Phytomedicine Plus 1 (2021) 100028

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Phytomedicine Plus
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Antioxidant and immunosuppressive activities of extracts of endophytic

fungi isolated from Psidium guajava and Newbouldia laevis
Nonye T. Ujam a,∗, Daniel L. Ajaghaku b, Festus B.C. Okoye c, Charles O. Esimone d
a
Department of Pharmaceutical Microbiology and Biotechnology, Faculty of Pharmaceutical Sciences, Enugu State University of Science and Technology, Enugu, Nigeria
b
Department of Pharmacology and Toxicology, Faculty of Pharmaceutical Sciences, Enugu State University of Science and Technology, Enugu, Nigeria
c
Department of Pharmaceutical and Medicinal Chemistry, Faculty of Pharmaceutical Sciences, Nnamdi Azikiwe University, Awka, Nigeria
d
Department of Pharmaceutical Microbiology and Biotechnology, Faculty of Pharmaceutical Sciences, Nnamdi Azikiwe University, Awka, Nigeria

a r t i c l e i n f o a b s t r a c t

Keywords: Background: Plant endophytic fungi have been recognized as an important and novel resource of natural bioactive
Newbouldia laevis products with potential application in different fields of life.
Psidium guajava
Endophytic fungi Purpose: The present work evaluated the antioxidant and immunosuppressive activities of the extracts of endo-
Bioactive compounds phytic fungi isolated from Psidium guajava and Newbouldia laevis.
Antioxidant
Study design/methods: Endophytic fungi were isolated from the selected plants and their secondary metabolites
Immunosuppressive activity
extracted with ethyl acetate after solid state fermentation on rice media for 21 days at 22°C. Free radical scav-
enging activity (DPPH assay) was used to determine the antioxidant capacity of the extracts and their IC50 were
calculated, using quercetin as the standard. Immunomodulatory activities of the extracts were evaluated using
cyclophosphamide induced myelosuppression model. Bioactive components of the extracts were examined using
High Performance Liquid Chromatography-Diode Array Detector (HPLC-DAD) analysis,
Results: Two endophytic fungi (PGS1 and NLL3) were isolated from P. guajava and N. laevis, respectively. The
fungal extract at 200 and 100 mg/mL significantly (p<0.05) reduced the total white blood cell count of the mice
after ten days treatment. The crude extracts of PGS1 and NLL3 exhibited potent inhibition of DPPH free radical
activity with IC50 values of 44.1 and 41.1 μg/mL, respectively. HPLC-DAD analysis revealed the presence of pro-
tocatechuic acid, asteric acid, citrinin, nidulalin, citreohybridinol, p-hydroxybenzoic acid, cyclopenin, nakijinol
previously reported to have antioxidant and immunosuppressant properties.
Conclusion: Extracts of endophytic fungi isolated from P. guajava and N. laevis possess antioxidant and im-
munosuppressive activities, which could be attributed to the presence of the secondary metabolites identified
by dereplication.

Introduction
Endophytic fungi reside within most tissues of the living plants and
are known to produce different rare and novel secondary metabolites
(Ibrahim et al., 2015). Although many endophytic fungi have been de-
scribed and explored from various terrestrial plants (Strobel and Di-
asy, 2003; Rodriguez et al., 2009; Okoye et al., 2013; Okoye et al., 2015;
Ujam et al., 2019; Ujam et al., 2020) only a limited number have been
studied, among approximately one million species worldwide (Ganley
et al., 2004). Plants, particularly those with medical significance tena-
ciously living under extreme conditions, such as in high-altitude moun-
tainous, oceanic, polar, and glacier areas, may harbor unique and di-
verse endophytic fungi (Yu et al., 2014). Schutz et al. (2001) notes
that certain microbial metabolites seem to be characteristic of certain
biotopes, on both an environmental as well as organismal level.
Bioactive compounds with sundry biological activities have been
isolated from plants and microbes. Protocatechuic acid (3, 4-
dihydroxybenzoic acid), a derivative of p-hydroxybenzoic acid which
can be synthesized chemically as well as isolated naturally1 also ex-
hibit antifungal activity, anti-inflammatory, anti-hepatotoxic, antiox-
idant, free radical scavenger, cytotoxic, chemopreventive, apoptotic,
anti-platelet aggregation, neuroprotective and LDL oxidation inhibitor
activities (Khadem and Marles, 2010; Manuja et al., 2013). Asteric acid
reportedly has antifungal, antibacterial, cytotoxic activity, fungicidal
activity, nematicidal activity (Iwahashi et al., 2007). Citrinin showed

∗
Corresponding author at: Department of Pharmaceutical Microbiology and Biotechnology, Faculty of Pharmaceutical Sciences, Enugu State University of Science
and Technology, P.M.B 01660 Ebeano, Enugu, Nigeria.
E-mail address: nonyetreasure@yahoo.com (N.T. Ujam).

https://doi.org/10.1016/j.phyplu.2021.100028
Received 17 November 2020; Received in revised form 15 January 2021; Accepted 17 January 2021
2667-0313/© 2021 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license
(http://creativecommons.org/licenses/by-nc-nd/4.0/)
N.T. Ujam, D.L. Ajaghaku, F.B.C. Okoye et al.
strong inhibitory action against yeast cells. It also has antimicrobial
activity (Iwahashi et al., 2007). Nidulalin has immunomodulatory and
antitumor activities (Tatsuta et al., 2009). Citreohybridinol has anti-
insectal activities (McMullin, 2014).
Secondary metabolites produced by endophytic fungi have been ex-
ploited in many disease areas. Among such disease, areas that have
shown promise are diseases associated with oxidative stress. Reactive
oxygen species (ROS), such as O2 − , H2 O2 , and OH, are generated
through biological reactions causing oxidative damage to biomolecules
and playing vital roles in programmed cell death and stress-response
signaling in conjunction with antioxidant production (Ravindran et al.,
2012). Under a situation of oxidative stress, reactive oxygen species such
as superoxide (O− 2
) hydroxyl (OH∙ ), and peroxyl (OOH∙ , ROO∙ ) radicals
are generated. These reactive oxygen species play an important role in
degenerative or pathological processes, such as aging, cancer, coronary
heart disease, alzheimer’s disease, atherosclerosis, cataracts, and inflam-
mation (Lawal et al., 2015a). The recent growth in knowledge of free
radicals and ROS in biological systems produces a medical revolution
that promises a new age of health (Tsado et al., 2016). The major roles
of antioxidants are in preventing the oxidation of other molecules by
inhibiting the instigation or promulgation of oxidizing chain reactions
by free radicals (Lawal et al., 2015b). Antioxidants, which inhibit the
oxidation of organic molecules, are very important, not only for food
preservation but also for the defense of living systems against oxidative
stress Masuda et al., 2003; Ullah et al., 2007).
Asymptomatic fungi as mediators can produce antioxidants that can
interrupt the chain reaction of ROS to help host plants respond to vari-
ous biotic and abiotic stresses (White and Torres, 2010; Hamilton et al.,
2012). As a result, some endophytic fungi with scavenging ROS activity
in vitro are isolated from special antioxidant plants (Zhao et al., 2012).
Some of these endophytes have also been shown in our previous stud-
ies to generate novel secondary metabolites with very high antioxidant
properties (Abonyi et al., 2018
A number of immunomodulatory compounds have been isolated
from endophytic fungi (Okoye et al., 2013, Okoye et al., 2015, Ujam
et al., 2019). These immunomodulatory compounds are mainly catego-
rized into immunosuppressive, immunostimulant, and immunoregula-
tory drugs. Immunosuppressive drugs are mainly used to prevent al-
lograft rejection in transplant patients and also to treat autoimmune
diseases such as insulin dependent diabetes and rheumatoid arthritis.
Consequently, researchers have mainly focused on the production of
these drugs from the alternative sources, and one of them could be en-
dophytes. An intensive search is on-going for more effective agents to
deal with immunological disorders related to graft rejection and vari-
ous other autoimmune diseases (Kaul et al., 2012). Hence, in this study,
endophytic fungi isolated from P. guajava and N. laevis were evaluated
for their antioxidant and immunosuppressive activity.
Material and methods
Plant collection: The leaves of N. laevis and stems of P. guajava were
collected from Akegbe-Ugwu, in Nkanu West Local Government Area of
Enugu State, South-Eastern, Nigeria and authenticated by a taxonomist,
Mrs. Anthonia U. Emezie of the Department of Pharmacognosy, Faculty
of Pharmaceutical Sciences, Nnamdi Azikiwe University, Awka, Nige-
ria, and deposited in the herbarium of the above-mentioned department
(With voucher specimen numbers PCG474/A/035 and PCG474/A/045
for N. laevis and P. guajava, respectively).
Experimental design
Animals
Swiss albino mice of either gender, weighing 25-30 g housed in stan-
dard conditions of temperature, humidity, and light, were used. They
were fed with a standard rodent diet and water ad libitum. All labora-
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Phytomedicine Plus 1 (2021) 100028
tory animals were treated according to the international regulations for
the usage and welfare of laboratory animals.
Standard drugs
Cyclophosphamide (500 mg) Cycloxan® (Biochem–Pharmaceutical
Industries Ltd., Mumbai) was used as a standard immunosuppressant.
NONI, a standard immunomodulatory drug. Dilutions were made using
sterile water for injection following manufacturer’s guidelines.
Isolation of endophytic fungi, fermentation, and extraction of the secondary
metabolites
Isolation of endophytic fungi from healthy leaves and stems of N. lae-
vis and P. guajava was carried out using methods previously reported by
Ujam et al. (2019) with modifications. The isolated pure fungal strains
were stored in malt extract agar (MEA). Solid-state fermentation was
carried out in 1 L Erlenmeyer flasks containing autoclaved rice medium
(100 g of rice and 200 mL of distilled water). Agar blocks containing
pure cultures of individual fungus were cut from MEA plates and inocu-
lated into the fermentation flasks. The inoculated flasks were incubated
at 28 ± 1°C for 28 days. The fungal secondary metabolites were ex-
tracted with ethyl acetate after the fermentation and concentrated with
a rotary evaporator at 40°C.
Identification of the isolated endophytic fungi
The isolated fungi were identified on the basis of the appearance
of their colonial morphology (cultural characteristics) and microscopic
examinations by staining with lactophenol blue and observing under a
photomicroscope. All the fungal isolates were maintained in test tubes
and Petri plates on SDA media.
Determination of antioxidant (scavenging) and the IC50s of the extracts of
the endophytic fungi using DPPH assay
The free radical scavenging activities of the ethyl acetate extracts of
the endophytic fungal isolates were evaluated by 1, 1-diphenyl-2-picryl-
hydrazyl (DPPH) free radical scavenging assay following previously re-
ported method (Shen et al., 2010). The IC50 (the concentration showing
50% inhibition) values were calculated.
Evaluation of immunomodulatory activity of the extracts of the isolated
endophytic fungi
The immunomodulatory potential was investigated using Cyclophos-
phamide (CA) -induced myelosuppression as previously reported Ujam
et al. (2019) with some modifications. The curative (treatment) and pro-
phylactic (preventive) experiments were carried out.
High-Performance Liquid Chromatography-Diode-Array Detection
(HPLC-DAD) assay
HPLC analysis of the endophyte extract was carried out using Dionex
attached to a photodiode array detector (UVD340S, DionexSoftron
GmbH, Germering, Germany) as previously described (Akpotu et al.,
2017). Two milligram (2 mg) of the extract was reconstituted with 2
mL of HPLC grade methanol, the mixture sonicated for 10 min and cen-
trifuged at 3000 rpm for 5 min. Afterward 100 𝜇L of the dissolved sam-
ples were transferred into HPLC vials containing 500 𝜇L of the HPLC
grade methanol for analysis. Detection was at 235, 254, 280, and 340
nm. The separation column (125 mm × 4 mm; length × internal di-
ameter) was pre-filled with Eurospher-10 C18 (Knauer, Germany) and
a linear gradient of nanopure water (adjusted to pH 2 by addition of
formic acid) and methanol was used as eluent. The absorption peaks for
each of the 10 dried fungal metabolite extract were analyzed by com-
paring with those in the HPLC-ultraviolet (UV)/visible database, which
contained scores of registered compounds.
N.T. Ujam, D.L. Ajaghaku, F.B.C. Okoye et al.
Fig. 2. The macroscopic (PGS1a and NLL3a) and microscopic (PGS1b and NLL3b) features of the Endophytic fungal isolates of P. guajava (1) and N. laevis (2),
respectively.
Table 1
Percentage (%) free-radical scavenging activity (DPPH assay).
S/N Extract code Conc. of extracts Mean Absorbance
1 PGS1 500𝜇g/mL 0.595
2 NLL3 500𝜇g/mL 0.462
3 Quercetin 500𝜇g/mL 0.009
Abs=Absorbance, Conc. = Concentration.
Statistical analysis
Data were presented as mean ± standard error of mean (SEM) of sam-
ple replicate, n=5. The analysis was done using statistical package for
social Sciences (SPSS) version 20 for windows. Statistical significance
was established when P<0.05. Graphical illustration was carried out us-
ing Microsoft excel, 2007.
Results
Endophytic fungi coded PGS1 and NLL3 were respectively isolated
from the stems and leaves of N. laevis and P. guajava respectively. P.
guajava and N. laevis (Fig. 1), and identified based on their colonial mor-
phology and microscopic appearances as Fusarium sp. and Cladosporium
sp. (Fig. 2). Their colonial characteristics and microscopic features are
shown in Fig. 2.
The results of the pilot study of the antioxidant activity extracts of
the fungal endophyte and the standard antioxidant agent quercetin are
shown in Table 1. A 500 μg/mL solution of PGS1 and NLL3 showed
69.84 and 76.58% inhibition, respectively compared to 99.5% inhibition
of quercetin at the same concentration. Extracts of PGS1 and NLL3 gave
IC50 of values of 44.07 and 41.14 μg/mL, respectively (Table 1).
The mean basal total white blood cell (TWBC) count of the mice
ranged from 4.20 - 6.00 × 103 / mm.3 Following administration of cy-
clophosphamide, the TWBC count decreased significantly p<0.005 rang-
ing from 1.40 to 2.60 × 103 / mm3 (Table 2). After five days of ad-
ministration of PGS1 and NLL3 extracts to the mice, rather than in-
crease the mean TWBC count, decreased ranged from 1.35 to 2.10 × 103
/mm3 . Further decrease of the TWBC counts was observed at the end ten
days administration of the extracts; mean TWBC count was from 1.30
to 2.12 × 103 /mm3 (Table 2).
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Phytomedicine Plus 1 (2021) 100028
Fig. 1. The studied plants: (a) N. laevis and (b) P. guajava.
% Inhibition of DPPH scavenging activity IC50 (mg/mL)
69.84 44.07
76.58 41.14
99.50 14.52
Comparing the effects of fungal extracts on the TWBC level of the
mice after five days of treatment with TWBC after suppression showed
a significant (p< 0.005) increase of the WBC by the positive control
while the extracts had no significant increase (p > 0.05), even after ten
days treatment. A higher dose (200 mg/kg) of the extracts decreased
the TWBC of the mice more compared to that of the lower dose (100
mg/kg).
The effects of PGS1 and NLL3 extracts on the differential leukocyte
(neutrophil) count are shown in Table 3. Basal neutrophil count ranged
from 20.00 to 23.00 × 103 , after suppression with CA the count sig-
nificantly (p<0.01 and p<0.005, respectively) decreased (8.67 to 13
.00 × 103 ). After 10 days of treatment, extracts of PGS1 and NLL3 fur-
ther suppressed the neutrophil count (8.21 – 12.00 × 103 ), comparable
to the effect of CA. On the other hand, the immunostimulant drug NONI
significantly increased the neutrophil count (p<0.001).
The result of the prophylactic study showed that pretreatment of the
mice with PGS1 and NLL3 extracts could not prevent cyclophosphamide
immunosuppression in the mice (Table 4). Percentage CA suppression
of TWBC inhibition by PGS1 and NLL3 extracts were 22% and 8% inhi-
bition, respectively (Table 4).
The extracts of extracts of Fusarium sp. (PGS1) and Cladosporium
sp. (NLL3) from P. guajava and N. laevis respectively represent a reli-
able source of bioactive compounds demonstrated by the wide range
of compounds with diverse biological properties present in these ex-
tracts. The HPLC-DAD analysis of the extracts revealed the presence of
protocatechic acid, p-hydroxybenzoic acid, nakijimol, chloramphenicol,
orthosporin, chloramphenicol, citreohybridinol, nidulalin and asterric
acid (Table 5).The HPLC chromatograms and UV-spectra of detected
compounds are presented in Figs. 3–6.
N.T. Ujam, D.L. Ajaghaku, F.B.C. Okoye et al. Phytomedicine Plus 1 (2021) 100028

Table 2
Mean total white blood cell counts (thousand /mm3 ) for the curative experiment.

Treatment groups Doses (mg/kg) Basal Ẋ +SEM After suppression Ẋ +SEM After 5 days treatment Ẋ +SEM After 10 days treatment Ẋ +SEM

Cyclophosphamide PGS1 200 4.20 ± 0.12bbb 1.50 ± 0.13 1.35 ± 0.07 1.30 ± 0.00
(30 mg/kg) 100 5.30 ± 0.24bbb 1.80 ± 0.08 1.70 ± 0.25 1.70 ± 0.17
NLL3 200 6.00 ± 0.30bbb 2.60 ± 0.20 1.99 ± 0.14 1.40 ± 0.16
200 4.20 ± 0.08bbb 2.20 ± 0.10 2.10 ± 0.08 2.12 ± 0.11
NONI 100 5.60 ± 0.14bbb 1.40 ±0.08 4.60 ± 0.20bbb 5.40 ± 0.34bbb
DW 10 ml/kg 4.50 ± 0.08bbb 2.10 ± 0.07 2.30 ± 0.12 2.43 ± 0.12

Values are presented as mean ± Standard Error of Mean (SEM) of five (5) replicates (n=5). b P<0.05, bb
P<0.01 and bbb
P<0.005 Significantly different from
WBC level at suppression. NONI =Positive control, DW (Distilled water) = Negative control.

Table 3
Mean neutrophil count (thousand /mm3 ) for the curative experiment.

Treatment Groups Doses (mg/kg) Basal Ẋ +SEM After suppression Ẋ + SEM After 5 days treatment Ẋ + SEM After 10 days treatment Ẋ + SEM
bb
Cyclophosphamide PGS1 200 23.00 ± 0.84 13.00 ± 1.64 11.00 ± 1.41 10.56 ± 0.71
(30 mg/kg) extract 100 23.00 ± 1.82bb 12.00 ± 0.71 12.00 ± 1.22 12.00 ± 1.22
NLL3 200 20.00 ± 1.00bbb 8.67 ± 0.71 8.00 ± 1.38 8.21.00 ± 1.38
extract 100 21.00 ± 0.95bbb 11.00 ± 0.95 12.00 ± 1.38 13.00 ± 1.22
NONI 10 ml/kg 22.00 ± 1.00bbb 9.40 ± 1.03 18.51 ± 1.05bbb 22.00 ± 1.05bbb
Distilled 10 ml/kg 20.00 ± 3.08bb 11.00 ± 1.00 12.00 ± 1.00 12.00 ± 1.00
water
Values are presented as mean (Ẋ) ± Standard Error of Mean (SEM) of five (5) replicates (n=5). Basal neutrophil significantly different b P<0.05.
bb
P<0.01 and bbb P<0.005 compared to neutrophil level at suppression and at the end of 10 days treatment. NONI= Standard immunomodulatory drug.

Table 4
Mean total white blood cells counts (thousand/mm3 ) for the prophylactic experiment/CA suppression inhibition.

After 10 days After suppression % CA Suppression

Treatment groups Doses Basal Ẋ ± SEM treatment Ẋ ± SEM Ẋ ± SEM Inhibition

PGS1 extract 200 mg/kg 7.10 ± 0.10 7.10 ± 0.07 3.30 ± 0.12 bbb 22
CA (50mg/kg) NLL3 extract 200 mg/kg 6.80 ± 0.07 7.90 ± 0.03 4.40 ± 0.13bbb 8
Distilled water 10 ml/kg 6.30 ± 0.13 8.30 ± 0.18 3.42 ± 0.16bbb 0

Values were presented as mean ± Standard error of mean (SEM) of five (5) replicates (n=5) Significantly, different from basal
WBC level. b P<0.05, bb P<0.01 and bbb P<0.005 Significantly different from WBC level on day 10. CA=Cyclophosphamide, Distilled
water = Negative control.

Table 5
Bioactive compounds detected in endophytic fungal extracts by HPLC-DAD analysis.

Fungal extracts Detected compounds

PGS1 Protocatechic acid, p-Hydroxybenzoic acid, Nakijimol B, Chloramphenicol, Orthosporin, Chloramphenicol

NLL3 Protocatechuic acid, Citreohybridinol, Nidulalin, Asterric acid

HPLC-DAD: High-performance liquid chromatography-diode-array detection.

Fig. 3. HPLC-DAD Chromatogram of the isolated Fusarium sp. (PGS1) extract showing detection of bioactive compounds A, B, C, D, E and F.

4
N.T. Ujam, D.L. Ajaghaku, F.B.C. Okoye et al. Phytomedicine Plus 1 (2021) 100028

Fig. 4. Ultraviolet(UV) spectra of compounds detected from Fusarium sp. (PGS1) extract.
A:Protocatechic acid, B: p-Hydroxybenzoic acid, C:Nakijimol, D: Chloramphenicol, E: Orthosporin and F:Chloramphenicol.

Fig. 5. HPLC-DAD chromatogram of extracts of the endophytic fungus Cladosprium sp. (NLL3) isolated from Newbouldia laevis.

5
N.T. Ujam, D.L. Ajaghaku, F.B.C. Okoye et al.
Fig. 6. Ultraviolet (UV) spectra of compounds detected from Cladosporium sp. (NLL3) extract.
A: Protocatechuic acid, B: Citreohybridinol, C: Nidulalin and D: Asterric acid.
Discussion
Accumulated evidence has suggested that most of the degenera-
tive diseases that afflict humanity have their origin in deleterious free
radical reaction (Florence, 1995; Khan et al., 2018). Hence, antiox-
idants find wide applications in the management of these disorders
in which free radicals are implicated. Plants have served as a source
of medicinal bioactive compounds against oxidative stress. In recent
years, microorganisms associated with plants rather than plants them-
selves have proved to be a potential source of bioactive compounds
with antioxidant activity that are being explored in drug discovery
(Gouda et al., 2016). In this study, endophytic fungi PGS1 and NLL3
from the stems and leaves of Psidium guajava and Newbouldia laevis, re-
spectively were shown to possess antioxidant activities. These activi-
ties should be attributed to the bioactive compounds present in their
metabolites.
Mounting evidence from both in vitro and in vivo studies demon-
strates that protocatechuic acid exerts potent antioxidant effects
(Semaming et al., 2015). Consistent with these reports, this study also
demonstrated that PSG1 and NLL3 metabolites containing these com-
pounds produced inhibition of free radical formation as measured by
the DPPH test. Other bioactive compounds like p-hydroxy benzoic acid,
which have also been reported to possess antioxidant activities may have
also contributed to the free radical scavenging activity of NLL3 metabo-
lite (Manuja et al., 2013).
Antioxidant compounds have also been suggested through numerous
scientific researches to improve a range of immune responses, support-
ing both innate and acquired immunity while also preserving the struc-
tural integrity of immune cells (Carr and Maggini, 2017). On the other
hand, many useful roles of free radicals and reactive oxygen species
(ROS) in the immune system have also been documented. These useful
roles include mediating key mechanisms in phagocyte recruitment, ad-
hesion, activation and phagocytosis (Halliwell and Gutteridge, 2007).
ROS have also been reported to mediate other signaling pathways in-
volved in immune response (Yarosz et al., 2018). Based on these avail-
able evidences, antioxidants may serve as a double-edged sword in im-
mune function and responses which may explain the antioxidant and
6
Phytomedicine Plus 1 (2021) 100028
immunosuppressive relationship exhibited by fungi metabolites of PGS1
and NLL3.
Since white blood cells, particularly neutrophils are key compo-
nents of inflammatory mediators that shape the immune responses
(Mócsai, 2013), the suppressive effect of PGS1 and NLL3 on these cell
types may also be attributed to the anti-inflammatory activity of their
bioactive compounds. Immunosuppressive activity of protocatechuic
acid on cell mediated immunity has been documented and attributed to
its anti-inflammatory activity (Rasne et al., 2018). Since this compound
is present in both PGS1 and NLL3, it may have accounted for both the an-
tioxidant and immunosuppressive activities of the fungi metabolites. Al-
ternatively, the presence of other non-antioxidant bioactive compounds
in the crude extracts that may suppress immune activity may also ac-
count for the immunosuppressive effect observed. Chloramphenicol, for
instance, was among the compounds detected in NLL3, which has been
reported to be associated with bone marrow depression resulting in the
reduction of hematological indices, including white blood cells (Skukla
et al., 2011).
These findings suggest that the metabolites of PGS1 and NLL3 could
be used as a complementary and alternative therapy in various chronic
degenerative diseases in which oxidative damage and inflammatory pro-
cesses play important roles in their pathogenesis and/or disease progres-
sion.
Conclusion
The present study reports that extracts of PGS1 and NLL3 isolated
from P. guajava and N. laevis respectively exhibited antioxidant and im-
munosuppressive activities, which could be attributed to the bioactive
compounds therein. Additional investigations on each of these com-
pounds are required to further elucidate their antioxidant and immuno-
suppressive mechanisms.
Authors’ contributions
This work was carried out in collaboration between all authors. COE
and FBCO designed and supervised the study. NTU managed the labo-
N.T. Ujam, D.L. Ajaghaku, F.B.C. Okoye et al.
ratory analyses and prepared the first draft of the manuscript. NTU and
DLA managed the data analysis and literature searches. All authors read
the final version and confirmed it for publication.
Funding/support
The study was self-sponsored. No grant was secured.
Declaration of Competing Interest
The authors declare no conflict of interest.
Acknowledgment
The authors thank the authorities of Nnamdi Azikiwe University,
Awka, Anambra Nigeria for the facilities. We are grateful Prof. Dr. Peter
Proksch and other members of staff of the Institute of Pharmaceutical
Biology and Biotechnology, Heinrich-Heine University, Düsseldorf, Ger-
many for allowing a part of my work to be done at the institution and
for their assistance and guidance all through the period.
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