen and Yang, 2013).

DMMP (59) also exhibited cytotoxicity against HeLa,

MDA-MB-231, MCF7 and MRC5 cell lines with

IC50 value of 36, 51, 81 and 147 lg/mL, respectively.

Moreover, DMMP was able to induce marked nuclear

apoptotic morphology in HeLa cells. DMMP (59)

induced apoptosis and loss of mitochondrial mem-

brane potential (DWm) in the HeLa cells. Although,

the activated forms of caspase-9 and caspase-3 in

HeLa cells were detected, pretreatment with caspase

inhibitors (Ac-DEVD-CHO and Z-VAD-FMK) failed

to attenuate MP-induced cell death (Chen et al. 2013).

Refermentation of P. photiniae isolated from the

Chinese Podocarpaceae plant P. macrophyllus and

further bio-assay guided fractionation of organic

extract yielded three new phthalide derivatives named

5-(30-methyl-20-butenyl)-2-hydroxy-3-methoxy-4-

methylbenzoic acid (60), 5-(30-carboxyl-30-methyl-

2E-allyloxy)-3-methoxy-4-methylphthalide (61) and

5-(30,30-dimethylallyloxy)-2-methoxycarbonyl-3-

methoxy-4-methylbenzoic acid (62) together with six

other known phthalide derivatives named 5-(30,30-dimethylallyloxy)-3-methoxy-4-methylphthalide

(63), zinnimidine (64), 5-(30,30-dimethylallyloxy)-3-

methoxy-4-methylphthalide (65), 5-(30,30-dimethylal-

lyloxy)-3-methoxy4-methylphthalic acid (66), zinniol

anhydride (67) and porriolide (68) (Fig. 5). Together

with mass spectroscopy data, two-dimensional NMR

spectroscopy employing HMQC and HMBC techqni-

ues were used to establish through-bond connectivities

Phytochem Rev

123

which led to the structural identification of the

compounds. Compounds (60–68) displayed signifi-

cant antifungal activities against three fungal strains

including Fusarium graminearum, B. cinerea and P.

nicotianae, with MIC values from 50.0 to 3.1 lg/mL

(the positive control ketoconazole showedMIC values

are 3.1 lg/mL) (Yang et al. 2011a).

Bioactive metabolites from Pestalotiopsis theae

Chloroisosulochrin (69), ficipyrone A (70) and pes-

theic acid (71) (Fig. 6) were obtained from endophytic

fungus Pestalotiopsis theae isolated from the fresh

leaves of the host Nigerian plant Fagara zanthoxy-

loides. Compound (69) was found to have the

Fig. 5 Chemical structures of metabolites isolated from the genus Pestalotiopsis photiniae

Phytochem Rev

123

strongest inhibition of the respiratory syncytial virus

(RSV) with IC50 of 4.22 ± 1.03 lM (ribavirin

4.91 ± 1.85 lM), while compound (70) and (71)

showed moderate inhibition of RSV with IC50 of

45.00 ± 0.98 and 146.20 ± 2.14 lM, respectively

(Uzor et al. 2016).

Phytotoxin (72) (?)-epiepoxydon (73), and oxys-

porone (74) (Fig. 6) were isolated from tea gray blight

fungi, P. longiesta and from P. theae. The threshold

concentrations of induced leaf necrosis for a cultivar

Yabukita by compounds (72–74) were 4, 60, and

15 lg/mL, respectively (Nagata et al. 1992). Solid-

substrate fermentation of plant pathogenic fungus P.

theae and its subsequent fractionation led to the

isolation of three new amides, pestalazine A (75),

pestalamide A (76) and asperazine (77) (Fig. 6).

Compounds (75–77) showed inhibitory effects on

HIV-1 replication in C8166 cells with EC50 values of

47.6, 64.2, and 98.9 lM, respectively (the CC50 values

for these compounds are all greater than 100 lM; the

positive control indinavir sulfate showed an EC50

value of 5.5 nM). Pestalamide A (76) displayed potent

antifungal activity against A. fumigatus (ATCC

10894) with IC50/MIC values of 1.50/57.8 lM (the

positive control fluconazole showed IC50/MIC values

of 7.35/163.4 lM) (Ding et al. 2008b). Pestalotheol C

(78) (Fig. 6) was obtained from cultures of an isolate

of P. theae. The structural analysis using NMR

showed the presence of four isolated proton spin

systems. The relative configuration of Pestalotheol C

(78) was assigned using homonuclear COSY and

NOESY correlations. The absolute configuration was

determined using modified Mosher method which led

to 2R, 4R, 5S, 6R, 9S spatial arrangement. Compound

(78) displayed an inhibitory effect on HIV-1LAIreplication in C8166 cells with an EC50 value of

16.1 lM (Li et al. 2008b).

Bioactive metabolites from other Pestalotiopsis

species

Compounds (79) and (80) (Fig. 7) were isolated from

the Pestalotiopsis virgatula, an endophytic fungus in

Fig. 6 Chemical structures of metabolites isolated from the genus Pestalotiopsis theae

Phytochem Rev

123

Fig. 7 Chemical structures of metabolites isolated from miscellaneous Pestalotiopsis species

Phytochem Rev

123

the branch of the Kandelia obovata. Compounds (79)

and (80), showed moderate selective inhibitory activ-

ities against five tumor cell lines using MTT method

(Li and Yang 2014). Penicillide derivatives (81–83)

(Fig. 7) and a-pyrone analogs (84–87) (Fig. 7) were

isolated from the solid culture on cooked rice of

Pestalotiopsis sydowiana from a halophyte, Phrag-

mites communis Trinus. The inhibitory activities

against the 20S proteasome were evaluated. Com-

pounds (81–87) inhibited the activity of the 20S

proteasome in a dose-dependent manner with IC50

values ranging from 1.2 ± 0.3 lM to 30.5 ± 1.5 lM(Xia et al. 2016). (±)-pestalachlorides E (88) and F

(89) (Fig. 7) were isolated from a marine-derived

Pestalotiopsis (ZJ-2009-7-6) fungus and their struc-

ture was determined using 1H-1H COSY and HMBC

correlations. Both the compounds showed potent anti-

fouling activities against the larval settlement of the

barnacle Balanus amphitrite at non-toxic concentra-

tions with EC50 values of 1.65 and 0.55 lg/mL,

respectively and anti-fouling activity was detected for

the first time for this class of metabolites (Xing et al.

2016).

Ambuic acid derivatives (90–92) (Fig. 7) were

isolated from the organic layer extract of the solid

culture of a plant pathogenic fungus Pestalotiopsis

neglecta. Routine 1D and 2D NMR experiments

together with mass spectroscopy data assisted in the

skeletal framework determination. NOESY and cir-

cular dichroism experiments were done to obtain the

relative and absolute configuration which led to total

assignment of the molecule. In the nitric oxide (NO)

inhibition assay, compounds (90–92) showed inhibi-

tory activity against the NO production in the

lipopolysaccharide (LPS)-induced macrophage with

IC50 values of 88.66, 11.20, and 20.80 lM,

Fig. 7 continued

Phytochem Rev

123

respectively (Qi et al. 2015). A polyhydroxylated

macrolide, namedmangiferaelactone (93) (Fig. 7) was

obtained from Pestalotiopsis manguiferae, isolated

from Hyptis dilatata, from the central region of

Panama. NMR based homo and heteronuclear con-

nectivities led to the determination of chemical

structure whose absolute configuration was deter-

mined using vibrational circular dichroism spec-

troscopy. Compound (95) showed MIC of 1.68 lg/mL against Listeria monocytogenes, and 0.55 lg/mL

against Bacillus cereus (Ortega et al. 2014).

Jesterone (94) and hydroxy-jesterone (95) (Fig. 7)

are novel highly functionalized cyclohexenone epox-

ides isolated from endophytic fungal species Pestalo-

tiopsis jesteri. Using DEPT and HSQC correlations,

several through-bond connectivities were established

for structure determination. Jesterone (94), in partic-

ular, displayed selective antimycotic activity against

the oomycetous fungi with MIC of 25, 6.5, 25, 6.5 lg/mL against P. ultimum, Aphanomyces sp., P. citroph-

thora, P. cinnamomi while hydroxy-jesterone (95)

displayed the antifungal activity against Aphanomyces

sp., P. citrophthora, P. cinnamomi with MIC of 125,

250 and 62.5 lg/mL, respectively (Li and Strobel

2001).

A novel phenolic compound, 4-(2,4,7-trioxa-bicy-

clo[4.1.0]heptan-3-yl) phenol (96) (Fig. 7), was iso-

lated from P. mangiferae, an endophytic fungus

associated with Mangifera indica Linn. The MIC of

compound (96) showed appreciable antibacterial and

antifungal activity against Bacillus subtilis, Klebsiella

pneumoniae and C. albicans (MIC 0.039 lg/mL),

Escherichia coli and M. luteus (MIC 1.25 lg/mL),

followed by Pseudomonas aeruginosa (MIC 5.0 lg/mL). The positive control (gentamycin) displayed

activity against B. subtilis, K. pneumoniae and M.

luteus (MIC 5.0 lg/mL), E. coli and P. aeruginosa

(MIC 10.0 lg/mL) and C. albicans (nystatin; MIC

10.0 lg/mL). The transmission electron microscope

study for the mode of inhibition of compound (96) on

bacterial pathogens revealed the destruction of bacte-

rial cells by cytoplasm agglutination with the forma-

tion of pores in cell wall membranes (Subban et al.

2013).

Bilobalide (97) (Fig. 7), a terpene trilactone

exhibiting neuroprotective effects, was isolated from

endophytic fungi Pestalotiopsis uvicola (strain

GZUYX13) from the leaves of medicinal plantGinkgo

biloba grown in Guizhou province, China. The amount

of bilobalide produced by this endophytic fungus was

quantified to be 106 lg/L via high-performance liquid

chromatography (HPLC) which was substantially

lower than the same produced by the host tissue (Qian

et al. 2016). Polyketide derivative pestalpolyol I (98)

(Fig. 7) was obtained from the endophytic fungus

Pestalotiopsis clavispora isolated from the mangrove

plant Rhizophora harrisonii. Compound (98) exhib-

ited strong cytotoxicity against the mouse lymphoma

cell line L5178Ywith an IC50 value of 4.10 lM (Perez

Hemphill et al. 2016). Prenylated phenols vaccinol I

(99) (Fig. 7), was isolated from endogenous Pestalo-

tiopsis vaccinii (cgmcc3.9199) of mangrove plant

Kandelia candel (L.) Druce (Rhizophoraceae). Com-

pound (99) exhibited potent COX-2 inhibitory activity

with an IC50 value of 16.8 lM (Wang et al. 2015c).

A naphthalene derivative vaccinal A (100) (Fig. 7)

was isolated from P. vaccinii (cgmcc3.9199) endoge-

nous with the mangrove plant K. candel (L.) Druce

(Rhizophoraceae), which is prevalent as a folk

medicine to treat rheumatoid arthritis. The organic

extract (ethyl acetate) of the fermentation broth,

grown over thirty days, displayed antiviral activity

against enterovirus 71 (EV 71). Bio-assay guided

separation of the organic extract yielded eight new

polyketide-derived metabolites of which compound

(100) exhibited in vitro anti-enterovirus 71 (EV71)

activity with IC50 value of 19.2 lM. Initial mass

spectrometry and NMR analysis suggested the pres-

ence of a bicyclic naphthalene skeleton having

aldehyde functionality. Proton COSY and HMBC

analysis were used for further assignments in the

chemical structure of compound (100). Additionally,

compound (100) showed potent COX-2 inhibitory

activity with IC50 value of 1.8 lM (Wang et al. 2014).

A new aromatic amine named pestalamine A (101)

(Fig. 7) was isolated from mangrove-derived endo-

phytic fungus Pestalotiopsis vaccinia which showed

moderate cytotoxicities against MCF-7, HeLa, and

HepG2 human cancer cell lines with IC50 values of

40.3, 22.0, and 32.8 lM, respectively (Zhou et al.

2014).

Pestalafuranones A–E (102–106) (Fig. 7) has been

obtained from the solid-substrate culture of an isolate

of Pestalotiopsis besseyi. Pestalafuranones A–C

(102–104) showed weak inhibitory effects on HIV-1

replication in C8166 cells with EC50 values of 10.52,

24.32 and 36.74 lg/mL, respectively (the EC50 values

of positive controls zidovudine (AZT) and indinavir

Phytochem Rev

123

sulfate (IDV) were 2.17 ng/mL and 3.92 ng/mL,

respectively). Pestalafuranone D (105) showed weak

activity against the plant pathogenic fungi Verticillium

dahiae (CGMCC 3758) with IC50 values of 24.5 lg/mL whereas pestalafuranone E (106) displayed mod-

erate activity against Alternaria longipes (CGMCC

2875) with IC50 values of 10.3 lg/mL (positive

control fluconazole showed IC50 value of 0.50 and

0.30 lg/mL) (Liu et al. 2012). A new oxysporone

derivative, pestalrone B (107) was obtained from the

endophytic plant fungus Pestalotiopsis karstenii iso-

lated from stems of Camellia sasanqua. Compound

(107) exhibited significant activities against HeLa,

HepG2 and U-251 with IC50 values of 12.6, 31.7 and

5.4 lg/mL, respectively (Luo et al. 2012).

Pestalotic acids C, G (108, 109) (Fig. 7) are new

furylidene tetronic acid derivatives obtained from a

plant endophyte Pestalotiopsis yunnanensis isolated

from the branches of Podocarpus macrophyllus. The

structural elucidation was done using NMR spec-

troscopy based 1D and 2D experiments that enabled

proton assignments as well as relative configuration

determination. The absolute configuration of these

structures was determined by circular dichroism

spectroscopy which was also aided by the crystal

structure of a closely related compound. Compounds

(108) and (109) showed significant activity against the

Gram-positive bacteria, S. aureus Col (CGMCC

1.2465) and S. pneumoniae (CGMCC 1.1692) with

MIC values of 6.35–12.76 lM while the positive

control ampicillin showed MIC values of 0.46 and

28.65 lM, respectively (Zhang et al. 2012).

New pyranes, scirpyranes A–C (110–112) (Fig. 7),

were isolated from solid cultures of the plant pathogen

Pestalotiopsis scirpina. Their structural assignments

were done using mass spectrometry and NMR tech-

niques which used both through bond (COSY) and

spatial (NOESY, NOE difference spectra) correla-

tions. The relative and absolute structures were

determined by NOESY and modified Mosher meth-

ods. Compounds (110–112), showed significant cyto-

toxicities towards MCF-7 cells with IC50 values of

5.84, 4.34, and 8.22 lM, respectively (the positive

control cisplatin showed an IC50 value of 11.9 lM) (Li

et al. 2012a). 20-acetyl-40, 4-dimethoxybiphenyl-2-

carbaldehyde (113) (Fig. 7) was obtained from the

plant endophytic fungus Pestalotiopsis zonata isolated

from Cyrtotachys lakka in Hainan, China which

showed antibacterial activity against E. coli, S. aureus,

P. aeruginosa, K. pneumoniae, methicillin resistant S.

aureus, Acinetobacter baumannii and vancomycin-

resistant Enterococcus faecium with IC50 values of

0.75, 0.75, 0.82, 0.81, 0.84, 0.90 and 0.87 lM/mL,

respectively (Yang et al. 2011b).

Bioactive metabolites from unidentified

Pestalotiopsis sp

A new ambuic acid analog (114) (Fig. 8) isolated from

Pestalotiopsis sp. cr013 was obtained from aecios-

pores of Cronartium ribicola. Compound (114) had

weak cytotoxicity against five cancer cell lines with

IC50 values 18.99 lM (HL-60), 17.68 lM (SMMC-

7721), 18.28 lM (A-549), 21.67 lM (MCF-7) and

12.27 lM (SW480) while the control experiment of

MW300 showed cytotoxicity against five cancer cell

lines with IC50 values 1.28 lM (HL-60), 6.72 lM(SMMC-7721), 6.15 lM (A-549), 16.33 mM (MCF-

7) and 12.86 lM (SW480) (Xie et al. 2014).

Ambuic acid (32) and its derivative (115) (Fig. 8)

have been isolated from endophytic fungus Pestalo-

tiopsis sp. inhabiting the lichen Multiclavula sp.

Compound (32) and (115) displayed antimicrobial

activity against the Gram-positive bacterium Staphy-

lococcus aureus (ATCC 6538) with IC50 values of

43.9 and 27.8 lM, respectively (the positive control

ampicillin showed an IC50 value of 1.40 lM) (Ding

et al. 2009b). A new caryophyllene sesquiterpenoid

named pestaloporinate B (116) (Fig. 8) has been

isolated from an endophytic fungus Pestalotiopsis sp.,

which was obtained from the fresh stem bark ofMelia

azedarach Linn. Pestaloporinate B (116) displayed

potent inhibitory activity with IC50 value of 19.0 lMduring the evaluation of nitric oxide (NO) inhibition in

lipopolysaccharide (LPS)-induced RAW264.7 macro-

phage cells (Liu et al. 2016).

Chlorinated benzophenone derivative (±)-pesta-

lachloride D (117) (Fig. 8), and (±)-pestalachloride C

(57) (Fig. 4) were obtained from the marine-derived

fungus Pestalotiopsis sp. isolated from a soft coral

Sarcophyton sp. collected in Yongxing Island in the

South China Sea. Chromatographic and structural

analysis showed the presence of a racemic mixture

whose structure was determined using NMR and

crystallographic techniques. Interestingly, compound

(117) did not exhibit any effect at a concentration of

50 lg/mL in the zebrafish embryo teratogenicity assay

Phytochem Rev

123

Fig. 8 Chemical structures of metabolites isolated from unclassified Pestalotiopsis species

Phytochem Rev

123

Fig. 8 continued

Phytochem Rev

123

while (57) led to abnormal growth effects in several

aspects of the embryonic development. Compound

(57) exhibited coagulated eggs (24 h), non-sponta-

neous movements (24 h), abnormal heartbeat (48 h),

tail (48 h), heart (48 h), notochord malformation

(72 h), delayed hatch (72 h) and embryo death

(72 h) with EC50 values of 16.3, 18.6, 6.3, 24.5, 8.2,

5.8, 7.4 and 12.6 lg/mL, respectively. Compounds

(117) and (57) both exhibited moderate antibacterial

activity against E. coli, Vibrio anguillarum and Vibrio

parahaemolyticus with the MIC values of 5.0, 10.0

and 20.0 lM, respectively (Wei et al. 2013).

A phthalide derivative pestalotiolide A (118), three

known analogs (7-Hydroxy-5-methoxy-4,6-dimethyl-

7-O-b-D-glucopyranosyl-phthalide (119) 7-Hydroxy-

5-methoxy-4,6-dimethyl-7-O-a-L-rhamnosyl-ph-

thalide (120) 7-Hydroxy-5-methoxy-4,6-dimethylph-

thalidc (121) along with 50-O-acetyl uridine (122)

(Fig. 8) were obtained from soft coral-derived fungus

Pestalotiopsis sp. which was isolated from a piece of

fresh tissue from the inner part of a soft coral Sarco-

phyton sp., in Yongxing Island in South China.

Compounds (118–122) possessed varying degrees of

antiviral activities. Compared with ribavirin

(IC50 418.0 lM), pestalotiolide A (118) exhibited

potent anti-EV71 activity, with an IC50 value of

27.7 lM. Compound (119) showed strong antiviral

activities against EV71, RSV, and HSV-1 with

IC50 values of 51.6 lM, 25.6 lM and 63.9 lM,

respectively. Compound (121) displayed pronounced

antiviral activities against Cox-B3 and RSV with

IC50 values of 19.6 lM and 21.0 lM which were

stronger than those of the positive control ribavirin

with IC50 values of 39.0 lM and 78.0 lM, respec-

tively. Compounds (122) and (120) showed similar

antiviral activities against EV71 (Jia et al. 2015a).

Two novel caprolactams, pestalactams A–B (123,

124) were isolated from the Pestalotiopsis sp. (BRRIP

39872) obtained from the plant Melaleuca quinquen-

ervia by static rice fermentation cultures. The metabo-

lite mixture isolated from the culture was initially

purified by reversed phase column chromatography

(C18) using water and methanol as eluents. Further

purification was done using reversed phase (C18, C8)

HPLC to yield the pure compounds. Two-dimensional

NMR experiments (HSQC, HMBC, and ROESY) and

X-ray crystallography were used to determine the

chemical structures. Compounds (123, 124) (Fig. 8)

displayed antimalarial activity against chloroquine

resistant (Dd2) and chloroquine sensitive (3D7) Plas-

modium falciparum with*16–41% growth inhibition

at 25 lM. The compounds displayed *3-fold selec-

tivity for MCF-7 breast cancer cells versus the NFF

control cells (Davis et al. 2010).

Pestalpolyols F–H (125–127) (Fig. 8) were

obtained from mycoparasite Pestalotipsis sp. PG52

isolated from aeciospore piles of Aecidium pourthiaea

and their structures were elucidated by NMR and

single crystal X-ray diffraction techniques. Compound

(125) had cytotoxicity against A-5495 cell line with an

IC50 value of 11.45 lM while compound (126)

showed weak activity against four cell lines with

IC50 values of 14.60 lM (HL-60), 27.46 lM (SMMC-

7721), 11.83 lM (A-549) and 18.50 lM (MCF-7).

Compound (127) had activity against three cell lines

with IC50 values of 22.85 lM (HL-60), 8.05 lM (A-

549) and 38.89 lM (MCF-7) (Xie et al. 2015).

Three novel polyketides, named pestalpolyols A

(128), B (129), and D (130) (Fig. 8) were obtained

from Pestalotiopsis sp. cr013 isolated from aecios-

pores of Cronartium ribicola collected in Yunnan

province, China. Compound (128) showed varying

IC50 values against different cell lines which were

10.4 lM (HL-60), 11.3 lM (SMMC-7721), 2.3 lM(A-549), 13.7 lM (MCF-7), and 12.4 lM (SW480),

respectively. Compound (129) showed an IC50 value

of 10.6 lM against A-549 cell line. Compound (130)

showed IC50 values of 15.7 lM (HL-60), 31.2 lM(SMMC-7721), 10.7 lM (A-549), 23.7 lM (MCF-7),

and 21.4 lM (SW480), respectively (Li et al. 2015b).

A pair of new enantiomeric alkaloid dimers, (?)-

and (-)-pestaloxazine A (131,132), (Fig. 8) with an

unprecedented symmetric spiro[oxazinane-piper-

azinedione] skeleton, having twenty-two carbons and

twelve heteroatoms respectively, were isolated from a

Pestalotiopsis sp. derived from a soft coral in South

China Sea. The enantiomers were separated by chiral

HPLC and extensive NMR analysis followed by single

crystal X-ray analysis helped in deducing their chem-

ical structures. (?)-Pestaloxazine A (131) exhibited

potent antiviral activity against EV71 with an IC50

value of 14.2 ± 1.3 lM, which was stronger than that

of the positive control ribavirin

(IC50 = 256.1 ± 15.1 lM) (Jia et al. 2015b). Ace-

toxydehydroaustin (133), two dihydroisocoumarins:

aspergillumarin A (134) and B (135) and penicillide

(136) (Fig. 8) were obtained from the seagrass-

derived fungus Pestalotiopsis sp. PSU-ES194 which

Phytochem Rev

123

was isolated from the leaves of seagrass, Enhalus

acoroides. Compounds (133) and (136) displayed

weak cytotoxicity to Vero cells with IC50 values of 48

and 84 lM. Only compound (135) displayed very

mild antifungal activity against C. albicans and

Cryptococcus neoformans with equal MIC values of

200 lg/mL (Arunpanichlert et al. 2015). Compounds

(134) and (135) exhibited weak antibacterial activity

against S. aureus and B. subtilis at a concentration of

50 lg/mL (Li et al. 2012b).

Five alkenyl phenol and benzaldehyde derivatives

pestalols A–E (137–141) trans-harzialactone A (142),

F (143), 3b, 5a, 9a-trihydroxy-7, 22-en-ergost-6-one(144) and 3b-hydroxy-sterol (145) (Fig. 8) were

isolated from endophytic fungus Pestalotiopsis sp.

AcBC2 derived from the Chinese mangrove plant

Aegiceras corniculatum. These compounds were iso-

lated from the organic extract (ethyl acetate) of culture

broth and mycelium by repeated purification using

reversed phase silica gel column chromatography as

well as preparative thin layer chromatography. Com-

pounds (137–141) displayed antiproliferative effects

in the range of 23.4–42.5 lMagainst ten human tumor

cell lines representing colon, lung, prostate, ovarian,

breast, cervical, pancreatic and melanoma malignan-

cies. Compounds (137–145) showed inhibitory activ-

ities against influenza A virus subtype (H3N2) and

swine flu (H1N1) viruses. Compound (145) was most

potent with IC50 of 4.7 lMfor virus H3N2 and 2.2 lMfor H1N1 virus. Compound (138) showed tuberculosis

inhibition compared with dimethyl sulfoxide control,

INH (isoniazid) and RIF (rifampin) as positive drugs.

(Sun et al. 2014)

Pestalotiopen A (146) (Fig. 8) was obtained from

Pestalotiopsis sp. isolated from the leaves of the

Chinese mangrove, Rhizophora mucronata which

exhibited moderate antimicrobial activity against E.

faecalis with an MIC value between 125 and 250 lg/mL (Hemberger et al. 2013).Two new sesquiterpenes,

1b,5a,6a,14-tetraacetoxy-9a-benzoyloxy-7bH-eudes-man-2b,11-diol (147) and 4a,5a-diacetoxy-9a-ben-zoyloxy-7bH-eudesman-1b,2b,11,14-tetraol (148)

(Fig. 8) were produced as stress metabolites in the

cultured mycelia of Pestalotiopsis sp. Z233 isolated

from the algae Sargassum horneri in response to

abiotic stress elicitation by CuCl2. Compounds (147,

148) (Fig. 8) showed tyrosinase inhibitory activities

with IC50 values of 14.8 lM and 22.3 lM (Wu et al.

2013).

Two isoprenylated epoxyquinol derivatives

pestaloquinol A (149) and B (150) (Fig. 8) were

obtained from the endophytic fungus Pestalotiopsis

sp. isolated from the branches of Podocarpus macro-

phyllus (Thunb.) D. Don in Kunming, People’s

Republic of China. Bio-assay guided fractionation of

solid-substrate fermentation culture followed by

exhaustive NMR analysis led to the identification of

the novel compounds. Compounds (149) and (150)

were tested for cytotoxicity against HeLa cells which

showed IC50 value of 8.8 lM (the positive controls

VP-16 and D-24851 showed IC50 values of 1.63 and

0.88 lM, respectively) (Ding et al. 2011).

A new chromone, named pestalotiopsone F (151)

(Fig. 8), was obtained from the mangrove endophytic

fungus Pestalotiopsis sp. which was isolated from

leaves of the Chinese mangrove plant R. mucronata.

Organic extraction of the mycelia and culture filtrate

followed by column chromatography and preparative

HPLC separation yielded pure compounds which were

characterized by NMR. Pestalotiopsone F (151) exhib-

ited moderate cytotoxicity against the murine cancer

cell line L5178Y, with an EC50 value of 8.93 lg/mL

(Xu et al. 2009). Two new phytotoxic c-lactones,pestalotines A and B (152, 153) (Fig. 8) were isolated

from the culture of Pestalotiopsis sp. HC02, a fungus

residing in the Chondracris rosea gut. Pestalotines A

and B (152–153) significantly inhibited the radical

growth of Echinochloa crusgalli with IC50 values of

1.85 9 10-4 and 2.50 9 10-4 M, respectively, com-

parable to that of 2-(2,4-dichlorophenoxy)acetic acid

(0.94 9 10-4 M) used as a positive control (Zhang

et al. 2008). RES-1214-1 and -2, (154–155) (Fig. 8)

novel and non-peptidic endothelin antagonists were

obtained from the culture broth of a fungus, Pestalo-

tiopsis sp. RE-1214 isolated from a soil sample

collected in Kanagawa Pref., Japan. RES-1214-1 and

RES-1214-2 (154, 155) selectively inhibited the ET-1

binding to endothelin type A receptor (ETA receptor)

with IC50 values of 1.5 lM and 20 lM, respectively.

RES-1214-1 and RES-1214-2 inhibited the increase in

intracellular Ca2? concentration elicited by 1 nMET-1

in A10 cells (Ogawa et al. 1995).

Overview of biological functions

As discussed before, metbolites reported from the

genus Pestalotiopsis display a range of biological

Phytochem Rev

123

activities which could significantly help in the dis-

covery and design of small molecules for curing fatal

ailments of current times e.g. cancer and HIV-AIDS.

However, a major limitation in the progress of taking

these metabolites to market has been the lack of a

focused and concerted approach for target validation

and subsequent structure–activity relationship based

design efforts. Except for the well-known case of

taxol, the majority of the biological activities reported

from the metabolites of Pestalotiopsis, do not shed

light on the biological mechanism or targets of these

compounds. For example, the anti-HIV activities of

petalafone F (5) and pestalafuranones A–E (102–106)

do not identify eventual molecular targets that inhibit

HIV replication process. In the absence of such details,

structural motifs that are essential to inhibitory

functions cannot be determined. Since few molecular

targets (such as reverse transcriptase, integrase in the

case of HIV) are structurally well defined and their

cellular activity is fairly well understood, molecules

that complement the molecular environment of the

active site can be tailored and tuned to elicit the

desired effects. One possible reason for the lack of

follow-up research to establish molecular targets and

to understand the underlying mechanism of interaction

is relatively very high inhibitory concentrations

required by the test molecules to achieve desired

activity in comparison to their positive controls. For

instance, Pestaloficiol J (19) displayed an EC50 value

8.0 lM, much poorer to the EC50 value of positive

control indinavir (8.2 nM). A similar gap in the

activity of test compounds and the positive controls

were also evident in the cases of Pestaloficiol A, B, D

(21–23).

Metabolites of Pestalotiopsis show cytotoxic activ-

ity with a wide range of IC50 values (Table 2). While

most of the studies lack standard compounds to define

their relative effectiveness to a positive control, some

that have a positive control (28, 50–54) show signif-

icant differences in the inhibitory concentrations. Such

large differences in the IC50 values make them

unlikely to be of significant therapeutic value. More-

over, the absence of a positive control in most of the

cytotoxic activity determinations does not provide a

reference point and such values do not represent the

true potential of those metabolites. This is especially

pertinent for antineoplastic agents where selectivity in

targeting a tumor cell over a normal cell is a key

determinant in the drug development. However, some

compounds have shown excellent ‘cytotoxic activity

against cancer cell lines. For example, chlorop-

upukeanolides (16) and (17) have shown nearly six

and four fold better cytotoxic acivity against cancer

cell line HeLa than the positive control 5-fluorouracil.

These compounds also showed nearly two and four

fold better activity against colon cancer cell line HT29

as compared with the positive control 5-fluorouracil.

Some of the other metabolites have also shown

promising antiviral activity. Pestalotiolide A (118)

and compound (121) displayed 15–20 fold higher

antiviral activities than the positive control ribavarin.

The antifungal activities displayed by these

metabolites, however, show far more significant

promise with compounds (3, 6, 7) displaying much

better antifungal activities than positive control

fluconazole. The excellent antifungal activities shown

by Pestalotiopsis metabolites clearly warrant further

studies to understand their mechanism of action.

While a major part of natural product based drug

discovery significantly relies on structural elucidation

of new metabolites, the identification of appropriate

molecular targets also provides thrust to undertake

detailed studies. Although, elucidation of chemical

structure, in some cases, can be extremely time-

consuming, the absence of defined molecular targets

can render them to bemolecules in search of functions.

It is noteworthy that nearly all of the literature

examples covered in this review, while reporting its

biological significance; do not provide a rational

approach towards its target validation. The anticancer,

anti-HIV or antifungal activities reported here are

arbitrarily chosen and in doing so, many of its other

possible targets might have been skipped. For exam-

ple, the structural diversity shown by the metabolites

of Pestalotiopsis highlights some features that favor

their nucleic acid binding. Pestalofones E, (50–52),

pestlazines (75–77) represent structural features that

might result in preferential nucleic acid binding than

proteins. This is due to the presence of fused ring

systems with extended unsaturation and appendages

that would have additional contacts in the form of

hydrogen binding and van der Walls interactions. In

addition to the possibility of end stacking and inter-

calative binding in duplex and triplex nucleic acid

structures, these molecules are highly likely to interact

with four stranded nucleic acid structures which form

a network of four guanines connected by eight

Hoogsteen hydrogen bonds resulting in a planar

Phytochem Rev

123

Ta

ble

2Novel

bioactivecompoundsreported

from

thegenusPestalotiopsis

Endophytic

fungal

strain

Host

plant(s)

(fam

ily),

Plantpartortissue/

Locality

ofhost

plants

Isolatedmetabolite

Testedsystem

sActivityresponse

References

Pestalotiopsisfici

Unidentified

plant

Branches

PestalofoneA

(4),B

(5),C

(6)andE(7)

HIV

-1replicationin

C8166cells

EC50values

of90.4,64.0,and

93.7

lM

Liu

etal.(2009a)

Hangzhou(Eastchina)

PestalofoneC(6)andE

(7)

A.fumigatus

IC50/M

ICvalues

of1.10/35.3,0.90/

31.2

lM,respectively(the

positivecontrolfluconazole

showed

IC50/M

ICvalues

of7.35/

163.4

lM)

P.fici

Camelliasinensis

(Theaceae)

Branches

Hangzhou,P.R.China

PestalofoneF(8)

HeL

aandMCF-7

cells

IC50values

of14.4

and11.9

lM,

respectively

Liu

etal.(2011a)

PestalodiolC

(9)

HeL

aandMCF-7

cells

IC50values

of16.7

and57.5

lM

P.fici

C.sinensis(Theaceae)

Branches

Hangzhou,P.R.China

PestalofoneJ(1

0)

HeL

a,T24,A549,and

MCF-7

celllines

IC50values

of44.3,39.3,35.3,and

38.3

lm

Wanget

al.(2016)

PestalofoneK

(11)

HeL

a,T24,A549,and

MCF-7

celllines

IC50values

of65.5,45.7,58.9,and

29.2

lM

Cisplatinshowed

theIC

50values

of

7.4,3.89,8.4,and6.4

lM,

respectively

P.fici

Unidentified

plant

Branches

Hangzhou,P.R.China

Chloropupukeananin

(12)

HIV

-1replicationin

C8166cells

IC50valueof14.6

lM

Liu

etal.(2008a)

HeL

aandHT29cells

S.aureus(A

TCC6538)

IC50values

of1.4

and6.7

lM,

IC50andMIC

values

of21.8

and

97.3

lM,

P.fici

Unidentified

plant

Branches

Hangzhou,P.R.China

ChloropestolideA

(13)

HeL

aandHT29

GI 50values

of0.7

and4.2

lM,

Liu

etal.(2009b)

P.fici

Unidentified

plant

Branches

Hangzhou,P.R.China

ChloropestolideB

(14)

CNE1-LMP1,A375and

MCF-7

IC50values

of16.4,9.9,and

23.6

lM,

Liu

etal.(2013a)

P.fici

Unidentified

plant

Branches

Hangzhou,P.R.China

Chloropupukeanolide

A(1

5)

HIV

-1replicationin

C8166cells

EC50valueof6.9

lMLiu

etal.(2010)

HeL

a,MCF-7

and

MDA-M

B-231cell

lines

IC50values

of16.9,15.5

and15.9

lM

P.fici

Unidentified

plant

Branches

Hangzhou,P.R.China

Chloropupukeanolide

C(1

6)andD

(17)

HeL

a(cervical

epithelium)and

HT29(colon)cell

lines,

IC50values

rangingfrom

1.2

to7.9

lM

Liu

etal.(2011b)

5-fluorouracil,whichgaveIC

50

values

of10.0

and15.0

lM

Phytochem Rev

123

Ta

ble

2continued

Endophytic

fungal

strain

Hostplant(s)

(fam

ily),

Plantpartortissue/

Locality

ofhost

plants

Isolatedmetabolite

Testedsystem

sActivityresponse

References

P.fici

C.sinensis

Branches

Hangzhou,P.R.China

Siccayne(1

8)

HeL

aandHT29

IC50values

of48.2

and33.9

lM

Liu

etal.(2013b)

P.fici

C.sinensis

Branches

Hangzhou,P.R.China

PestaloficiolJ(1

9)

HIV

-1replicationin

C8166cells

EC50valueof8.0

lM

(theCC50

valueisgreater

than

100lM;the

positivecontrolindinavirsulfate

showed

anEC50valueof8.2

nM)

Liu

etal.(2009c)

PestaloficiolL(2

0)

HeL

aandMCF7cells

IC50values

of8.7

and17.4

lM,(5-

fluorouracilIC

50values

of10.0

and15.0

lM,).

P.fici

Unidentified

plant

Branches

Hangzhou,P.R.China

PestaloficiolA

(21),B

(22)andD

(23)

HIV

-1replicationin

C8166cells

EC50values

of26.0,98.1,and64.1

lM,respectively(allthree

compoundsshowed

CC50values

ofgreater

than

200lM;the

positivecontrolindinavirsulfate

showed

anEC50valueof

8.81nM).

Liu

etal.(2008b)

P.fici.

Unidentified

plant

Branches

Hangzhou,P.R.China

PestaloficiolN

(24)

HIV

-1replicationin

C8166cells

Liu

andLiu,(2010)

PestaloficiolO

(25),P

(26)

HeL

acellline

PestaloficiolO

(25)

A.fumigatus

P.fici

C.sinensis

Branches

Hangzhou,P.R.China

FicipyroneA

(27)

G.zeae(CGMCC

3.2873)

IC50valueof15.9

lM

(thepositive

controlketoconazole

showed

an

IC50valueof6.02lM)

Liu

etal.(2013c)

P.fici

C.sinensis

Branches

Hangzhou,P.R.China

PestalotriolB

(28),

HeL

acells

IC50valueof87.0

lM

(cisplatin

showed

anIC

50valueof7.4

lM).

Liu

etal.(2015)

Pestalotiopsis

microspora

Artificially

infected

Florida

torreya(Torreya

taxifolia)

Inner

barkof

symptomless

trees,

NorthAmerica

Pestalopyrone(2

9),

Hydroxypestalopyrone

(30),andpestaloside

(31)

Cladosporium

sp.

sterilehyphomycete

R.solani,G.

candidurn

andA.

compestris,

Lee

etal.(1995)

P.microspora

Manytropical

plantspecies

Sydney,

Ausralia

Ambuic

acid

(32),

P.ultimum

MIC

valueof7.5

lg/m

LLiet

al.(2001)

Phytochem Rev

123

Ta

ble

2continued

Endophytic

fungal

strain

Host

plant(s)

(fam

ily),

Plantpartortissue/

Locality

ofhost

plants

Isolatedmetabolite

Testedsystem

sActivityresponse

References

P.microspora

Taxuschinensis

Branch

Yichang,Hubei

Province,China

LL-P880c(3

3)

Showed

significant

gibberellin

synergisticactivity

towardsDistylium

chinense

seeds,with

thesubstrate

LL-

P880cconcentration

of0.6

mg/L.The

germinationrate

of

theseedswas

85.56%

Liet

al.(2015a)

P.microspora

-PapuaNew

Guinea

Pestacin(3

4)

P.ultimum

Exhibitsantioxidantactivity11

times

greater

than

thevitam

inE

derivativetrolox

MIC

of*

10lg/m

L

Harper

etal.(2003)

P.microspora

Terminaliamorobensis

NorthcoastofPapua

New

Guinea

Isopestacin(3

5)

P.ultimum,

Totalinhibitionat

40mg/m

Lat

48h.

Antioxidantscavengingboth

superoxideandhydroxyfree

radicals

Strobel

etal.(2002)

P.microspora

Torreyataxifolia

-Torreyanic

acid

(36)

Protein

kinaseC(PKC)

agonists

IC50values

rangefrom

3.5

(NEC)

to45(A

549)lg/m

Lwithamean

valueof9.4

lg/m

Lfor25

differentcelllines

andcausescell

death

byapoptosis..Torreyanic

acid

also

showsG1arrest

ofG0

synchronized

cellsat

the1–5lg/

mLlevel

dependingonthecell

line

Lee

etal.(1996)

Pestalotiopsis

foedan

Bruguiera

sexangula

Branch

Hainan,China

(-)-(4S,8S)-

Foedanolide(3

7)HeL

a,A549,U251,

HepG2,MCF-7

IC50values

of15.8,296.0,159.0,

22.8,70.2

lg/m

L

YangandLi(2013)

(?)-(4R,8R)-

foedanolide(3

8)

HeL

a,A549,U251,

HepG2,MCF-7

IC50values

of5.4,67.9,53.0,19.0,

20.8

lg/m

L

P.foedan

Bruguiera

sexangula.

Branch

Hainan,China

(3R,4R,6R,7S)-7-

hydroxyl-3,7-

dim

ethyl-oxabicy-

clo[3.3.1]nonan-2-

one(3

9),(3R,4R)-3-

(7-m

ethylcyclo-

hexenyl)-propanoic

acid

(40)

B.cinerea

andP.

nicotianae

MIC

values

of3.1

and6.3

lg/m

LXuet

al.(2016a)

Phytochem Rev

123

Ta

ble

2continued

Endophytic

fungal

strain

Host

plant(s)

(fam

ily),

Plantpartortissue/

Locality

ofhost

plants

Isolatedmetabolite

Testedsystem

sActivityresponse

References

(3R,4R)-3-(7-

methylcyclo-

hexenyl)-propanoic

acid

(40)

C.albicans

MIC

valueof50lg/m

L

P.foedan.

Unidentidfied

tree

Branches,

Dongzai,

Hainan

Province

PestafolideA

(41)

A.fumigatus(A

TCC

10894)

Zoneofinhibitionof10mm

at

100lg

/disk

Dinget

al.(2008a)

PestaphthalideA

(42)

C.albicans(A

TCC

10231),

Zoneofinhibitionof13mm

at100

lg/disk

PestaphthalideB(4

3)

G.candidum(A

S2.498)

11mm

zoneofinhibitionat

100lg

/disk

Pestalotiopsis

adusta

Clerodendrum

canescens

Stems

ZhejiangProvince,

People’s

Republicof

China

(10S)-12,16-epoxy-

17(15?

16)-abeo-

3,5,8,12,15-

abietapentaen-

2,7,11,14-tetraone

(44),teuvincenoneF

(45),uncinatone(4

6),

coleonU

(47)and

coleonU-12-M

e

ether

(48)

HL-60

IC50values

of12.54,25.06,15.66,

57.60,and66.41lM,

respectively(CisplatinwithIC

50

of9.20lM

)

Xuet

al.(2016b)

P.adusta(L416),

Unidentified

tree

Stem

Hainan

Province,

People’s

Republicof

China

PestalachlorideA

(49)

F.culmorum

IC50valueof0.89lM

Liet

al.(2008a)

PestalachlorideB

(50)

G.zeae

IC50valueof1.1

lM,.

PestalachlorideC

(51)

F.culmorum,G.zeae,

andV.aibo-atrum

IC50valueof[

100lM

Pestalotiopsis

foedan

Roystonea

regia

Hainan

Province,P.

R.China.

Photinides

A-F

(52–57)

MDA-M

B-2311

Withinhibitory

ratesof24.4,24.2,

23.1,24.4,and24.6%,

respectively,when

tested

at

10lg/m

L

Dinget

al.(2009a)

Pestalotiopsis

photiniae(L461)

Roystonea

regia

(H.B.K.)

Cook

Hainan

Province,P.

R.China

PhotipyroneB

(58)

MDA-M

B-231

Withinhibitory

rate

at25.0%

and

23.0%,respectively,when

tested

at10lg/m

L

Dinget

al.(2012)

P.photiniae

Podocarpusmacrophyllus

MP[4-(30 ,30 -

dim

ethylallyloxy)-5-

methyl-6-

methoxyphthalide]

(59)

HeL

a,MDA-M

B-231,

MCF-7

andMRC5

celllines

IC50valueof36,51,81and

147lg

/mL

Chen

andYang(2013)and

Chen

etal.(2013)

Phytochem Rev

123

Ta

ble

2continued

Endophytic

fungal

strain

Host

plant(s)

(fam

ily),

Plantpartortissue/

Locality

ofhost

plants

Isolatedmetabolite

Testedsystem

sActivityresponse

References

P.photiniae

P.macrophyllus

Branch,

Hainan,P.R.China

5-(30 -methyl-20 -

butenyl)-2-hydroxy-

3-m

ethoxy-4-

methylbenzoic

acid

(60),5-(30 -carboxyl-

30 -methyl-2E-

allyloxy)-3-m

ethoxy-

4-m

ethylphthalide

(61),5-(30 ,3

0 -dim

ethylallyloxy)-2-

methoxycarbonyl-3-

methoxy-4-

methylbenzoic

acid

(62)5-(30 ,3

0 -dim

ethylallyloxy)-3-

methoxy-4-

methylphthalide(6

3),

zinnim

idine(6

4),

5-(30 ,30 -

dim

ethylallyloxy)-3-

methoxy-4-

methylphthalide(6

5),

5-(30 ,30 -

dim

ethylallyloxy)-3-

methoxy-4-

methylphthalic

acid

(66),zinniol

anhydride(6

7)and

porriolide(6

8)

F.graminearum,B.

cinerea

andP.

nicotianae

MIC

values

from

50.0–3.1

lg/m

L

(thepositivecontrol

ketoconazole

showed

MIC

values

are3.1

lg/m

L)

Yanget

al.(2011a)

Pestalotiopsis

theae

Fagara

zanthoxyloides

Fresh

leaves

Nsukka,

Eastern

Nigeria

Chloroisosulochrin(6

9)

InhibitionoftheRSV

InhibitionoftheRSV

IC50valueof4.22±

1.03lM

Uzoret

al.(2016)

FicipyroneA

(70)and

Pestheicacid

(71)

IC50valueof45.00±

0.98and

146.20±

2.14lM

P.theae.

Tea

grayblightfungi

PTtoxin

(72)(?

)-

Epiepoxydon(7

3),

Oxysporone(7

4)

Thethreshold

concentrationsof

inducedleaf

necrosis

foracultivar

YabukitabyII,I,and

oxysporone(III)were

4,60,and15lg/m

L,

respectively

Nagataet

al.(1992)

Phytochem Rev

123

Ta

ble

2continued

Endophytic

fungal

strain

Host

plant(s)

(fam

ily),

Plantpartortissue/

Locality

ofhost

plants

Isolatedmetabolite

Testedsystem

sActivityresponse

References

P.theae

Plantpathogenic

fungus

Branches,Hangzhou,

ZhejiangProvince,

People’s

Republicof

China

PestalazineA

(75),

PestalamideA

(76),

Asperazine(7

7)

HIV

-1replicationin

C8166cells

EC50values

of47.6,64.2,and

98.9

lM,(theCC50values

for

thesecompoundsareallgreater

than

100lM;thepositivecontrol

indinavirsulfateshowed

anEC50

valueof5.5

nM)

Dinget

al.(2008b)

PestalamideA

(76)

A.fumigatus(A

TCC

10894),

IC50/M

ICvalues

of1.50/57.8

lM

(thepositivecontrolfluconazole

showed

IC50/M

ICvalues

of7.35/

163.4

lM)

P.theae

Unidentified

tree

Branches,Hainan

Province,P.R.China

PestalotheolC

(78)

HIV

-1LAIreplicationin

C8166cells

EC50valueof16.1

lM

Liet

al.(2008b)

Pestalotiopsis

virgatula

K.obovata

Branch

2-(1-m

ethoxy-1H-

indole-3-yl)ethanol

(79),2-(1-m

ethoxy-

1H-indole-3-yl)

acetic

acid

(80)

A549,MCF-7,HeL

A,

U251,HepG-2

LiandYang(2014)

Pestalotiopsis

sydowiana

Halophyte,Phragmites

communis

Trinus.

Rhizome,

Suncheon,South

Korea

Compounds

(81,8

2,8

3,8

4,

85,8

6,8

7)

Compounds1–3,5,8,

9–10inhibited

the

activityofthe20S

proteasomein

adose-

dependentmanner,

IC50values

rangingfrom

1.2

±

0.3

lMto

30.5

±

1.5

lM

Xia

etal.(2016)

Pestalotiopsis(ZJ-

2009-7-6)

Marine-derived,softCoral-

Sarcophytonsp.

South

ChinaSea

(±)-pestalachlorideE

(88)andF(8

9)

Potentantifouling

activitiesagainst

the

larval

settlementof

thebarnacle

B.

amphitrite

at

nontoxic

concentrationswith

EC50values

of1.65

and0.55lg/m

L,

respectively

Xinget

al.(2016)

Pestalotiopsis

neglecta

C.sinensis

Twig,

Fujian

Province

of

China

Ambuic

acid

derivatives

(90–92)

Nitricoxide(N

O)

inhibitionassayin

the

lipopolysaccharide

(LPS)-induced

macrophage

IC50values

of88.66,11.20,and

20.80lM,respectively

Qiet

al.(2015)

Pestalotiopsis

manguiferae

Hyptisdilatata

Central

provincesof

Panam

a

Mangiferaelactone(9

3)

L.monocytogenes

MIC

valueof1.68lg/m

LOrtegaet

al.(2014)

B.cereus

MIC

values

of0.55lg

/mL

Phytochem Rev

123

Ta

ble

2continued

Endophytic

fungal

strain

Host

plant(s)

(fam

ily),

Plantpartortissue/

Locality

ofhost

plants

Isolatedmetabolite

Testedsystem

sActivityresponse

References

Pestalotiopsis

jesteri

Fragraea

bodenii

SouthernHighland

Province

ofPapua

New

Guinea

Jesterone(9

4)

P.ultimum,

Aphanomyces

sp.,P.

citrophthora,P.

cinnamomi

MIC

valueof25,6.5,25.6.5

lg/

mL

LiandStrobel

(2001)

Hydroxy-jesterone(9

5)

Aphanomyces

sp.,P.

citrophthora,P.

cinnamomi

MIC

valueof125,250and

62.5

lg/m

L,respectively

Pestalotiopsis

mangiferae

Mangiferaindica

Tam

ilNaduProvince,

India

4-(2,4,7-trioxa-

bicyclo[4.1.0]heptan-

3-yl)phenol(9

6)

B.subtilis,K.

pneumoniaeandC.

albicans

MIC

values

of0.039lg/m

LSubban

etal.(2013)

E.coliandM.luteus

MIC

values

of1.25lg/m

L

P.aeruginosa

MIC

values

of5.0

lg/m

Lpositive

control(gentamycin)displayed

activityagainst

B.subtilis,K.

pneumoniaeandM.luteus(M

IC

5.0

lg/m

L),E.coliandP.

aeruginosa

(MIC

10.0

lg/m

L)

andC.albicans(nystatin;MIC

10.0

lg/m

L).

Pestalotiopsis

uvicola

(strain

GZUYX13)

Ginkgobiloba

Leaves

Bilobalide( 9

7)

Qianet

al.(2016)

Pestalotiopsis

clavispora

R.harrisonii

Petioles,

PortHarcourt(N

igeria)

PestalpolyolI(9

8)

L5178Y

IC50valueof4.10lM

Perez

Hem

phillet

al.(2016)

P.vaccinii

(cgmcc3.9199)

K.candel

SouthernChina

VaccinolI(9

9)

COX-2

inhibitory

activity

IC50valueof16.8

lM

Wanget

al.(2015c)

Pestalotiopsis

vaccinii

(cgmcc3.9199)

K.candel

SouthernChina

Vaccinal

A(1

00)

Anti-enterovirus71

(EV71)

IC50valueof19.2

lM

Wanget

al.(2014)

P.vaccinii

(cgmcc3.9199)

K.candel

SouthernChina

Vaccinal

A(1

00)

COX-2

inhibitory

IC50valueof1.8

lM

Wanget

al.(2014)

P.vaccinia

Branch,southernChina

PestalamineA

(101)

MCF-7,HeL

a,and

HepG2human

cancer

celllines

IC50values

of40.3,22.0,and

32.8

lM,

Zhouet

al.(2014)

Phytochem Rev

123

Ta

ble

2continued

Endophytic

fungal

strain

Host

plant(s)

(fam

ily),

Plantpartortissue/

Locality

ofhost

plants

Isolatedmetabolite

Testedsystem

sActivityresponse

References

Pestalotiopsis

besseyi.

-DonglingMountain,

Beijing

Pestalafuranones

A–E

(102–106)

PestalafuranoneD

(105)

PestalafuranoneE

(106)

HIV

-1replicationin

C8166cells

EC50values

of10.52,24.32and

36.74lg/m

L,(theEC50values

of

positivecontrolzidovudine

(AZT)andindinavirsulfate

(IDV)were2.17ng/m

Land

3.92ng/m

L,respectively)

Liu

etal.(2012)

V.dahiae(CGMCC

3758)

IC50values

of24.5

lg/m

L

A.longipes

(CGMCC

2875)

IC50values

of10.3

lg/m

L

Pestalotiopsis

karstenii

Camelliasasanqua.

pestalroneB

(107)

HeL

a,HepG2and

U-251

IC50values

of12.6,31.7

and

5.4

lg/m

L,

Luoet

al.(2012)

Pestalotiopsis

yunnanensis

Podocarpusmacrophyllus

Branches

Kunming,People’s

RepublicofChina

Pestaloticacid

C-G

(108–109)

S.aureusCol(CGMCC

1.2465)andS.

pneumoniae

(CGMCC

1.1692)

MIC

values

of6.35–12.76lM

Zhanget

al.(2012)

Pestalotiopsis

scirpina

Myristica

yunnanensis

Branch,

Yunnan,People’s

RepublicofChina

ScirpyraneA-C

(110,1

11,1

12)

MCF-7

IC50values

of5.84,4.34,and

8.22lM,

Liet

al.(2012a)

Pestalotiopsis

zonata

Cyrtotachys

lakka

Hainan,China

20 -acetyl-40 ,

4-dim

ethoxy-

biphenyl-2-

carbaldehyde(1

13)

E.coli,S.aureus,P.

aeruginosa,K.

pneumoniae,

methicillinresistant

S.aureus,A.

baumanniiand

vancomycin-resistant

E.faecium

IC50values

of0.75,0.75,0.82,

0.81,0.84,0.90and0.87lM/m

L

Yanget

al.(2011b)

Phytochem Rev

123

Ta

ble

2continued

Endophytic

fungal

strain

Hostplant(s)

(fam

ily),

Plantpartortissue/

Locality

ofhost

plants

Isolatedmetabolite

Testedsystem

sActivityresponse

References

Pestalotiopsissp.

cr013

AeciosporesofCronartium

ribicola

Kunming,Yunnan

Province,People’s

Republicof

China

Ambuic

acid

analog

(114)

HL-60,

SMMC-7721

A-549,

MCF-7

SW480

Control-MW300

HL-60,

SMMC-7721

A-549,

MCF-7

SW480

IC50values

of18.99lM

17.68lM

18.28lM

21.67lM

12.27lM

1.28lM,

6.72lM

6.15lM

16.33lM

12.86lM

Xie

etal.(2014)

Pestalotiopsissp.

Lichen

Multiclavula

sp.

Hainan

Province,

People’s

Republicof

China

Ambuic

acid

(32)and

S.aureus(A

TCC

6538),with

IC50values

of43.9

and27.8

lM,

respectively(thepositivecontrol

AMPshowed

anIC

50valueof

1.40lM)

Dinget

al.(2009b)

Ambuic

acid

derivative

(115)

Pestalotiopsissp.,

M.azedarach

Stem

bark

Jiangsu

Province,People’s

RepublicofChina

Pestaloporinate

B(1

16)

Nitricoxideinhibition

inlipopolysaccharide

(LPS)-inducedRAW

264.7

macrophage

cells

IC50valueof19.0

lMLiu

etal.(2016)

Pestalotiopsissp.

SoftcoralSarcophytonsp.

YongxingIslandin

the

South

ChinaSea

(±)-pestalachlorideC

(57),

Zebrafish

embryo

teratogenicityassay

Led

toabnorm

algrowth

effectsin

several

aspects

oftheem

bryonic

development

Wei

etal.(2013)

Pestalotiopsissp.

SoftcoralSarcophytonsp.

YongxingIslandin

the

South

ChinaSea

(±)-pestalachlorideD

(117),and

(±)-

PestalachlorideC

(57),

E.coli,V.anguillarum

andV.

parahaem

olyticus

MIC

values

of5.0,10.0

and

20.0

lM,

Wei

etal.(2013)

Phytochem Rev

123

Ta

ble

2continued

Endophytic

fungal

strain

Hostplant(s)

(fam

ily),

Plantpartortissue/

Locality

ofhost

plants

Isolatedmetabolite

Testedsystem

sActivityresponse

References

Pestalotiopsissp.

SoftcoralSarcophytonsp.

South

ChinaSea

PestalotiolideA

(118),

EV71activity

IC50valueof27.7

lM.

Jiaet

al.(2015a)

7-H

ydroxy-5-m

ethoxy-

4,6-dim

ethyl-7-O

-b-

D-glucopyranosyl-

phthalide(1

19)

EV71,RSV,andHSV-

1

IC50values

of51.6

lM,25.6

lM

and63.9

lM

7-H

ydroxy-5-m

ethoxy-

4,6-dim

ethyl-7-O

-a-

L-rham

nosyl-

phthalide(1

20)

EV71,Cox-B3

IC50values

of111lM

and

95.9

lM

7-H

ydroxy-5-m

ethoxy-

4,6-

dim

ethylphthalidc

(121)

Cox-B3andRSV

IC50values

of19.6

lM

and

21.0

lM

50 -O-acetyluridine

(122)

EV71,Cox-B3

IC50values

of110lM

and

127lM

Compared

toribavirin

(IC50418.0

lM),

Pestalotiopsissp.

Melaleuca

quinquenervia

Toohey

Forest,

Queensland,

Australia

Pestalactam

sA-B

(123–124)

Chloroquineresistant

(Dd2)and

chloroquinesensitive

(3D7)Plasm

odium

falciparum

*16–41%

inhibitiongrowth

at

25lM

Davis

etal.(2010)

*3-fold

selectivityforMCF-7

breastcancercellsversusthe

NFFcontrolcells

Pestalotipsissp.

PG52

Aeciospore

piles

of

Aecidium

pourthiaea

Yunnan

Province,People’s

RepublicofChina

PestalpolyolF,(1

25)

A-5495

IC50valueof11.45lM

Xie

etal.(2015)

G(1

26)

HL-60

SMMC-7721A-549

MCF-7

IC50values

of14.60,27.46,11.83,

and18.50lM

H(1

27)

HL-60

A-549

MCF-7

IC50values

of22.85,8.05,and

38.89lM

Phytochem Rev

123

Ta

ble

2continued

Endophytic

fungal

strain

Host

plant(s)

(fam

ily),

Plantpartortissue/

Locality

ofhost

plants

Isolatedmetabolite

Testedsystem

sActivityresponse

References

Pestalotiopsissp.

cr013

AeciosporesofCronartium

ribicola

collectedfrom

Pinusarm

andii

Yunnan

Province,

People&sRepublic

ofChina

PestalpolyolA

(128)

HL-60SMMC-7721

MCF-7

A-549

SW480

IC50values

of10.4,11.3,2.3,13.7,

and12.4

lM

Liet

al.(2015b)

Pestalpolyol

B(1

29)

A-549

IC50valueof10.6

lM.

PestalpolyolD

(130)

HL-60

SMMC-7721

A-549

MCF-7

SW480

IC50values

of15.7,31.2,10.7,

23.7,and21.4

lM

Pestalotiopsissp.

Softcoral

South

ChinaSea

(?)-PestaloxazineA

(131)

EV71

IC50valueof14.2

±1.3

lM

positivecontrolribavirin

(IC50=

256.1

±15.1

lM)

Jiaet

al.(2015b)

Pestalotiopsissp.

PSU-ES194

Seagrass,Enhalus

acoroides

Leaves,

Thailand

Acetoxydehydroaustin

(133)andPenicillide

(136)

Verocells

IC50values

of48and84lM.

Arunpanichlertet

al.(2015)

Pestalotiopsissp.

PSU-ES194

Seagrass,E.acoroides

Leaves,

Thailand

Aspergillumarin

B

(135)

C.albicansandC.

neoform

ans

MIC

values

of200lg/m

L.

Arunpanichlertet

al.(2015)

Aspergillumarin

Aand

B(1

34–135)

S.aureusandB.

subtilis

Exhibitweakantibacterial

activity

ataconcentrationof50lg/m

L

Liet

al.(2012b)

Pestalotiopsissp.

AcB

C2

Aegicerascorniculatum

SouthernChina

Pestalols

A-E

.(1

37–141)

TransharzialactoneA,

(142)

TransharzialactoneF

(143)3b,

5a,9a-

trihydroxy-7,22-en-

ergost-6-one(1

44),

3b-hydroxy-sterol

(145).

Influenza

Avirus

subtype(H

3N2)and

swineflu(H

1N1)

viruses

Sunet

al.(2014)

Pestalotiopsissp.

AcB

C2

A.corniculatum

Guangdongprovince,

China

3b-hydroxy-sterol

(145),

H3N2

H1N1

IC50valueof4.7

lM

Sunet

al.(2014)

IC50valueof2.2

lM

Phytochem Rev

123

Ta

ble

2continued

Endophytic

fungal

strain

Hostplant(s)

(fam

ily),

Plantpartortissue/

Locality

ofhost

plants

Isolatedmetabolite

Testedsystem

sActivityresponse

References

Pestalotiopsissp.

AcB

C2

A.corniculatum

Guangdongprovince,

China

PestalolB(1

38)

Inhibitionto

tuberculosis

compared

with

dim

ethylsulfoxide

controlwiththeIN

H

(isoniazid)andRIF

(rifam

pin)as

positive

drugs

Sunet

al.(2014)

Pestalotiopsissp.

Rhizophora

mucronata

Leaves

Hainan

Island,China

Pestalotiopen

A(1

46)

Enterococcusfaecalis

MIC

valuebetween125and250

lg/m

L

Hem

berger

etal.(2013)

Pestalotiopsissp.

Z233

Algae

Sargassum

horneri

Wenzhou,China

1b,5a,6a,14-

tetraacetoxy-9a-

benzoyloxy-7bH-

eudesman-2b,11-diol

(147)and4a,5a-

diacetoxy-9a-

benzoyloxy-7bH-

eudesman-

1b,2b,11,14-tetraol

(148)

Tyrosinaseinhibitory

activities

IC50valueof14.8

lMand

22.3

lMWuet

al.(2013)

Pestalotiopsissp.

Podocarpusmacrophyllus

Branches

Kunming,People’s

RepublicofChina

PestaloquinolA

(149)

andB

(150)

HeL

a(cervical

epithelium)

IC50valueof8.8

lMDinget

al.(2011)

Positivecontrols

VP-16and

D-24851showed

IC50values

of

1.63and0.88lM

Pestalotiopsissp.,

R.mucronata

Leaves

Hainan

Island,China

PestalotiopsoneF(1

51)

L5178Y,

EC50valueof8.93lg

/mL

Xuet

al.(2009)

Pestalotiopsissp.

HC02,

Chondracris

roseagut

JiangxiProvince,P.

R.China

PestalotineA

andB

(152–153)

E.crusgalli

Inhibited

theradical

growth

ofwith

IC50values

of1.859

10-4and

2.509

10-4M,respectively,

comparable

tothat

of2-(2,4-

dichlorophenoxy)aceticacid

(0.949

10-4M)usedas

a

positivecontrol

Zhanget

al.(2008)

Phytochem Rev

123

assembly (termed G-tetrads) (Ranjan et al. 2010) .

Previous reports have shown the molecules that

contain both a planar unit and moieties that favor

interaction in the nucleic acid grooves can be used to

target a variety of nucleic acid structures and develop

probes for nucleic acid recognition (Xue et al. 2011;

Watkins et al. 2013; Ranjan et al. 2013). However,

such target validation approaches must be compre-

hensive and easy to perform. In this regard, assays

such as competition dialysis could provide a rapid

approach to determine optimum targets of such new

metabolites (Chaires et al. 2005). Alternate

approaches which do not rely on the intrinsic fluores-

cence of the test metabolites such as fluorescence

intercalator displacement assay, thermal melting of

nucleic acid mixtures, fluorescence quenching assays

could also complement target validation efforts.

Conclusion and perspectives

The findings indicate that species of genus Pestalo-

tiopsis are prolific producers of bioactive metabolites.

They have the potential of producing compounds

which have anticancer, antiviral, antibacterial, anti-

fungal activities and some of them exhibit strong

potential to be developed as a new drug. Efforts are

needed to take these compounds forward for drug

development. These compounds should be evaluated

for their modes of action and toxicity. Since there is a

large chemical diversity, they should be evaluated for

other unscreened targets and modified chemically to

improve their activity. The whole genome sequences

are needed for checking their potential for producing

unreported compounds. Molecular approaches such as

transfer of biosynthetic gene clusters to a vector

suitable for large-scale fermentation could be used.

Liu (2011) reported seventy new natural products from

different biosynthetic routes using, P. fici highlighting

the vast possibility of uncovering several such new

compounds. Pestalotiopsis is reported from various

substrates and environment hence there is a need of

culture collection of specialized cultures of industrial

importance.

Acknowledgement Authors are thankful to Dr. Alok

Adholeya, Senior Director, Biotechnology and Bioresouces

Division, The Energy and Resources Institute, Darbari Seth

Block, IHC Complex, Lodhi Road, New Delhi, India, for help

and continuous support.Ta

ble

2continued

Endophytic

fungal

strain

Hostplant(s)

(fam

ily),

Plantpartortissue/

Locality

ofhost

plants

Isolatedmetabolite

Testedsystem

sActivityresponse

References

Pestalotiopsissp.

RE-1214.

Soilsample

collectedin

Kanagaw

aPref.,Japan

RES-1214-1

(154)and

RES-1214–2(1

55)

ET-1

bindingto

endothelin

typeA

receptor(ETA

receptor)

IC50values

of1.5

lM

and20lM,

respecively

Ogaw

aet

al.(1995)

RES-1214-1

(154)and

RES-1214-2

(155)

Inhibited

theincrease

inintracellularCa2

?

concentration.

elicited

by1nM

ET-

1in

A10cells

Ogaw

aet

al.(1995)

Phytochem Rev

123

Author contributions SKD, VP, and NR reviewed the con-

tents critically. VP and NR drew chemical structures and

assisted in the preparation of Table 2. SKD and NR wrote the

review.

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