characterization of antimicrobial metabolites produced by
salvadora persica and analysis of its chemical compounds
using gc-ms and ftir
rafid hadi hameed1, ghaidaa jihadi mohammed2, imad hadi hameed3
1ministry of health, mesan health office, mesan city, iraq 2department of biology, college of science,
university of al-qadisiyah, hillah city, iraq 3biomedical science department, university of babylon,
college of nursing, hillah city, iraq
abstract
meswak (salvadora persica) is one of the most commonly used medicinal plants for oral hygiene among
global muslim community. the objectives of this study were analysis of the secondary metabolite products
using gc-ms and ftir and evaluation of antimicrobial activity. the ftir analysis proved the presence of
alkyl halides, alkanes, and alkenes which shows major peaks at 960.55, 1029.99, 1097.50, 1141.86, 1321.24,
1373.32, 1616.35, 1723.65, 2852.72 and 2922.16. our research recorded maximum zone formation was
against staphylococcus aureus (6.635±0.25). salvadora persica was very highly active against aspergillus
terreus 6.77±0.24. twenty one bioactive compounds were identified in the methanolic extract of salvadora
persica. gc-ms analysis of salvadora persica revealed the existence of the 5?-androstan-16-one ,
cyclic ethylene mercaptole , glycerin , 3,5-dithiahexanol 5,5-dioxide , 1h-pyrazole-1-carbothioamide
,3,5-dimethyl- , 17-octadecynoic acid , benzenepropanoic acid , ?-(hydroxyimino)- , indan-1,2-dione ,
4-methyl- , butanoic acid , 4,4’-dithiobis[2-amino-, [s-(r*,r*)]- , 5-hydroxymethylfurfural , 1h-pyrrole
, 1-(phenylmethyl)- , cyclopentanone , 2-cyclopentylidene- , phenol ,2-methoxy-6-(1-propenyl)- ,
2,7-diphenyl-1,6-dioxopyridazino[4,5:2’,3’]pyrrolo[4’,5’-d]pyrida , 2-(4-(but-2-yl)phenyl)propanoic acid ,
3h-cyclodeca[b]furan-2-one , 4,9-dihydroxy-6-methyl-3,10-dim , phenol ,2,6-dimethoxy-4-(2-propenyl)- ,
butylaldehyde ,4-benzyloxy-4-[2,2,-dimethyl-4-dioxolanyl]- , benadryl , 9-hexadecenoic acid , i-propyl
11,12-methylene-octadecanoate and 2,6-bis[2-[2-s-thiosulfuroethylamino ] ethoxy]pyrazine.
keywords: gc-ms, ft-ir, salvadora persica, pseudomonas aerogenosa, proteus mirabilis, staph. aureus,
klebsiella pneumonia
corresponding author:
imad hadi hameed
biomedical science department,
university of babylon, college of nursing,
hillah city, iraq
phone: 009647716150716
e-mail: imad_dna@yahoo.com
introduction
salvadora persica has antiurolithiatic properties.
used for centuries as a natural toothbrush, its fibrous
branches have been promoted by the world health
organization for oral hygiene use1-6. research suggests
that it contains a number of medically beneficial properties
including abrasives, antiseptics, astringent, detergents,
enzyme inhibitors, and fluoride. previous studies have
reported that s. persica extracts were effective against
streptococcus mutans and streptococcus faecalis,
even using low extract concentrations. plaque is found
preferentially at protected and stagnant surfaces, and
these are at the greatest risk of disease7-15. moreover,
the attachment, growth, removal and reattachment of
bacteria to the tooth surface are a continuous and dynamic
process. dental plaque, biofilms of microorganisms on
tooth surface, plays an important role in the development
of caries and periodontal disease16-23. plaque is found
preferentially at protected and stagnant surfaces, and
these are at the greatest risk of disease. it was established
that mutans group of streptococci are the key agents
doi number: 10.5958/0976-5506.2018.00216.4
242 indian journal of public health research & development, march 2018, vol.9, no. 3
causing dental caries24-29. according to studies, mutans
streptococci can colonize the tooth surface and
initiate plaque formation by their ability to synthesize
extracellular polysaccharides from sucrose, using
glucosyl transferase. this sucrose dependent adherence
and accumulation of cariogenic streptococci is critical to
the development of pathogenic plaque30-33. some types
of interactions are thought to be of primary importance
in the colonization of the periodontal environment. the
further accumulation of plaque around the gingival and
subgingival region may lead to a shift in its microbial
composition from streptococcus-dominated to a larger
number of actinomyces spp., and an increased number
of capnophilic and obligatory anaerobic bacteria, such as
porphyromonas gingivalis. several in vitro studies have
indicated that salvadora persica contains substances
that possess dental plaque inhibiting properties against
oral microbes34-38. the fresh leaves can be eaten as part
of a salad and are used in traditional medicine for cough,
asthma, scurvy, rheumatism, piles and other diseases.
the flowers are small and fragrant and are used as a
stimulant and are mildly purgative39-42. the berries are
small and barely noticeable they are eaten both fresh
and dried. the aims of our research were analysis of
the secondary metabolite products and determination of
antibacterial and antifungal activity.
material and method
gas chromatography–mass spectrum and fourier
transform infrared spectrophotometer analysis:
salvadora persica gc–ms analysis were carried out
in a gc system (agilent 7890a series, usa). the
flow rate of the carrier gas, helium (he) was set to
beat 1 ml min?1, split ratio was 1:50. interpretation
of mass spectrum was conducted using the database of
national institute of standards and technology (nist,
usa). the powdered sample of salvadora persica was
treated for ftir spectroscopy (shimadzu, ir affinity
1, japan)43,44. the sample was run at infrared region
between 400 nm and 4000 nm. results of the study were
based on analysis of variance (anova) using statistica
software. a significance level of 0.05 was used for all
statistical tests.
antibacterial and antifungal activity of natural
compounds of salvadora persica: the test pathogenic
bacteria were swabbed in müller-hinton. seventy ml of
plant extract was loaded on the bored wells. antifungal
activity was evaluated by measuring the zone of
inhibition against the test fungi. methanol was used as
solvent control. amphotericin b and fluconazole were
used as reference antifungal agent. the tests were carried
out in triplicate. the antifungal activity was evaluated
by measuring the inhibition-zone diameter observed
after 48 h of incubation45-47.
table 1: major phytochemical compounds identified in methanolic extract of salvadora persica
molecular exact mass pharmacological actions
phytochemical compound rt (min) weight s.
no.
5?-androstan-16-one , cyclic 3.264 350 350.210194 antimicrobial activity
1. ethylene mercaptole
2. glycerin 3.310 92 92.047344 anti-inflammatory
3. 3,5-dithiahexanol 5,5-dioxide 3.379 170 170.007136 anti-tumour activity
1h-pyrazole-1-carbothioamide 3.625 155 155.051719 anti-inflammatory
4. ,3,5-dimethyl5.
17-octadecynoic acid 3.768 280 280.24023 anti- diarrheal
benzenepropanoic acid , 4.975 179 179.058243 antimicrobial activity
6. ?-(hydroxyimino)-
antimicrobial, antitumor,
7. indan-1,2-dione , 4-methyl- 5.353 160 160.052429 anti-inflammatory, antiviral
butanoic acid , 4,4’-dithiobis[2- 5.759 268 268.05515 anti-inflammatory agents
8. amino-, [s-(r*,r*)]-
anti-inflammatory
9. 5-hydroxymethylfurfural 6.366 126 126.031694 properties
10. 1h-pyrrole , 1-(phenylmethyl)- 7.424 157 157.089149 anti-candida activities
indian journal of public health research & development, march 2018, vol.9, no. 3 243
anti-inflammatory,
8.002 150 150.1044655 analgesic, anticonvulsant cyclopentanone ,
11. 2-cyclopentylidenephenol
,2-methoxy-6-(1- 8.425 164 164.08373 antibacterial
12. propenyl)-
9.930 355 355.106924 new chemical compound
2,7-diphenyl-1,6-
dioxopyridazino[4,5:2’,3’]
pyrrolo[4’,5’-d]pyrida
13.
anti-inflammatory agent
10.325 206 206.13068 with analgesic properties 2-(4-(but-2-yl)phenyl)propanoic
14. acid
3h-cyclodeca[b]furan-2-one , 10.617 264 264.13616 antioxidant activity
15. 4,9-dihydroxy-6-methyl-3,10-dim
phenol ,2,6-dimethoxy-4-(2- 11.092 194 194.094295 new chemical compound
16. propenyl)-
butylaldehyde ,4-benzyloxy-4- 12.597 278 278.15181 anti-inflammatory
17. [2,2,-dimethyl-4-dioxolanyl]-
18. benadryl 13.123 255 255.162314 antihistamine
19. 9-hexadecenoic acid 13.507 254 254.22458 anti-inflammatory
i-propyl 11,12-methylene- 13.501 338 338.318481 antibacterial activity
20. octadecanoate
14.222 478 478.032047 anti-bacterial activity
2,6-bis[2-[2-sthiosulfuroethylamino
] ethoxy]
pyrazine
21.
result s and discussion
identification of biochemical compounds: analysis
of compounds was carried out in methanolic extract of
salvadora persica, shown in table 1. chromatogram
gc-ms analysis of the methanol extract of salvadora
persica showed the presence of thirty one major peaks
and the components corresponding to the peaks were
determined as follows. all peaks were determined to
be 5?-androstan-16-one, cyclic ethylene mercaptole,
glycerin , 3,5-dithiahexanol 5,5-dioxide, 1h-pyrazole-
1-carbothioamide ,3,5-dimethyl-, 17-octadecynoic acid,
benzenepropanoic acid , ?-(hydroxyimino)- , indan-
1,2-dione, 4-methyl-, butanoic acid, 4,4’-dithiobis[2-
amino-, [s-(r*,r*)]-, 5-hydroxymethylfurfural,
1h-pyrrole, 1-(phenylmethyl)-, cyclopentanone,
2-cyclopentylidene-, phenol, 2-methoxy-6-(1-propenyl)-,
2,7-diphenyl-1,6-dioxopyridazino[4,5:2’,3’]pyrrolo[4’,
5’-d]pyrida, 2-(4-(but-2-yl)phenyl)propanoic acid,
3h-cyclodeca[b]furan-2-one, 4,9-dihydroxy-6-methyl-
3,10-dim, phenol, 2,6-dimethoxy-4-(2-propenyl)-,
butylaldehyde, 4-benzyloxy-4-[2,2,-dimethyl-4-
dioxolanyl]-, benadryl, 9-hexadecenoic acid, i-propyl
11,12-methylene-octadecanoate and 2,6-bis[2-[2-sthiosulfuroethylamino]
ethoxy]pyrazine. the ftir
analysis of salvadora persica leaves proved the presence
of alkyl halides, alkanes, and alkenes which shows major
peaks at 960.55, 1029.99, 1097.50, 1141.86, 1321.24,
1373.32, 1616.35, 1723.65, 2852.72 and 2922.16.
in the current study, the anti-microbial activity of
salvadora persica methanolic extract was determinate
by zone of inhibition against many bacteria, fungi
and yeast. clinical pathogens klebsiella pneumonia,
escherichia coli, staphylococcus aureus, salmonella
typhi, bacillus subtilis, streptococcus faecalis,
proteus mirabilis and streptococcus pyogenes were
selected for antibacterial activity. our research found
maximum zone formation was against staphylococcus
aureus (6.635 ± 0.25). antifungal activities against
aspergillus niger, penicillium expansum, trichoderma
horzianum, aspergillus terreus, candida albicans,
trichoderma viride, saccharomyces cerevisiae, and
microsporum canis. salvadora persica recorded high
antifungal activity against against aspergillus terreus
(6.77±0.24). medicinal properties of salvadora persica
extract is due to presence of bioactive metabolites.
twenty one phytoconstituents were identified by gas
chromatography – mass spectrum and fourier transform
infrared spectrophotometer. this plant derived bioactive
compounds used as source of antibiotic properties and
pharmaceutical industries used for drug formulation.
244 indian journal of public health research & development, march 2018, vol.9, no. 3
conclusion
twenty one bioactive chemical compounds were
identified by gc-ms and ft-ir analysis. this salvadora
persica derived bioactive compounds used as source of
antibiotic properties and pharmaceutical industries used
for drug formulation.
financial disclosure: there is no financial disclosure.
conflict of interest: none to declare.
ethical clearance: in this study, all experimental
protocols were approved under the department of
biology, college of science for women, university
of babylon, hillah city, iraq and all experiments were
carried out in accordance with approved guidelines.
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