Plant Archives Volume 20 No. 2, 2020 pp.

9253-9264 e-ISSN:2581-6063 (online), ISSN:0972-5210

ANTIMICROBIAL EFFECT OF ASPARAGUS OFFICINALIS L.

EXTRACTS

S.F. Desoukey1, Shereen E.M. El-Nahas2, Atef Z. Sabh1, Zeinab K. Taha1

and Hattem M. El-Shabrawi3
1
Agricultural Botany Dept., Faculty of Agriculture, Cairo University, Giza, Egypt.
2
Plant Pathology Research Institute, Agricultural Research Center, Giza, Egypt.
3
Plant Biotechnology Dept., National Research Center, Giza, Egypt.

Abstract
This experiment was carried out in the tissue culture laboratory, Agricultural Botany Dept., Faculty of Agriculture, Cairo
University, Giza, Egypt and Plant Pathology Research Institute, Agricultural Research Center, Giza, Egypt, during the two
years 2018 and 2019 with the aim of assessing the antimicrobial effect of different biomass extracts of Asparagus officinalis
L. plant (Mary Washington 500 w cultivar) from family Asparagaceae such as callus, fresh shoot, fresh root, dry shoot and
dry root using different solvents (methanol, ethyl acetate, acetone, chloroform and petroleum ether) and estimating the
inhibition zones. The antifungal activity of the different extracts has been verified in vitro by antifungal assays against five
phytopathogenic fungi (Alternaria tenuissima, Botrytis cinerea, Fusarium oxysporum, Macrophomina phaseolina and
Rhizoctonia solani). The results showed that the inhibition percent of mycelial growth increased with increasing concentration
of dry biomass extracts for all fungi used except Botrytis cinerea. Macrophomina phaseolina was less affected by different
biomass extracts compared with all fungi used. The results indicated that dry biomass with ethyl acetate extracts showed
better inhibition percent against all fungi used at most concentrations. Regarding the antibacterial activity, ethyl acetate
extract was superior in inhibition of Gram-positive bacterial strains of all biomass types used. While dry shoot and root
methanol extracts as well as fresh shoot and callus ethyl acetate extracts were the most inhibiting of Gram-negative bacteria
strains.
Key Words: Asparagus officinalis L., plant extracts, antifungal activity, antibacterial activity, phytopathogenic fungi.

Introduction alternatives such as fungicides of fungal infections is very

Asparagus (Asparagusofficinalis L.) is one of the important to enhance resistance to common antifungal
promising nontraditional horticultural crops in Egypt. agents. Natural chemical strategies for controlling crop
However, it’s considered one of the most important diseases are of considerable interest because of
vegetable crops in some Asian, African, European and environmental and health concerns about the widespread
American countries (Hassan, 2001). Pathogenic fungi use of chemical pesticides (Brauer et al., 2019 and Zhang,
are one of the vital problems of crops (Xie et al., 2017 et al., 2020). The potential of secondary metabolites to
and Jain et al., 2019). Fungal pathogens are proved to be protect plants can be used in a more modern and
a common and popular contaminant of agro-ecosystem environmentally friendly alternative strategy (Pino et al.,
that approximately causes 70–80% of total microbial crop 2013 and Marutescu et al., 2017). Shrestha et al., (2018)
loss (Moore et al., 2000 and Santra and Banerjee 2020). stated that phytochemical screening revealed the presence
Common chemical fungicides have long been used to of coumarin, flavonoid, catecholic tannin and reducing
control fungal diseases. These chemical fungicides compounds in the alcoholic extract of A. racemosus.
increase the risk of environmental pollution and adverse Çoban et al., (2009) observed that the ether extract of
effects on biodiversity (Aktar et al., 2009 and Queyrel et A. officinalis L., had an antimicrobial effect against ten
al., 2016). Therefore, the search for new, more effective pathogen bacteria and five yeasts. Patel and Patel (2013)
and environmentally friendly approaches and less toxic and Kishor et al., (2019) pointed out that Asparagus
9254 S.F. Desoukey et al.

racemosus have potent antimicrobial activity against equilibrated. Then grown in nutrient agar, the test bacteria
Gram-positive and Gram- negative bacteria. (0.1 ml) were streaked on nutrient medium plates using a
sterile cotton swab. Agar was allowed to holes numbers
Materials and methods were cut using a sterile cork borer and distribution of
Collection of plant material holes in petri dishes. In each hole, the different extract
In vivo plant material of A. officinalis L. (Mary was loaded. Then, the petri dishes were left at room
Washington 500 w cultivar) was collected from the temperature for 2 h to allow diffusion of the test sample.
experimental field in Fac. Agri., Cairo University, Giza, Antibacterial activities of plant extracts were evaluated
Egypt, in the growing season, 2018, as well as in vitro using a well diffusion method on nutrient agar (Das, et
plant material of the same species was collected from al., 2010). The inhibition zones were reported in millimeter
Plant Tissue Culture Laboratory, Agricultural Botany (mm). R. solanacearum (-ve) and B. subtilis (+ve) were
Department, Faculty of Agriculture, Cairo University, used as references for the antibacterial assay. Briefly,
Giza, Egypt. nutrient agar plates were inoculated with bacterial strain
under aseptic conditions and wells (diameter=6mm) were
Preparation of plant material
filled with 25 µl or 50 µl of the test samples and incubated
• The plant samples (in vivo and in vitro grown plants at 30°C for 24–72 hours. After the incubation period, the
as well as callus tissues) were thoroughly washed well diameters of growth inhibition zones were measured and
and weighed. at three replicates for each test.
• The plant samples (in vivo grown plants) were Statistical analysis
dried completely at 50-60°C for 48 h in a hot air oven
Data was subjected to appropriate statistical and
then crushed using a mechanical grinder.
conventional methods of analysis of variance according
Preparation of plant extracts to Snedecor and Cochran 1989. The mean differences
The plant material (15g) was soaked in 200 ml of the were compared by least significant difference test (LSD)
solvent at room temperature in the dark for one week at P  0.05.
before being filtered. Plant material extraction (in vivo
and in vitro grown plants as well as callus tissues) was Result and Discussion
extracted with methanol, ethyl acetate, acetone, Antifungal activity
chloroform and petroleum ether. The filtered solutions Effect of different dry biomass of A. officinalis
were evaporated to dry by being placed in a water bath L. on linear growth of some fungi on PDA medium
at 40°C overnight. Plant extracts were concentrated and by using different solvents
preserved at 4°C until required for the experiments.
Effect of A. officinalis L. on the linear growth of R.
Antifungal activity assay solani on the PDA medium represented in table 1 and
Antifungal screening: The five phytopathogenic Fig. 1 showed that different extracts caused a significant
fungi, including Alternaria tenuissima, Botrytis cinerea, inhibition on the linear growth of R. solani compared to
Fusarium oxysporum, Macrophomina phaseolina and the control. Different root extracts were more effective
Rhizoctonia solaniwere isolated from infected samples. than shoot extracts at most concentrations used. Both
Samples of different plants, i.e. strawberry, tomato and shoot petroleum ether extract and root ethyl acetate
cucumber were collected from Beheira and Qalyubia extract at the concentration of 600 µg ml-1 were superior
governorates, Egypt. The isolated fungi were purified out in inhibiting the linear growth of R. solani with inhibition
either by hyphal tip or single spore technique (Dhingra percent of 100%.
and Sinclair, 1985). Pure cultures were maintained during Data in table 2 and Fig. 2 revealed that the different
the experiments on potato dextrose agar (PDA 200g extracts caused a significant inhibition on the linear growth
grated potato, 20g dextrose, 20g agar), In vitro antifungal of F. oxysporum compared to the control. Different shoot
assays of prepared against the fungus were performed and root extracts caused significant inhibition on the linear
with the poisoned plate technique (Das et al., 2010). growth at all concentrations used compared to the control.
Antibacterial activity assay Different root extracts significantly inhibited linear growth
Prior to the test, bacterial cultures Bacillus subtilis at all concentrations used compared to shoot extracts
and Ralstonia solanacearum were prepared as follows: except chloroform and petroleum ether extracts. Shoot
The bacterial cultures were sub-culture in nutrient broth chloroform extracts caused non-significant inhibition on
30°C for 24-27 h. The broth culture turbidity of was the linear growth at both used concentrations compared
 Antimicrobial Effect of Asparagus officinalis L. Extracts 9255

Table 1: Effect of dry biomass extracts of (A. officinalis L.) on Table 2: Effect of dry biomass extracts of (A. officinalis L.) on
the linear growth of R. solani on the PDA medium by the linear growth of F. oxysporum on the PDA medium
using different extraction solvents. by using different extraction solvents.
Concentration µg ml -1 Concentration µg ml -1
Bio- 400 600 Bio- 400 600
mass Solvent Linear Inhi- Linear Inhi- mass Solvent Linear Inhi- Linear Inhi-
type growth biti- growth biti- type growth biti- growth biti-
(cm) onb(%) (cm) onb(%) (cm) onb(%) (cm) onb(%)
Shoot Methanol 4.95 45 2.05 77.22 Shoot Methanol 5.75 36.11 5.05 43.89
Ethyl acetate 5.09 43.44 3.69 59 Ethyl acetate 5.40 40 4.60 48.89
Petroleum ether 5.37 40.33 0 100 Petroleum ether 3.05 66.11 1.10 87.78
Acetone 2.75 69.44 1.82 79.78 Acetone 4.00 55.56 2.85 68.33
Chloroform 5.83 35.22 3.87 57 Chloroform 5.30 41.11 5.05 43.89
Root Methanol 2.38 73.56 1.35 85 Root Methanol 4.80 46.67 0 100
Ethyl acetate 2.43 73 0 100 Ethyl acetate 4.55 49.44 3.65 59.44
Petroleum ether 3.13 65.22 0.89 90.11 Petroleum ether 4.60 48.89 4.35 51.67
Acetone 0.75 91.67 0.47 94.78 Acetone 3.80 57.78 1.00 88.89
Chloroform 5.74 36.22 3.78 58.00 Chloroform 5.40 40 5.05 43.89
Control 9 — 9 — Control — 9 — 9 —
LSD(P  0.05) solvent(S) 0.29 — 1.36 — LSD(P0.05) solvent (S) 1.42 — 0.66 —
LSD(P  0.05) Type (T) 0.17 — 0.79 — LSD(P0.05) Type (T) 0.82 — 0.38 —
LSD(P  0.05) S x T 0.41 — 1.93 — LSD(P0.05) S x T 2.01 — 0.93 —
a
a
Mean of three replicates b Inhibition % = (Control-Treatment)/ Mean of three replicates b Inhibition % = (Control-Treatment)/
control)*100) control)*100).
with root chloroform extracted. Root methanol extract significant inhibition on the linear growth at all
at the concentration of 600 µg ml -1 had the highest concentrations used in comparison with shoot extracts.
inhibition percent of 100%. There were no differences between values of shoot and
Regarding table 3 and Fig. 3, different extracts caused root petroleum ether extracts and shoot chloroform extract
significant inhibition on the linear growth of M. at the low concentration. Shoot ethyl acetate extract at
phaseolina compared to the control. Different shoot and 400 µg ml-1 and 600 µg ml-1 recorded the highest inhibition
root extracts caused significant inhibition on the linear value 73.89 and 82.22%, respectively in comparison with
growth at all concentrations used compared to control. the other treatments.
Root methanol and chloroform extracts resulted in a Data in table 4 and Fig. 4 showed that different

Fig. 1: Effect of dry biomass extracts of (A. officinalis L.) on inhibition zone of R. solani on the PDA medium by using different
extraction solvents.
9256 S.F. Desoukey et al.

Fig. 2: Effect of dry biomass extracts of (A. officinalis L.) on inhibition zone of F. oxysporum on the PDA medium by using
different extraction solvents.

Fig. 3: Effect of dry biomass extracts of (A. officinalis L.) on inhibition zone of M. phaseolina on the PDA medium by
using different extraction solvents.
extracts caused significant inhibition on the linear growth acetone extract at 600 µg ml-1 as well as shoot ethyl
of A. tenuissima compared with control. Different shoot acetate extract at 400 and 600 µg ml-1 gave the superior
and root extracts caused a significant inhibition on the inhibition effect (100%) than the other treatments.
linear growth at all concentrations used in comparison The different dry biomass types of A. officinalis L.
with control. Different root extracts gave inhibition values were extracted by absolute methanol, ethyl acetate,
more than shoot extracts on the linear growth at all acetone, chloroform and petroleum ether as solvents differ
concentrations used. The highest inhibition value (100%) in their polarities. The purpose was to evaluate their
was recorded with root chloroform extract at the bioactivity against some pathogenic fungi using the
concentration of 600 µg ml-1. poisoned plate technique as found in Asparagus
Effect of A. officinalis L. on the linear growth of B. racemosus, Sangvikar (2012) and Parveen (2020). Shoot
cinerea on the PDA medium in table 5 and Fig. 5 showed and root extracts tables 1 to 5 significantly inhibited all of
that different extracts caused a significant inhibition on the studied pathogenic fungi compared to control. Root
linear growth. Root acetone extracts caused a significant extracts had a more inhibitory effect on the A. tenuissima
inhibition on the linear growth at all concentrations used than shoot extracts. Root and shoot extracts had a
in comparison with shoot acetone extracts. Shoot fluctuating inhibitory effect on B. cinerea, F.oxysporum,
chloroform extract was non-significant compared with M. phaseolina and R. solani. Growth reduction
root extract at low and high concentration used. Root percentages ranged between (36.22 to 91.67%) and (35.22
 Antimicrobial Effect of Asparagus officinalis L. Extracts 9257

Table 3: Effect of dry biomass extracts of (A. officinalis L.) on Table 4: Effect of dry biomass extracts of (A. officinalis L.) on
the linear growth of M. phaseolina on the PDA the linear growth of A. tenuissima on the PDA medium
medium by using different extraction solvents. by using different extraction solvents.
Concentration µg ml -1 Concentration µg ml -1
Bio- 400 600 Bio- 400 600
mass Solvent Linear Inhi- Linear Inhi- mass Solvent Linear Inhi- Linear Inhi-
type growth biti- growth biti- type growth biti- growth biti-
(cm) onb(%) (cm) onb(%) (cm) onb(%) (cm) onb(%)
Shoot Methanol 7.15 20.56 6.05 32.78 Shoot Methanol 5.55 38.33 5.00 44.44
Ethyl acetate 2.35 73.89 1.60 82.22 Ethyl acetate 6.50 27.78 5.35 40.56
Petroleum ether 7.50 16.67 5.30 41.11 Petroleum ether 5.20 42.22 1.75 80.56
Acetone 5.50 38.89 3.50 61.11 Acetone 4.20 53.33 2.90 67.78
Chloroform 7.50 16.67 6.10 32.22 Chloroform 6.20 31.11 0.80 91.11
Root Methanol 6.00 33.33 4.60 48.89 Root Methanol 4.10 54.44 3.20 64.44
Ethyl acetate 6.10 32.22 4.05 55 Ethyl acetate 5.35 40.56 5.25 41.67
Petroleum ether 7.50 16.67 4.10 54.44 Petroleum ether 2.80 68.89 1.30 85.56
Acetone 6.85 23.89 4.70 47.78 Acetone 3.50 61.11 0.80 91.11
Chloroform 6.20 31.11 3.00 66.67 Chloroform 5.00 44.44 0 100
Control 9 — 9 — Control 9 — 9 —
LSD(P0.05) solvent (S) 0.24 — 0.88 — LSD(P0.05) solvent (S) 0.6 — 0.87 —
LSD(P0.05) Type (T) 0.14 — 0.51 — LSD(P0.05) Type (T) 0.34 — 0.5 —
LSD(P0.05) S x T 0.33 — 1.25 — LSD(P0.05) S x T 0.84 — 1.23 —
a
Mean of three replicates b Inhibition % = (Control-Treatment)/ a
Mean of three replicates b Inhibition % = (Control-Treatment)/
control)*100) control)*100)

Fig. 4: Effect of dry biomass extracts of(A. officinalisL.) on inhibition zone of A. tenuissima on the PDA medium by using
different extraction solvents. 100%) for B. cinerea in the case of root and shoot
to 69.44%) for R. solani; (40.56 to 68.89%) and (27.78 extracts, respectively, at the low concentration. Battu and
to 53.33%) for A. tenuissima and (40.00 to 57.78%) and Kumar (2010) and Tinrat and Sila-asna (2017) found that
(36.11 to 66.11%) for F. oxysporum in the case of root biological activity may be due partly to the presence of
and shoot extracts, respectively, at the low concentration. various phytochemical compounds in A. racemosus like;
On the other hand, the different dry biomass (root and cardiac glycosides, flavonoids, phenolics, saponins,
shoot) extracts showed different patterns of effects on steroids and terpenoids.
M. phaseolina and B. cinerea as inhibition percentages Inhibition percent caused significant or non-significant
ranged between (16.67 to 33.33%) and (16.67 to 73.89%) decreases in mycelial growth by increasing the
for M. phaseolina and (16.67 to 86.67%) and (16.67 to concentration of the extracts. Shrestha et al., (2018)
9258 S.F. Desoukey et al.

Table 5: Effect of dry biomass extracts of (Asparagus Table 6: Effect of different fresh biomass types extracts of (A.
officinalis L.) on the linear growth of B. cinerea on officinalis L.) plant on the linear growth of R. solani
the PDA medium by using different extraction on the PDA medium by using different extraction
solvents. solvents.
Concentration µg ml -1 Concentration µg ml -1
Bio- 400 600 Bio- 400 600
mass Solvent Linear Inhi- Linear Inhi- mass Solvent Linear Inhi- Linear Inhi-
type growth biti- growth biti- type growth biti- growth biti-
(cm) onb(%) (cm) onb(%) (cm) onb(%) (cm) onb(%)
Shoot Methanol 5.75 36.11 3.10 65.56 Shoot Methanol 5.5 38.89 1.3 85.56
Ethyl acetate 0 100 0 100 Ethyl acetate 1.35 85 0 100
Petroleum ether 6.85 23.89 6.10 32.22 Petroleum ether 6.3 30 3.85 57.22
Acetone 5.60 37.78 2.30 74.44 Acetone 7.35 18.33 4.8 46.67
Chloroform 7.50 16.67 7.50 16.67 Chloroform 6.5 27.78 5.3 41.11
Root Methanol 7.50 16.67 7.50 16.67 Root Methanol 6.3 30 4.5 50
Ethyl acetate 1.20 86.67 0.65 92.78 Ethyl acetate 1.55 82.78 0 100
Petroleum ether 7.50 16.67 0.45 95 Petroleum ether 6.55 27.22 2.3 74.44
Acetone 1.40 84.44 0 100 Acetone 3.25 63.89 3.65 59.44
Chloroform 7.50 16.67 7.50 16.67 Chloroform 6.95 22.77 5.45 39.44
Control 9 — 9 — Callus Methanol 5.5 38.89 2.4 73.33
LSD(P0.05) solvent (S) 0.2 — 0.28 — Ethyl acetate 2.1 76.67 0 100
LSD(P0.05) Type (T) 0.11 — 0.16 — Petroleum ether 4.05 55 3.1 65.55
LSD(P0.05) S x T 0.28 — 0.40 — Acetone 6.1 32.22 5.55 38.33
a
Mean of three replicates b Inhibition % = (Control-Treatment)/ Chloroform 5.8 35.56 5 44.4
control)*100) Control 9 — 9 —
LSD(P0.05) solvent (S) 0.7 — 0.86 —
stated that A. racemosus has shown selected
LSD(P0.05) Type (T) N.S — N.S —
antimicrobial effects against S. cerevisiae and C.
LSD(P0.05) S x T 1.2 — 1.5 —
albicans with inhibition zone of 25 mm in an average.
a
Shukla (2018) mentioned that the antimicrobial activity Mean of three replicates b Inhibition % = (Control-Treatment)/
of hydroalcoholic extract (80% ethanol) of Glycyrrhiza control)*100)
glabra root and A. racemosus leaves due to the presence fungi on the PDA medium by using different
of different phytochemicals with biological activity like extraction solvents
saponin, phenol, carbohydrates, flavonoids, proteins and Effect of different fresh biomass types extracts of
amino acids. A. officinalis L. plant on the linear growth of R. solani
Effect of different fresh biomass types extracts of on the PDA medium are clarified in table 6. Different
A. officinalis L. plant on the linear growth of some solvent extracts caused a significant inhibition on the linear

Fig. 5: Effect of dry biomass extracts of (A. officinalis L.) on inhibition zone of B. cinerea on the PDA medium by using different
extraction solvents.
 Antimicrobial Effect of Asparagus officinalis L. Extracts 9259

growth. Different types of extracts caused non-significant significant inhibition on the linear growth of M.
inhibition on the linear growth. It showed an interaction phaseolina compared to the control. The inhibition
between fresh biomass type and solvent type. Ethyl percent ranged from 16.67 to 28.89% and 16.67 to 52.78%
acetate extracts of different fresh biomass types recorded in the case of different fresh biomass types and solvents
the highest inhibition values on the linear growth being; at low and high concentrations, respectively. Callus
(100, 100, 100%) and (85, 82.78, 76.67%) for shoot, root methanol extract caused significant inhibition (28.89%)
and callus at the concentration of 600 and 400 µg ml-1, on the linear growth at the low-use concentration in
respectively. comparison with different fresh biomass types and
Data in table 7 revealed that different extracts caused solvents. The inhibition percent was the highest (52.78%)
a significant inhibition on the linear growth of F. in the case of root acetone extract at high concentration
oxysporum compared to the control. Different shoot, root compared to different fresh biomass types and solvents.
and callus extracts significantly inhibited linear growth in Data in table 9 indicated the major differences on
all concentrations used compared to the control. Different the linear growth of A. tenuissima between different
root extracts significantly inhibited linear growth in most extracts within the fresh biomass type and the solvent.
concentrations used compared to the shoot and callus Different shoot, root and callus extracts significantly
extracts. Callus methanol and chloroform extracts caused inhibited linear growth in all concentrations used compared
significant inhibition on the linear growth in all to control. Callus chloroform extract showed the highest
concentrations used compared to the shoot and root inhibition percent (69.44%), followed by shoot petroleum
extracts except for shoot methanol extract at high ether extract (58.33%), then shoot chloroform extract
concentration. (49.44%) at low concentration. Different callus extracts
Regarding table 8, different extracts caused showed the highest inhibition percent at high concentration
Table 7: Effect of different fresh biomass types extracts of (A. Table 8: Effect of different fresh biomass types extracts of (A.
officinalis L.) plant on the linear growth of F. officinalis L.) plant on the linear growth of M.
oxysporum on the PDA medium by using different phaseolina on the PDA medium by using different
extraction solvents. extraction solvents.
Concentration µg ml -1 Concentration µg ml -1
Bio- 400 600 Bio- 400 600
mass Solvent Linear Inhi- Linear Inhi- mass Solvent Linear Inhi- Linear Inhi-
type growth biti- growth biti- type growth biti- growth biti-
(cm) onb(%) (cm) onb(%) (cm) onb(%) (cm) onb(%)
Shoot Methanol 5.7 36.67 0.55 93.89 Shoot Methanol 7 22.22 6.25 30.56
Ethyl acetate 6 33.33 4.1 54.44 Ethyl acetate 7.5 16.67 6 33.33
Petroleum ether 5.65 37.22 4.05 55 Petroleum ether 6.95 22.78 4.85 46.11
Acetone 5.85 35 4.6 48.89 Acetone 7.5 16.67 7.5 16.67
Chloroform 5.95 33.89 5.45 39.44 Chloroform 7.5 16.67 6.15 31.67
Root Methanol 5.7 36.67 4.6 48.89 Root Methanol 7.05 21.67 4.75 47.22
Ethyl acetate 4.9 45.56 4.05 55 Ethyl acetate 6.55 27.22 5.85 35
Petroleum ether 4.75 47.22 3.5 61.11 Petroleum ether 7.5 16.67 7.4 17.78
Acetone 5.3 41.11 4.2 53.33 Acetone 7.25 19.44 4.25 52.78
Chloroform 6.3 30 5.1 43.33 Chloroform 7 22.22 5.6 37.78
Callus Methanol 5.25 41.67 4.8 46.67 Callus Methanol 6.4 28.89 5.25 41.67
Ethyl acetate 5 44.44 4.35 51.67 Ethyl acetate 7.5 16.67 6.45 28.33
Petroleum ether 5.7 36.67 4.55 49.44 Petroleum ether 7.5 16.67 7.5 16.67
Acetone 5.95 33.89 4.6 48.89 Acetone 7.3 18.89 7.15 20.56
Chloroform 3.65 59.44 0 100 Chloroform 7.5 16.67 6.35 29.44
Control 9 — 9 — Control 9 — 9 —
LSD(P0.05) solvent (S) 0.56 — 0.5 — LSD(P0.05) solvent (S) 0.27 — 0.55 —
LSD(P0.05) Type (T) 0.4 — 0.36 — LSD(P0.05) Type (T) N.S — 0.39 —
LSD(P0.05) S x T 0.97 — 0.87 — LSD(P0.05) S x T 0.46 — 0..95 —
a
Mean of three replicates b Inhibition % = (Control-Treatment)/ a
Mean of three replicates b Inhibition % = (Control-Treatment)/
control)*100) control)*100)
9260 S.F. Desoukey et al.

used compared to shoot and root extracts with the the antimicrobial activity of different ethanol extracts of
exception of methanol extract. A. officinalis (in vivo plant, in vitro and callus tissues)
Effect of A. officinalis L. extracts on the linear against some gram-positive and gram negative. Showed
growth of B. cinerea on the PDA medium given in table that only antimicrobial activity was obtained from in callus
10 showed that different extracts caused a significant tissue against B. cereus (+ve). Flavonoids are phenolic
inhibition on the linear growth compared to the control. compounds that possess a wide range of biological
Petroleum ether and acetone extracts in the case of activities, including antifungal activity obtained from
different fresh biomass types caused non-significant “triguero” asparagus by-products against F. oxysporum
inhibition of each other on the linear growth at different (Rosado- Álvarez et al., 2014).
concentrations used with the exception of callus at high Antibacterial activity
concentration caused a significant inhibition of each other Inhibition effect of different dry biomass of A.
on the linear growth. Root ethyl acetate extract showed officinalis L. on growth of two bacteria (+ve and -
the highest inhibition percent (91.67 and 100%) at the ve) on PDA medium by using different solvents
concentration of 400 and 600 µg ml-1, respectively, in
In this study, the different dry shoot and root extracts
comparison with the same solvent and the other fresh
of A. officinalis L. were tested for their antibacterial
biomass types. Different callus extracts showed the
activity against Gram-negative and positive bacteria.
highest inhibition percent at high used concentration in
Strains of R. solanacearum and B. subtilis were used.
comparison with shoot and root extracts except for ethyl
Dry shoot and root methanol extracts showed antibacterial
acetate and chloroform extracts. Study on A. officinalis
activity (positive and negative Gram (+ve & -ve)) similar
showed that these bioactivities differ between in vitro
to the control tables 11 and 12. Dry shoot ethyl acetate
and in vivo grown plants. Khorasani et al., 2010 studied
Table 9: Effect of different fresh biomass types extracts of (A. Table 10: Effect of different fresh biomass types extracts of
officinalis L.) plant on the linear growth of A. (A. officinalis L.) plant on the linear growth of B.
tenuissima on the PDA medium by using different cinerea on the PDA medium by using different
extraction solvents. extraction solvents.
Concentration µg ml -1 Concentration µg ml -1
Bio- 400 600 Bio- 400 600
mass Solvent Linear Inhi- Linear Inhi- mass Solvent Linear Inhi- Linear Inhi-
type growth biti- growth biti- type growth biti- growth biti-
(cm) onb(%) (cm) onb(%) (cm) onb(%) (cm) onb(%)
Shoot Methanol 5.95 33.89 5.65 37.22 Shoot Methanol 7.5 16.67 5.45 39.44
Ethyl acetate 6.25 30.56 4.15 53.89 Ethyl acetate 6 33.33 0.5 94.44
Petroleum ether 3.75 58.33 3.6 60 Petroleum ether 7.5 16.67 7.5 16.67
Acetone 6.05 32.78 3.8 57.78 Acetone 7.5 16.67 7.5 16.67
Chloroform 4.55 49.44 0.9 90 Chloroform 7.5 16.67 1 88.89
Root Methanol 5.55 38.33 4.6 48.89 Root Methanol 1.6 82.22 1 88.89
Ethyl acetate 5.6 37.78 5.1 43.33 Ethyl acetate 0.75 91.67 0 100
Petroleum ether 6.7 25.56 5.75 36.11 Petroleum ether 7.5 16.67 7.5 16.67
Acetone 6.25 30.56 5.7 36.67 Acetone 7.5 16.67 7.5 16.67
Chloroform 5.4 40 4.1 54.44 Chloroform 6.55 27.22 4.55 49.44
Callus Methanol 6.4 28.89 5.55 38.33 Callus Methanol 1.5 83.33 0.95 89.44
Ethyl acetate 5.25 41.67 1.45 83.89 Ethyl acetate 7.3 18.89 6.05 32.78
Petroleum ether 6.5 27.78 1.75 80.56 Petroleum ether 7.5 16.67 6.2 31.11
Acetone 5.45 39.44 3.3 63.33 Acetone 7.45 17.22 2.5 72.22
Chloroform 2.75 69.44 0 100 Chloroform 7.5 16.67 7.5 16.67
Control 9 — 9 — Control 9 — 9 —
LSD(P0.05) solvent (S) 0.44 — 0.71 — LSD(P0.05) solvent (S) 0.65 — N.S —
LSD(P0.05) Type (T) 0.31 — 0.5 — LSD(P0.05) Type (T) 0.46 — 0.65 —
LSD(P0.05) S x T 0.77 — 1.2 — LSD(P0.05) S x T 1.12 — 1.59 —
a
Mean of three replicates b Inhibition % = (Control-Treatment)/ a
Mean of three replicates b Inhibition % = (Control-Treatment)/
control)*100) control)*100)
 Antimicrobial Effect of Asparagus officinalis L. Extracts 9261

Table 11: Antibacterial activity of different types of dry extract caused inhibition against Gram- positive bacteria
biomass extracts of (A. officinalis L.) plant against (+ve) but dry root ethyl acetate extract caused inhibition
selected strains of bacteria. against Gram- positive and negative bacteria (+ve & –
Biomass Solvent Conc. Strains of bacteria ve). Dry shoot and root petroleum ether extracts were
type R. solana- B. subti- not active against Gram-positive and negative bacteria
cearum(-ve) lis(+ve) (+ve & –ve), except shoot petroleum ether extract against
Shoot Methanol Low + + Gram- negative and positive bacteria (-ve & +ve) at high
High + + concentration. Dry shoot acetone extract inhibited only
Ethyl acetate Low - + Gram-positive bacteria (+ve) but dry root acetone extract
High - + inhibited both types of bacteria. Dry shoot chloroform
Petroleum ether Low - - extract was active against Gram-positive and negative
High + + bacteria (+ve and –ve) at high and low concentrations,
Acetone Low - + except dry shoot chloroform extract against Gram-
High - + positive bacteria (+ve) at low concentration. Dry root
Chloroform Low + - chloroform extract was only active against Gram-positive
High + + bacteria (+ve) at high concentration.
Root Methanol Low + +
Inhibition effect of different fresh biomass of A.
High + +
officinalisL. on growth of two bacteria (+ve and -
Ethyl acetate Low + +
ve) on PDA medium by using different solvents
High + +
Petroleum ether Low - - Tables 13 and 14 show the effect of fresh shoot, root
High - - and callus extracts against the two Gram-positive and
Acetone Low + + negative bacterial strains mentioned before. Fresh shoot
High + + and root methanol extracts at low and high concentrations
Chloroform Low - - did not affect the two bacterial strains (+ve & -ve). While,
High - + in callus, it gave inhibitory effect at both concentrations
Table 12: Inhibition zones of different concentrations of dry against Gram-positive bacteria (+ve) as well as at high
biomass extracts of (A. officinalis L.) plant against concentration against Gram-negative bacteria (-ve).
selected strains of bacteria. Callus chloroform extract gave inhibition effect against
Solvent Conc. Strains of bacteria two bacterial strains (+ve & -ve) at both low and high
Biomass
type R. solana- B. subti-
concentrations, while in fresh root chloroform extract,
cearum(-ve) lis(+ve) this effect exhibited only in Gram-positive bacteria (+ve).
Shoot Methanol Low 0.4 1.1 On the other hand, fresh shoot chloroform extract did
High 1.2 1.6 not affect the bacterial activity. In the case of fresh shoot
Ethyl acetate Low 0 0.67 acetone extract, the effect was only valid against Gram-
High 0 0.77 negative bacteria at high concentration but had no effect
Petroleum ether Low 0 0 against Gram-positive bacteria. Fresh root and callus
High 1.2 1.1 acetone extracts had no effect against both bacterial
Acetone Low 0 1.2 strains at low and high concentrations. Fresh shoot and
High 0 1.9 callus ethyl acetate extracts gave inhibitory effect against
Chloroform Low 0.9 0 both strains (+ve & -ve) at low and high concentrations
High 1.13 1 but root ethyl acetate extract affected only gram-positive
Root Methanol Low 1.1 1.07 bacteria (+ve) at both concentrations. Fresh root
High 1.63 1.23 petroleum ether extract did not record any inhibitory effect
Ethyl acetate Low 0.43 1.1 against both bacterial strains (+ve & -ve). While shoot
High 1.23 1.67 and callus extracts at high concentration were valid
Petroleum ether Low 0 0 against gram-negative and positive bacteria (-ve & +ve),
High 0 0 respectively.
Acetone Low 0.83 1.03 Nair and Chanda (2006) mentioned that the aqueous
High 1.6 1.27 and ethanol extracts of A. racemosus showed the least
Chloroform Low 0 0 antibacterial activity compared with medicinal various
High 0 0.9 plants against seven gram-negative and five gram-positive
9262 S.F. Desoukey et al.

Table 13: Antibacterial activity of different types of fresh Table 14: Inhibition zones of different concentrations of fresh
biomass extracts of (A. officinalis L.) plant against biomass extracts of (A. officinalis L.) plant against
selected strains of bacteria. selected strains of bacteria.
Biomass Solvent Conc. Strains of bacteria Biomass Solvent Conc. Strains of bacteria
type R. solana- B. subti- type R. solana- B. subti-
cearum(-ve) lis(+ve) cearum(-ve) lis(+ve)
Shoot Methanol Low - - Shoot Methanol Low 0 0
High - - High 0 0
Ethyl acetate Low + + Ethyl acetate Low 0.63 0.73
High + + High 1 1.03
Petroleum ether Low - - Petroleum ether Low 0 0
High + - High 1.6 0
Acetone Low - - Acetone Low 0 0
High + - High 1.2 0
Chloroform Low - - Chloroform Low 0 0
High - - High 0 0
Root Methanol Low - - Root Methanol Low 0 0
High - - High 0 0
Ethyl acetate Low - + Ethyl acetate Low 0 0.7
High - + High 0 1.07
Petroleum ether Low - - Petroleum ether Low 0 0
High - - High 0 0
Acetone Low - - Acetone Low 0 0
High - - High 0 0
Chloroform Low - + Chloroform Low 0 0.6
High - + High 0 1.6
Callus Methanol Low - + Callus Methanol Low 0 0.67
High + + High 0.3 1.5
Ethyl acetate Low + + Ethyl acetate Low 0.6 0.7
High + + High 1.2 1.37
Petroleum ether Low - - Petroleum ether Low 0 0
High - + High 0 0.9
Acetone Low - - Acetone Low 0 0
High - - High 0 0
Chloroform Low + + Chloroform Low 0.6 0.3
High + + High 0.9 0.67

bacteria. Aqueous extracts and ethanol extracts of A. bacteria (B. subtilis, Micrococcus luteus and S. aureus)
racemosus showed no effect against different used and gram-negative bacteria (E. coli, Pseudomonas
bacteria (+ve and –ve) except Bacillus cereus (+ve). aeruginosa, two types of Shigella bacteria ( S.
Mishra et al., (2014) used roots methanol extract of (A. dysenteriae and S. flexneri) and Vibrio cholerae). Patel
racemosus) against gram-positive and negative bacteria and Patel (2013) used that A. racemosus leaves extracts
which showed significant in vitro antibacterial efficacy with different solvents like acetone, chloroform, ethyl
against gram-negative bacteria (Escherichia coli, three acetate, methanol, petroleum ether and water. Recorded
types of Shigella bacteria (Shigella dysenteriae, S. that different extract of A. racemosus leaves have
sonnei, S. flexneri), Vibrio cholerae, two types of antimicrobial activity against Gram positive as well as
salmonella bacteria (Salmonella typhi, S. typhimurium) Gram negative bacteria.Shevale et al., (2015)
and Pseudomonas putida) and gram-positive bacteria demonstrated in vitro antibacterial activity of crude
(B. subtilis and Staphylococcus aureus). Sinha and ethanol and acetone extract of A. racemosus roots
Biswas (2011) mentioned that bactericidal activity of crude against gram-positive (S. aureus and B. subtilis) and
extracts from A. racemosus roots was screened against gram-negative (E. coli, Klebsiella pneumoniae and S.
eight pathogenic strains belonging to gram-positive typhi). Kishor et al., (2019) also found that in vitro
 Antimicrobial Effect of Asparagus officinalis L. Extracts 9263

shoot and root were used. Also, the callus ethyl acetate
extract gave the same result against R. solani. Acetone
extract gave positive results against A. tenuissima, R.
solani and B. cinerea when dry root was used. When
petroleum ether extract was used, the result was more
than 91% against R. solani and B. cinerea when dry
shoot and dry root were used, respectively. In the case
of chloroform extract, the realized result was more than
or equal 90% against A. tenuissima when callus, fresh
shoot, dry shoot and dry root were used at high
concentration. M. phaseolina gave the least inhibition
1: Shoot 2: Root 3: Callus N: Control M: Methanol A: percent.
Acetone P: Petroleum C: Chloroform E: Ethyl The concentration of the extract has a positive
acetate relationship with inhibiting activity against bacterial growth
Fig 6: Inhibition zones of different concentrations of fresh
for example; fresh shoot petroleum ether extract against
and dry biomass extracts of (A. officinalis L.) plant
gram negative bacteria (–ve). Also, the bacterial type is
against B. subtilis (+ve) at high concentration.
considered one of the important factors for example; fresh
root ethyl acetate extract has inhibitory effect against
gram positive bacterial (+ve). Using different types of A.
officinalis L. biomass extracts as an antibacterial led to
positive results. In the case of using dry root ethyl acetate,
acetone ether or chloroform extract, they had an inhibitory
effect against Gram- negative bacteria (-ve) compared
to dry shoot of the same solvents extracts. Also, methanol
extract for dry shoot and root had an inhibitory effect on
bacterial strains (+ve & -ve).
Further studies are needed using isolates from other
fungi and bacteria. Other in vivo experiments are also
needed to confirm the effect of plant extracts.
1: Shoot 2: Root 3: Callus N: Control M: Methanol A:
Acetone P: Petroleum C: Chloroform E: Ethyl References
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Fig. 7: Inhibition zones of different concentrations of fresh
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