Lawsozaheer, a new chromone produced by an endophytic fungus Paecilomyces variotii isolated from Lawsonia Alba Lam. inhibits the growth of Staphylococcus aureus

Article abstract of DOI:10.1080/14786419.2020.1729148

One new chromone, lawsozaheer (1), and five known compounds 4-(2-hydroxyethyl) phenol (2), viriditoxin (3), stigmasta-4,6,8(14),22-tetraen-3-one (4), β-sitosterol (5) and stigmasterol (6) were isolated from the fungal broth of Paecilomyces variotii. Their

Full text of DOI:10.1080/14786419.2020.1729148

Natural Product Research
Formerly Natural Product Letters
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Lawsozaheer, a new chromone produced by an
endophytic fungus Paecilomyces variotii isolated
from Lawsonia Alba Lam. inhibits the growth of
Staphylococcus aureus
Zaheer Abbas, Bina Shaheen Siddiqui, Saleem Shahzad, Samia Sattar, Sabira
Begum, Anum Batool & Mohammad Iqbal Choudhary
To cite this article: Zaheer Abbas, Bina Shaheen Siddiqui, Saleem Shahzad, Samia Sattar,
Sabira Begum, Anum Batool & Mohammad Iqbal Choudhary (2020): Lawsozaheer, a new
chromone produced by an endophytic fungus Paecilomycesꢀvariotii isolated from Lawsonia
Alba Lam. inhibits the growth of Staphylococcusꢀaureus, Natural Product Research, DOI:
10.1080/14786419.2020.1729148
To link to this article: https://doi.org/10.1080/14786419.2020.1729148
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NATURAL PRODUCT RESEARCH
https://doi.org/10.1080/14786419.2020.1729148
Lawsozaheer, a new chromone produced by an
endophytic fungus Paecilomyces variotii isolated from
Lawsonia Alba Lam. inhibits the growth of
Staphylococcus aureus
Zaheer Abbasa, Bina Shaheen Siddiquia, Saleem Shahzadb, Samia Sattarb,
Sabira Begumc, Anum Batoola and Mohammad Iqbal Choudharya,c
aHEJ Research Institute of Chemistry, International Center for Chemical and Biological Sciences,
b
University of Karachi, Karachi, Pakistan; Department of Agriculture & Agribusiness Management,
c
University of Karachi, Karachi, Pakistan; Department of biochemistry, faculty of science, Icing
Abdnlaziz university, Jeddah, Saudia Arabia
ABSTRACT
ARTICLE HISTORY
Received 10 July 2019
Accepted 9 February 2020
One new chromone, lawsozaheer (1), and five known compounds
4-(2-hydroxyethyl) phenol (2), viriditoxin (3), stigmasta-
4,6,8(14),22-tetraen-3-one (4), b-sitosterol (5) and stigmasterol (6)
were isolated from the fungal broth of Paecilomyces variotii. Their
structures were elucidated using spectroscopic data. The configur-
ation of 1 was determined by Horeau’s method. The broth extract
KEYWORDS
Lawsonia alba Lam;
Paecilomyces variotii;
antimicrobial activity;
Staphylococcus aureus;
Candida albicans
and compound
1 showed highly selective activity against
Staphylococcus aureus (NCTC 6571) bacterium with 83.19 and
84.26% inhibition respectively at 150mg/mL, comparing well with
that of standard drug ofloxacin (87.013% inhibition at 100mg/mL).
Broth extract also showed 75, and 40% inhibition of Candida albi-
cans and Fusarium lini, respectively.
1. Introduction
Lawsonia (family Lythraceae) is a monotypic genus characterized by Lawsonia inermis
Linn. (syn. L. alba Lam.). It is known as henna in Arabic, mehndi in Hindi and Urdu. It
is a native of North Africa and South-West Asia, and partially cultivated in West Africa
(Sastri 1962; Zafar et al. 2006). The natural products isolated from this plant include
lawsone (Cox 1938), aromatic compounds (Bhardwaj et al. 1976; Siddiqui et al. 2003;
Uddin et al. 2011), saponins, triterpenoids (Khan et al. 1996; Siddiqui et al. 2005) and
CONTACT Bina Shaheen Siddiqui
siddiqui_bina@yahoo.com
Supplemental data for this article can be accessed at https://doi.org/10.1080/14786419.2020.1729148.
ß 2020 Informa UK Limited, trading as Taylor & Francis Group

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Z. ABBAS ET AL.
dioxin derivatives (Siddiqui et al. 2009). Previous investigations revealed that L. alba
exhibits a wide range of biological activities, such as antibacterial (Habbal et al. 2011),
antifungal (Begum et al. 2007), antiviral (Chaudhary et al. 2010), anticancer (Priya et al.
2011; Pradhan et al. 2012; Singh and Luqman 2014), hepatoprotective (Latha et al.
2005) and enzyme inhibition (Uddin et al. 2013) activity.
Endophytic microorganisms are fungi or bacteria which animate inside the plant tis-
sues during their life cycle at any moment, without causing damage to their host.
Endophytic fungi are significant source of a number of novel bioactive secondary
metabolites, possibly valuable as human medicines, possessing antimicrobial, anti-
cancer and several other activities (Amirita et al. 2012; Mousa and Raizada 2013) and
could also serve as a prospective source of agrochemical and industrial products
(Souza et al. 2011). Previously a few studies on Lawsonia alba endophytic secondary
metabolites have been undertaken (Sarang et al. 2017) but there is no record on the
chemical constituents of secondary metabolites of the fungus Paecilomyces variotii
from Lawsonia alba. Therefore present studies were undertaken on the fungal broth
extract from this source. Separation of broth extract using ethyl acetate extraction and
various chromatographic techniques led to isolation of one new ((lawsozaheer, 1) and
five known compounds. The broth extract and compound 1 showed pronounced
activity against Gram-positive bacterium Staphylococcus aureus (NCTC 6571; and mod-
erate activity against two fungi Candida albicans and Fusarium lini.
2. Results and discussion
One new chromone, lawsozaheer (1) and five known compounds 4-(2-hydroxyethyl)
phenol (2) (Kimura and Tamura 1973), viriditoxin (3) (Silva et al. 2013), stigmasta-
4,6,8(14),22-tetraen-3-one (4) (Kobayashi et al. 1992), b-sitosterol (5) and stigmasterol
(6) (Kamboj and Saluja 2011) were isolated in the present studies (Figure 1). The broth
extract and compound 1 were evaluated for their antimicrobial activity. It may be
noted that viriditoxin (2) has earlier been shown to possess broad spectrum antibac-
terial activity against clinically relevant Gram-positive pathogens including methicillin-
resistant S. aureus (Silva et al. 2013). However this is a new source of its isolation.
2.1. Structure elucidation of lawsozaheer (1)
1 was isolated as white powder; [a] 27: ꢀ 43.89 (c, 0.09, MeOH). Its IR spectrum (KBr)
D
showed characteristic bands at 3421, 3376 (OH) and 1614, 1662 (sh.), (C ¼ O) cmꢀ1
and UV spectrum (MeOH) showed kmax at 243, 248 and 260 nm. Its EI-MS displayed
molecular ion peak at m/z 234 ((Figure S1) and peak matching of the molecular ion
gave exact mass m/z 234.0892 corresponding to the molecular formula C13H14O4 (calc.
234.08913) displaying seven unsaturations. A significant peak at m/z 190 was due to
1
loss of ethyl alcohol. A benzopyrone ring was deduced from analysis of the H- and
13C-NMR data (Supplementary material, Table S1; Figures S2-S7) which showed two
meta coupled aromatic protons at d 6.63 (1 H, d, J ¼ 2.4, H-6) and 6.65 (1 H, d, J ¼ 2.4,
H-8) showing connectivity to aromatic carbons at dC 118.0 (C-6) and 101.7 (C-8) in the
HSQC spectrum (Supplementary material, Figures S6); an olefinic proton singlet at d

NATURAL PRODUCT RESEARCH
3
Figure 1. Structures of compounds 1–6.
6.05 for H-3 connected to olefinic carbon at d 112.4 (C-3); a carbonyl carbon at dC
182.0 (C-4), a quaternary carbon at d 167.1 (C-2). The 13C-NMR spectrum further
showed four quaternary aromatic carbons at dC 163.2 (C-7), 143.6 (C-5), 161.5 (C-9)
and 115.7 (C-10). The attached hydroxypropyl chain was recognized from proton sig-
nals at dH 4.19 (1 H, m, H-2’), 1.27 (3 H, d, J ¼ 6.0, H-3’) and two diastereotropic protons
at dH 2.64 (1 H, dd, J ¼ 14.4, 7.8, H-1’a) and 2.70 (1 H, dd, J ¼ 14.4, 6.8, H-1’b). Their
respective carbons were identified at dC 66.3 (C-2’), 23.5 (C-3 ’) and 44.2 (C-1’) in the
HSQC spectrum (Supplementary material, Figures S6). Signals for an aromatic CH3
group were noted at dH 2.71 (3 H, s)/dC 23.2. The types of carbons were identified
from broad band (Figure S3), DEPT 90 (Supplementary material, Figure S4) and DEPT
135 (Supplementary material, Figure S5) spectra and the protonated carbons were
identified with the help of HSQC spectrum. The placement of various groups was con-
firmed from HMBC NMR spectrum (Supplementary material, Figure S7). Thus interac-
tions were observed between H-3 (d 6.05) and C-2 (d 167.1), C-1’ (d 44.2) and C-10 (d
115.7); H-1’b to C-2, H-1’a (d 2.64) and C-3 (d 112.4), C-3’ (d 23.5); H-3’ (d 1.27) and C-1’
(d 44.2), C-2’ (d 66.3); H-11 (d 2.71) and C-6 (d 118.0), C-7 (d 163.2) and C-10 (d 115.7);
H-8 (d 6.65) and C-9 (d 161.5) (Supplementary material, Figure S8). The stereochemistry
at C-2’ was determined by Horeau’s method (Horeau and Kagan 1964; vide
Supplemental). 1 (4.5 mg) was acylated with racemic phenyl-a-butyric anhydride
(20 mg) in dry pyridine. Ice cold water was added to the mixture to decompose excess
anhydride and the mixture was extracted with ethyl acetate which was again extracted
with 5% sodium bicarbonate. The alkaline layer was acidified with 10% hydrochloric
acid and extracted with chloroform. The chloroform layer was worked up in the usual

4
Z. ABBAS ET AL.
27
manner to yield free phenyl-a-butyric acid [a]D ¼ þ0.38 (MeOH, c ¼ 3.5) which led
to determination of configuration at C-2’ as R. In the light of above data, the com-
pound was identified as 2- (2’R-hydroxypropyl) ꢀ5-methyl-7-hydroxychromone (Figure
1). It is a new 2’R isomer of earlier reported 2- (2’S-hydroxypropyl) ꢀ5-methyl-7-
hydroxychromone (Kashiwada et al. 1984).
Compounds 2-6 were identified through comparison of their spectral data with
those reported in literature (loc.cit).
2.2. Antimicrobial activity
Initial antimicrobial activities were determined by Micro plate Alamar Blue Assay
(MABA). Antimicrobial activity of the broth extract was determined against 15 Gram
–positive bacteria including multi drug resistant strains Staphylococcus aureus,
Streptococcus pyogenes, Streptococcus fecalis, Bacillus cereus, Bacillus subtilis, Bacillus
thuringiensis, Micrococcus luteus, Corynebacterium xerosis, Corynebacterium hoffmanii,
Streptococcus pneumonia, Staphylococcus aureus clinical isolate, Staphylococcus aureus
13277, Staphylococcus aureus 13143, Staphylococcus aureus EMRSA-17, and
Staphylococcus aureus VRSA 700699; twelve Gram-negative bacteria Shigella boydii,
Salmonella typhi, Salmonella typhi para A, Salmonella typhi para B, Shigella flexneri,
Proteus mirabilus, Proteus vulgaris, Escherichia coli, Klebsiella pneumonia, Shigella dysen-
teriae, Enterobacter, and Pseudomonas aeruginosa PA0286 and thirteen fungi
Trichopyton rubrum, Microsporum canis, Trichopyton mentegrophyte, Microsporum gyp-
sium, Trichopyton tonsurans, Saccharomyces cerevisiae, Candida albicans, Helementho
sporain, Aspergillus flavus, Aspergillus niger, Penicillium sp, Rhizopus sp., and
Fusarium sp.
The broth extract (LA-B-EA) and new compound (1) showed highly significant activ-
ity against Staphylococcus aureus (NCTC 6571) with 83.19 and 84.26% inhibition
respectively at 150 mg/mL. The standard drug used was ofloxacin which showed
87.013% inhibition at the concentration of 100 mg/mL (Supplementary material, Table
S2). Since compound 1 showed significant activity at 150 mg/mL it was evaluated at
further increasing concentrations in order to determine the MIC90 via MABA which
was found to be 225 lg/mL (Supplementary material, Table S3).
Compound 1 did not show any activity against other strains of S. aureus although
the broth extract (LA-B-EA) showed moderate activity against S. aureus clinical isolate
(39.76%) and S. aureus VRSA 700699 ((37.81%) (Supplementary material, Table S2).
The broth extract showed moderate activity against two fungi Candida albicans
(75% inhibition) and Fusarium lini (40% inhibition) at 400 mg/mL (Supplementary
material, Table S4). The activity of the extract and compound 1 against rest of the bac-
teria and fungi was marginal and not shown in the table.
3. Conclusions
The broth extract (LA-B-EA) and new compound (1) showed highly selective activity
against Staphylococcus aureus (NCTC 6571) bacterium with 83.19 and 84.26% inhibition
respectively at 150 mg/mL, comparing well with that of standard drug ofloxacin

NATURAL PRODUCT RESEARCH
5
(87.013% inhibition at 100 mg/mL). Broth extract also showed 75 and 40% inhibition of
Candida albicans and Fusarium lini, respectively. The discovery provides a direction for
further studies to obtain and develop potent antimicrobial agents from endophytic
metabolites of Lawsonia alba Lam.
Disclosure statement
The authors confirm that this article content has no conflict of interest.
Funding
This work was supported by grants from the Pakistan Academy of Science by the grant # 5-9/
PAS/52.
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