ReseaRch PaPeR Journal of Agricultural and Marine Sciences 2020, 25(2): 58–66 DOI: 10.24200/jams.vol25iss2pp58-66 Received 18 Mar 2020 Accepted 08 June 2020 Antibacterial Activity and Chemical Composition of Crude Extract and Oil of Zygophyllum (Fagonia) luntii (Baker) 1894 (Family Zygophyllaceae) Riaz Shah1*, Suad J.A. Alabri2, Ameera S.M Ashehi2, Nasser S.S. Asiyabi2, Wafa K.A. AlMamari2, Jamal N. AlSabahi3 Huda Al-Ruqaishi3 Riaz Shah1*( ) riazshah@squ.edu.om,1Department of Plant Sciences, College of Agricultural and Marine Sciences, Sultan Qaboos University, Al-Khoud-123, Muscat, Sultanate of Oman. 2College of Science, Sultan Qaboos University, Al-Khoud-123, Muscat, Sultanate of Oman. 3Cen- tral analytical laboratory, College of Agricultural and Marine Sciences, Sultan Qaboos University, Al-Khoud-123, Muscat, Sultanate of Oman. Introduction Around 80% of the world population use tradi-tional medicine which is often based on medici-nal plants (Martins, 2013; Oyebode et al., 2016). Around 75% of commercial drugs launched in the world النشاط املضاد للبكترياي والرتكيب الكيميائي للمستخلص اخلام وزيت زيغوفيللوم )فاجونيا( لونيت )بيكر( 1894 )عائلة زيغوفيلالسي( رايض شاه 1* ، سعاد ج. العربية 2 ، أمرية س.م. الشحية 2 ، انصر س.س. السيايب 2 ، وفاء ك.أ. املعمرية 2 ، مجال ن. الصباحي 3 ، هدى خ. الرقيشية 3 Abstract. Wild plants such as Zygophyllum luntii, from the Zygophyllaceae family, have traditionally been used for medicinal purposes in Oman. The present study investigated (i) the antibacterial activity of the crude extracts (leaves, stem and roots) and the oil (leaves); and (ii) the hydrocarbon contents and fatty acid methyl ester (FAME) components from Z. luntii. These extracts were tested against Escherichia coli, Staphylococcus aureus, and Pseudomonas aeruginosa using well diffusion assays utilizing Muller Hinton Agar (MHA). Antibacterial activity was observed with the Z. luntii leaf extract and significant differences (F=14.06, df=2, P=0.002) were found among E. coli, P. aeruginosa and S. aureus. The highest inhibition was observed against P. aeruginosa, with an inhibition zone of 15.5 ± 2.6 mm, followed by E. coli and S. aureus with inhibition zones of 11.3 ± 1.5 mm and 3.5 ± 4.7 mm, respectively. The Z. luntii extracts showed effec- tiveness within 50-60% against E. coli and P. aeruginosa as compared to Ciprofloxacin. The hydrocarbon contents and the FAME components of the extracts were determined with Gas Chromatography-Mass Spectrometry (GC-MS). A total of 20, 19, and 8 compounds were detected from leaf, stem, and root extracts, respectively. Heneicosane, docosane, and tricosane were found in the highest concentration in the leaves, HOP-22(29)-EN-3.BETA.-OL and β-sitosterol were found in the stems, and docosane and tricosane were found in the roots of Z. luntii. Nine types of fatty acids methyl esters were detected in the oil extracted from leaves with methyl esters of palmitic acid, linolenic acid, and oleic acid constituting 90% of the oil. This is the first report on antibacterial activity and chemical composition of Z. luntii. Keywords: Zygophyllum luntii, Antibacterial activity, Gas Chromatography- Mass Spectrometry (GC-MS), Fatty Acid Methyl Ester (FAME) Zygophyl- املســماة الســقطرية ، مــن عائلــة زيغوفيلالســي Zygophyllum luntii املس��تخلص يتــم اســتخدام النبــااتت الربيــة مثــل زيغوفيللــوم لونــي: laceae بشــكل تقليــدي يف عمــان لألغــراض الطبيــة. حبثــت هــذه الدراســة عــن النشــاط املضــاد للبكتــرياي للمســتخلصات اخلــام مــن األوراق والســاق .Z. luntii واجلذوروالزيــت املســتخلص مــن األوراق. مت الكشــف علــى مركبــات مــن اهليدروكربــوانت واألمحــاض الدهنيــة يف مســتخلص األوراق لنبتــة seudomonas aerugi� و Staphylococcus aureus و Escherichia coli مت اختبــار هــذه املســتخلصات ضــد بعــض األنــواع مــن البكتــرياي مثــل nosa بطريقــة فحوصــات االنتشــار اجليــد إبســتخدام أجــار مولــر هينتــون املناســب لنمــو هــذه األنــواع مــن البكتــرياي. لوحــظ نشــاط مضــاد للجراثيــم مــن P. فقد لوحظ أعلى تثبيط ضد . S. aureus و P. aeruginosa و E. coli ووجدت فروق ذات داللة إحصائية بني Z. luntii مستخلصات نبات aeruginosa مع منطقة تثبيط 15.5 ± 2.6 مم ، تليها E. coli و S. aureus مع مناطق تثبيط 11.3 ± 1.5 مم و 3.5 ± 4.7 مم على التوايل. أظهــرت مســتخلصات Z. luntii فعاليــة يف حــدود 50-60 ٪ ضــد E. coli و P. aeruginosa مقارنــة مــع النمــوذج التجــاري سيربوفلوكساســني. مت تعريــف مركبــات اهليدروكربــوانت واألمحــاض الدهنيــة يف املســتخلصات إبســتخدام جهــاز الفصــل الكروماتوجــرايف الغــازي املرتبــط مبستشــعر الطيــف الكتلــي (GC-MS) . مت الكشــف عــن ماجمموعــه 20 و 19 و 8 مــن املركبــات مــن مســتخلصات األوراق والســاق واجلــذور علــى التــوايل. مت العثــور .3-EN- )29( 22-HOP علــى اهليدركربــوانت هينيكوســان و دوكوســان وتريكوســان كأعلــى تراكيــز للمركبــات يف األوراق ، ومت العثــور علــى مركــب BETA.-OL ومركب بيتا سيستريول يف السيقان ، كما مت العثور على مركبات دوكوسان وتريكوسان يف اجلذور املوجودة يف Z. luntii . مت الكشف عــن تســعة أنــواع مــن األمحــاض الدهنيــة يف الزيــت املســتخرج مــن األوراق مــع نســب عليــا جملمــوع تراكيــز محــض الباملتيــك ومحــض اللينولينيــك ومحــض األوليــك شــكلت مانســبته 90٪ مــن الزيــت. يعتــرب هــذا هــو التقريــر األول علــى مســتوى الدراســات البحثيــة عــن النشــاط املضــاد للبكتــرياي والرتكيــب .Z. luntii الكيميائــي لـــنبات ، (GC-MS) لكلم��ات املفتاحي��ة: زيغوفيللــوم لونــي ، النشــاط املضــاد للبكتــرياي ، جهــاز الفصــل الكروماتوجــرايف الغــازي املرتبــط ابالطيــف الكتلــي (FAME) إســرت امليثيــل الدهنيــة األمحــاض 59Research Paper Shah, Alabri, Ashehi, Asiyabi, AlMamari, AlSabahi, Al-Ruqaishi global market yearly are extracted or isolated from nat- ural resources and about 25% of the prescribed phar- maceutical drugs are based on plant chemicals (Orhan, 2012). Plants are the major source of secondary metab- olites, which are used to treat various diseases (Hossain et al., 2013; Akhtar et al., 2017; Raqiya and Hossain, 2017; Asma et al., 2017; Hossain, 2018; Said et al., 2018). Medicinal plants are found in many places in the world; however, they are found more in tropical regions (Al- Salt, 2012). Family Zygophylllaceae includes many medicinal- ly important plants spices including several species of Zygophyllum. Zygophyllum have antitumor, antioxidant and analgesic properties, and have been used for the treatments of cancer, fever, asthma, urinary discharges, toothache, stomach problems and kidney diseases (Ah- san et al., 2007; Satpute et al., 2009). Zygophyllum spe- cies were found to be potent antifungal and antibacterial agents (Zhang et al., 2008; Gupta et al., 2009) and con- tained many biologically active chemical constituents, such as alkaloids, saponins, terpenoids, sterols, flavo- noids, coumarins and trace elements (Beier, 2005). Zygophyllum luntii distribution is restricted to the Horn of Africa region, including Djibouti, Oman, Soma- lia and Yemen (Beier, 2005). It grows on sand as well as gravel, from sea level up to 1950 m altitude. In Oman, Z. luntii is found in the foot of Dhofar mountains along with several other species of Zygophyllum (Z. bruguieri, schweinfurtii, indica, mahrana and ovalifolia) (Mosti, et al., 2012). The current study explored two objectives. Firstly, an- tibacterial activity of the Z. luntii extract (leaves, stems and roots) and leaf oil against Escherichia coli, Staphylo� coccus aureus and Pseudomonas aeruginosa were inves- tigated. Secondly, hydrocarbon contents of the plant ex- tracts and lipids fatty acid methyl ester were determined (FAME) components of the oil extracted from leaves by gas chromatography- mass spectrometer (GC-MS). Materials and Methods Collection and Preparation of Plant Materials Roots, leaves, and stems of Z. luntii were gathered from the Botanical Garden at Sultan Qaboos University, Oman. These parts were cleaned with tap water followed by distilled water to remove any dust and soil. The plant parts were then further divided into two portions; one was dried in the oven at 70°C for 8 h and then ground into a fine powder and other portion was kept fresh in a refrigerator at 4°C. Plant Extracts Preparation for Anti-bacterial Test The dried powder of the leaves, stem and roots was dissolved in 70% methanol (1:3, w/v) and then kept in a shaker for extraction at room temperature for 24 h. Then, methanol was dissipated from the sample using oven to get the crude extract which was re-suspended in dimethylsulfoxide (DMSO) for application in the an- tibacterial test. Lipid Extraction from Leaves Around 470g of the fresh leaves with 1 L of distilled wa- ter were grinded by a blender. The solution was mixed with the solvent (chloroform: methanol in 2:1 ratio) to separate the lipids and then evaporated in a rotary evap- orator. The extract was filtered through charcoal. The obtained oil (1 g) was kept in storage at 4°C until utiliza- tion for further tests. Anti-bacterial Assay Muller Hinton Agar CM0337 from Oxiod (Part of Ther- mo Fisher Scientific) was used in well diffusion assay. MHA contains beef dehydrated infusion 300.0 g/L, ca- sein hydrolysate 17.5 g/L, starch 1.5 g/L and agar 17.0 g/L. Amount of 38 g of MHA was suspended in 1 L of distilled water, boiled and sterilized by autoclaving at 121°C for 15 min. Three strains of pathogenic bacteria E. coli ATCC 25922, P. aeruginosa ATCC 27853, and S. aureus ATCC 25923 were utilized as test microorganisms. These clini- cal isolates were obtained from Microbiology Laboratory at Sultan Qaboos University Hospital. These strains are for antibiotic testing and fall under the American type collection culture (ATCC). Furthermore, these strains were sub cultured in liquid broth for a period of 6 to 8 hours. The well diffusion assay was conducted utilizing Muller Hinton Agar (MHA). The assay of leaves, stem and roots extract activity was carried out in nine MHAs plates which were replicated three times. For every plate, four discs were used one each for leaves, stem, roots and an antibiotic standard (Ciprofloxacin) as positive con- trol. Discs of 6 - 8 mm diameter were removed from agar with a sterile glass pasture pipette and filled with 30 μl of the sample extract or standard. At the same time, three MHAs plates were used for oil using a similar procedure where three discs were used for every plate; two discs for oil and one disc for antibiotic standard. Zone inhibition was investigated after incubating agar plates at 37°C. Sample Preparation and Extraction for GC-MS Analysis Fresh leaf, stem and root samples were weighed and grinded using a mechanical grinder. Then, 50 mL of 70% methanol was added to each grinded sample and placed in an ultrasonic water bath working at 50–60 kHz with power of 350 W for 30 min at room temperature. By us- ing a rotary vacuum evaporator, the methanol was evap- orated and the extracts were concentrated. The crude extracts were dissolved in hexane, filtered by microfil- tration (0.45 µl syringe) and injected to GC-MS. 60 SQU Journal of Agricultural and Marine Sciences, 2020, Volume 25, Issue 2 Antibacterial Activity and Chemical Composition of Crude Extract and Oil of Zygophyllum (Fagonia) luntii (Baker) 1894 (Family Zygophyllace- ae) Perkin Elmer Clarus 600 GC System, fitted with a Rtx- 5MS capillary column (30 m × 0.25 mm i.d. × 0.25 μm film thickness; maximum temperature, 350°C), coupled to a Perkin Elmer Clarus 600C MS. Ultra-high puri- Gas Chromatography-Mass Spectrometry (GC/ MS) Analysis GC-MS conditions for samples extracted from leaves, stem and roots: GC-MS analysis was performed on a Figure 1. Comparison of anti-bacterial activity (inhibition zone in mm) among Escherichia coli ATCC 25922, Staphylococ� cus aureus ATCC 25923 and Pseudomonas aeruginosa ATCC 27853 in well diffusion assays exposed to Zygophyllum luntii extracts and Ciprofloxacin. Bars designated by the same letters are not statistically significant at α0.05 Figure 2. Comparison of anti-bacterial activity (inhibition zone in mm) of the Zygophyllum luntii extracts and Ciproflox- acin against Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853 and Staphylococcus aureus ATCC 25923 well diffusion assays. Bars designated by the same letters are not statistically significant at α0.05 61Research Paper Shah, Alabri, Ashehi, Asiyabi, AlMamari, AlSabahi, Al-Ruqaishi ty helium (99.9999%) was used as carrier gas at a con- stant flow of 1.0 mL/min. The injector, transfer line and ion source temperatures were 280°C, 270°C and 270°C, respectively. The ionizing energy was 70 eV. Electron multiplier (EM) voltage was obtained from auto tune. All data were obtained by collecting the full-scan mass spectra within the scan range 40-550 amu. The injected sample volume was 1 μl with a split ratio of 10:1. The oven temperature program was 60°C at a rate of 80°C per minute to 280°C hold for 25 minutes. The total run time was 53.5 minutes. GC/MS conditions for the oil extracted from leaves: Fatty Acid Methyl Ester (FAME) compounds were de- tected by the GC-MS equipment as mentioned above. The helium gas flow rate was 0.7 mL/min. The injector, transfer line and ion source temperatures were 250°C, 250°C and 220°C, respectively. The initial oven tempera- ture was set at 50°C (holds for 8 minutes) and increased to 250°C in a rate of 40°C per minute. All data were ob- tained by collecting the full-scan mass spectra within the scan range 35-500 amu. The unknown compounds were identified by comparing the spectra obtained with mass spectrum libraries (NIST 2011 v.2.3 and Wiley, 9th edition). Statistical Analysis Data on antibacterial assays from leaves, stems and roots, and the three bacterial strains were analysed sep- arately using one-way analysis of variance (ANOVA) in Past (https://past.en.lo4d.com/windows). Means were separated by Dunn’s Multiple Comparison Test and dif- ferences were considered significant at p<0.05. Table 1. Compounds identified in the leaf, stem and root ex- tracts of Zygophyllum luntii by GC-MS Name of compound Retention Time (min) Percent (%) Leaves Pentadecane 13.64 0.33 Tetradecane, 2,6,10-trimethyl- 16.43 0.68 Oxalic acid, allyl octyl ester 16.81 0.32 Unidentified 17.02 0.61 Octadecane 17.79 0.34 Nonadecane 18.73 0.90 Eicosane 19.02 5.84 Heneicosane 21.57 12.30 Docosane 22.60 12.72 Tricosane 23.26 12.93 Tetracosane 24.24 9.94 Pentacosane 25.36 10.42 Hexacosane 26.21 7.63 Heptacosane 27.13 5.84 Octacosane 27.87 5.82 Nonacosane 28.70 5.03 Triacontane 29.62 4.10 Hentriacontane 30.71 1.84 Dotriacontane 31.99 2.42 Tetratriacontane 36.24 0.13 Stem Hexadecanal 18.33 2.60 Phytol 21.50 5.22 Unidentified 22.22 2.70 Unidentified 23.20 0.38 Unidentified 24.20 0.27 Unidentified 25.18 0.47 Heptacosane 26.90 0.35 Octacosane 27.80 0.22 Supraene 28.14 0.95 Unidentified 28.60 0.46 Triacontane 29.50 0.29 Hentriacontane 30.64 0.38 Vitamin E acetate 31.40 1.24 Dotriacontane 31.93 0.64 Name of compound Retention Time (min) Percent (%) Tritriacontane 33.40 1.23 β-Sitosterol 34.36 11.02 Heptatriacontane 35.40 0.80 Hop-22(29)-En-3.Beta.-Ol 35.90 69.95 Octatriacontane 37.70 0.84 Roots Docosane 22.60 14.04 Tricosane 23.49 14.36 Tetracosane 24.43 12.54 Pentacosane 25.36 4.02 Pentacosane 25.54 5.33 Hexacosane 26.24 6.67 Octacosane 27.13 43.05 Tetratriacontane 36.24 1.15 62 SQU Journal of Agricultural and Marine Sciences, 2020, Volume 25, Issue 2 Antibacterial Activity and Chemical Composition of Crude Extract and Oil of Zygophyllum (Fagonia) luntii (Baker) 1894 (Family Zygophyllace- ae) Result and Discussion Antibacterial Assays Significant antibacterial activity was observed in the Z. luntii leaves extract and significant differences (F=14.06, df=2, P=0.002) were found among E. coli, P. aerugino� sa and S. aureus (Figure 1). The highest inhibition was observed against P. aeruginosa (15.5 ± 2.64 mm) then E. coli (11.25 ± 1.50mm) and S. aureus (3.50±4.72 mm). The inhibition zones of the stem (F=2.03, df=2, P=0.187) and root (F=0.68, df=2, P=0.530) extracts did not dif- fer significantly among the three bacterial strains. Oil from leaves did not produce any inhibition in E. coli but had significantly higher inhibition (F=229.4, df=2, P<0.001) in P. aeruginosa (9.5±0.58mm) and S. aureus (6.75±0.96mm). Ciprofloxacin had significantly dif- ferent inhibition zones (F=73.3, df=2, P<0.001) among the three bacterial strains. The highest inhibition zone by the commercial antibiotic was against P. aeruginosa (30.75± 0.96 mm) then E. coli of (25.0± 0.82 mm) and lowest against S. aureus (19.50±1.91 mm). The inhibition in the stem, root and leaves extracts, and oil from leave was significantly lower than Cipro- floxacin against E. coli (F=29.8, df=4, P<0.001), P. aerugi� nosa (F=11.7, df=4, P<0.001) and S. aureus (F=9.9, df=4, P<0.001) (Figure 2). The stem extract was more active against P. aeruginosa while root extract was more active against S. aureus. Leaves extract showed more activity against E. coli. The oil showed some antibacterial activ- ity against P. aeruginosa and S. aureus but not E. coli. Overall, the Z. luntii extracts were active against both Gram-positive (S. aureus) and Gram-negative bacteria (E. coli and P. aeruginosa), though they were more active against the latter. The ethanol extracts of intact leaf of Z. arabica showed an inhibition zone of only 6.08 mm against E. coli and did not inhibit the growth of S. aureus (Alam et al., 2010). The ethanol whole plant extract of F. cretica produced inhibition zones of 15 mm, 15 mm and 14 mm against E. coli, P. aeruginosa and S. aureus, re- spectively (Sajid et al., 2011) which are similar to our re- sults. The crude extract of Z. arabica from Sinai showed broad antimicrobial spectrum against Gram-positive, Gram-negative, spore-forming and acid-fast bacteria (El-Hefnawi, 1999). It is obvious that leaves extracts from Zygophyllum spp including Z. luntii have antibac- terial properties. The percent effectiveness of the Z. luntii extracts were compared to Ciprofloxacin against E. coli, P. aeru� ginosa and S. aureus by subtracting the extract inhibi- tion zone sizes from the Ciprofloxacin (Figure 3). The calculated percent effectiveness against E. coli was 47.0±4.8%; against P. aeruginosa was 57.1±8.1%; and against S. aureus was 19.4±24.2%. The percent effective- ness against E. coli and P. aeruginosa was significantly higher compared to S. aureus (F=10.54, df=2, P=0.005). Alam et al. (2010) produced callus of Z. arabica by tissue culture and found that the callus extract was more effec- tive against Serratia marcescens, E. coli and Acetobacter aceti subsp. liquefaciens (inhibition zones = 32.67, 33.92 and 34.83 mm respectively) than the crude extract of the intact leaf (IZ=6.08 mm) suggesting higher antibacterial effects of callus extract against Gram – ve bacteria. Several pathogens are increasingly developing resis- tance, particularly to broad-spectrum antibiotics (Kunin, 1993). Resistant E. coli isolates have been reported from humans using disk diffusion method against ciproflox- acin (22% with the highest of 52% reported from Iran), cefotaxime (31.2%–58%) and ceftazidime (10%–57.4%) (Pormohammad et al., 2019). Some of the gram-positive drug resistant bacteria include S. aureus, Streptococcus Table 2. Compounds detected in the oil extracted from Zygo� phyllum luntii leaves by GC-MS Name of compound Retention Time (min) Percent (%) Leaves Caprylic acid methyl ester 11.58 0.12 Capric acid methyl ester 18.27 0.19 Lauric acid, methyl ester 24.29 0.74 Myristic acid, methyl ester 29.83 0.81 Palmitic acid, methyl ester 34.93 34.55 Palmitoleic acid, methyl ester 35.90 1.70 Oleic acid, methyl ester 39.78 23.99 Linoleic acid, methyl ester 40.57 5.81 Linolenic acid, methyl ester 41.73 32.09 Figure 3. Percent effectiveness of the Zygophyllum luntii extracts compared to Ciprofloxacin against Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 25923 and Pseudomonas aeruginosa ATCC 27853, and S. aureus well diffusion assays. Bars designated by the same letters are not statistically significant at α0.05 63Research Paper Shah, Alabri, Ashehi, Asiyabi, AlMamari, AlSabahi, Al-Ruqaishi Figure 4. Chromatogram of compounds in leaves (A), stem (B) and roots (C), and fatty acids methyl esters (FAME) in oil (D) extracted from Zygophyllum luntii leaves detected by GC/MS. 64 SQU Journal of Agricultural and Marine Sciences, 2020, Volume 25, Issue 2 Antibacterial Activity and Chemical Composition of Crude Extract and Oil of Zygophyllum (Fagonia) luntii (Baker) 1894 (Family Zygophyllace- ae) pneumoniae, and Enterococcus spp., and the gram-neg- ative drug resistant bacteria Acinetobacter baumannii, Klebsiella pneumoniae, E. coli, and P. aeruginosa (Lister et al., 2009). Our results also indicated reduced sensi- tivity of S. aureus and E. coli to Ciprofloxacin. The Z. luntii callus extract may help in managing antibacterial resistant pathogens of different strains Gas Chromatography-Mass Spectrometry (GC/ MS) Analysis Higher number of compounds was identified in the leaves (n=20) and stem (n=19) extract compared to roots (n=8) extracts of Z. luntii while 6 compounds could not be identified (Table 1). The retention time (RT) of all compounds varied between 13.64 to 37.7 minutes. The unidentified compounds were present in a relatively low amount. Heneicosane, Docosane and Tricosane were present in higher quantities in the leaves extract. Hop- 22(29)-En-3.Beta.-Ol (69.95%) was present in higher quantities in the stem extract while the root extract had octacosane in high quantities. Palmitic acid, linolenic acid and oleic acid were the main components of oil ex- tracted from leaves (Table 2). Heptacosane, Heneicosane, Tetradecane have been reported with antimicrobial activity (Elshiekh and Ab- delmageed, 2015). Beta-acids are an important compo- nent of hops (Humulus lupulus L. family Cannabaceae) soft resins and usually isolated as by-products during hop processing (McCallum et al., 2019). Hexahydro-β acids showed strong antibacterial activity and good sta- bility (Liu et al., 2019). Noticeably, HOP-22(29)-EN-3. BETA.-OL had the highest percentage compared to oth- er compounds. Close retention time of 22-27 minutes in roots com- pounds showed their similar affinity to stationary phase. Both retention time and peak area (%) for Docosane (C22) and Tricosane (C23) (22.60 minutes and 23.26 minutes, and 12.72% and 12.93%, respectively) did not show wide variation. These two compounds were pres- ent in leaves and roots but not in stems. The n-alkane fractions (hydrocarbons C22-C35) was detected in the leaves extract of Z. luntii which have been detected in vegetable oils by GC/MS (Troya et al., 2015). These com- pounds are more common in plant extracts. Species of Zygophyllum have been found to contain saponins (Abdel-Khalik et al., 2001), alkaloids (Sharawy and Alshammari, 2009), terpenoids (Perroni et al., 2007), sterols (Shoeb et al., 1994), flavonoids (Ibrahim et al., 2008), proteins and amino acids (Sharma et al., 2010), coumarins (Alam et al., 2010) and trace elements (Fati- ma et al., 1999). The presence of such chemical ingredi- ents in Zygophyllum spp. Would contribute to the medi- cal properties, including stimulating the immune system in humans, treating and preventing the development of chronic diseases (Beier, 2005), and the vitality to resist such types of pathogenic bacteria used in this study. Oil Extract from Leaves Nine different types of fatty acids methyl esters (FAME) were found in oil extracted from Z. luntii leaves. Pal- mitic acid, Linolenic acid, and Oleic acid were the main compounds and represented 90% of the oil (Fig.4). There was no study found on Z. luntii that explored the fatty acid content. However, seven fatty acids including the Oleic acid, Palmitic acid and Linoleic acid were found in other Zygophyllum species e.g. F. arabica L (Alam et al., 2010) and F. cretica (Soad, 1994). Conclusion Extracts from leaves, stem, and roots of Z. luntii had sig- nificant antibacterial activity against E. coli, P. aerugino� sa and S. aureus. The extracted oils had activity against only P. aeruginosa and S. aureus. The commercial antibi- otic Ciprofloxacin had reduced activity against S. aureus and E. coli. The Z. luntii extracts showed about 50-60% effectiveness against E. coli and P. aeruginosa com- pared to Ciprofloxacin. HOP-22(29)-EN-3. BETA.-OL, Hexahydro-β acids, and FAME compounds could have contributed to the antibacterial activity. With further research on callus production, improving extraction process and antimicrobial activity assays against more pathogenic bacterial (including antibiotic resistant) spe- cies, Z. luntii can be promoted as a source of traditional medicine in Oman Acknowledgement The corresponding author is grateful to postgraduate students at Sultan Qaboos University (SQU) for the GC/ MS analysis, Ruqiya Al Hattali and Hibatallah Al Hab- si from the central laboratory from Animal Health for assistance with antibacterial test, Microbiology Labora- tory at Sultan Qaboos University Hospital for providing clinical isolates and Dr. Annette Patzelt from Oman Bo- tanic Garden, Diwan of Royal Court for identification of Zygophyllum luntii. References Abdel-Khalik SM, Miyase T, Hanan EA, Melek FR. (2000). Triterpenoid saponins from Fagonia cretica. Phytochemistry 54: 853-859. Ahsan H, Muhammad Z, Bushra M. (2007). Cytotoxic and antitumor potential of Fagonia cretica L. Turkish Journal of Biology 31(1): 19-24. Akhtar SM, Hossain MA, Sadri SA. (2017). Isolation and characterization of antimicrobial compound from the stem-bark of the traditionally used medicinal plant Adenium obesum. Journal of Traditional Com- plementary Medicine 7: 296–300. Alam EA, Amin GH, ElAyouty YM, Abdel-Hady MS. (2010). Chemical composition and antibacterial ac- 65Research Paper Shah, Alabri, Ashehi, Asiyabi, AlMamari, AlSabahi, Al-Ruqaishi tivity studies on callus of Fagonia arabica L. Academ- ic Arena 2(12): 91-106. Al-Salt, J. (2012). Antimicrobial activity of crude extracts of some plant leaves. Research Journal of Microbiol- ogy 7: 59-67. Asma HS, Moza TG, Hossain MA. (2017). Brine shrimp toxicity of various polarities leaves and fruits crude fractions of Ziziphus jujuba native to Oman and their antimicrobial potency. Sustainable Chemical Phar- macology 5: 122-125. Beier BA. (2001). Itvo new unifoliolate species of Fago- nia (Zygophyllaceae) from the Horn of Africa region, and the resurrection of F. subinermis from Iran. Nor- diac Journal of Botany 21(5): 449-455. Beier BA. (2005). A revision of the desert shrub Fagonia (Zygophyllaceae). Systematics and Biodiversity 3(3): 221-263. Bobbarala V. (2015). Concepts, Compounds and the Alternatives of Antibacterials. InTech Open. DOI: 10.5772/59522. eBook. El-Hefnawi HN. (1994). Screening of some Sinai plants for their antimicrobial activity. Al-Azhar Journal of Microbiology 43: 1-6. Elshiekh YH, Abdelmageed MAM. (2015). Gas chro- matography-mass spectrometry analysis of Pulicaria crispa (whole plant) petroleum ether extracts. Amer- ican Journal of Research Communication 3(3): 58-67. Fatima K, Khaula S, Kalhoro MA, Muhammad Q, Yas- meen B. (1999). Trace elements in indigenous medic- inal plants (Rhazya stricta, Vinca rosea and Fagonia cretica). Phytochemistry 42(4): 182-183. Gupta V, Sharma S, Josef I, George M. (2009). Analgesic and antimicrobial activities of Fagonia indica. Phar- macology online 3. Hossain MA. (2018). A Review on Adenium Obesum: A Potential Endemic Medicinal Plant in Oman. Be- ni-Suef University Journal of Basic and Applied Sci- ence 7 (4): 559–63. Hossain MA, AL-Mijizy ZH, Al-Rashdi KK, Weli AM, Al-Riyami Q. (2013). Effect of temperature and ex- traction process on antioxidant activity of various leaves crude extracts of Thymus vulgaris. Journal of Coastal Life Medicine 1 (2): 118-122. Ibrahim LF, Kawashty SA, El-Hagrassy AM, Nassar ML, Mabry TJ. (2008). A new kaempferol triglycoside from Fagonia taeckholmiana: cytotoxic activity of its extracts. Carbohydrate Research 343(1): 155-158. Kunin CM. (1993). Resistance to antimicrobial drugs a worldwide calamity. Annals of Internal Medicine 118:557–561. Lister PD, Wolter DJ, Hanson ND. (2009). Antibacteri- al-Resistant Pseudomonas aeruginosa: clinical im- pact and complex regulation of chromosomally en- coded resistance mechanisms. Clinical Microbiology Review 22(4): 582–610. Liu Y, Lu N, Tang J. (2019). Synthesis, characterization, crystal structure, and antioxidant activity of hexa- hydro-β-acids. Journal of Molecular Structure 1175: 721–727. Martins E. (2013). The growing use of herbal medicines: issues relating to adverse reactions and challenges in monitoring safety. Frontiers in Pharmacology 4: 177- 179. McCallum JL, Nabuurs MH, Gallant ST, Kirby CW, Mill AA. (2019). Phytochemical characterization of wild hops (Humulus lupulus ssp. lupuloides) germplasm resources from the maritimes region of Canada. Frontiers in Plant Science 10:1438. Mosti S, Raffaelli M, Tardelli M. (2012). Contribution to the flora of central-Southern Dhofar (Sultanate of Oman). Webbia 67(1): 65-91. Orhan IE. (2012). Biotechnological production of plant secondary metabolites. Bentham e-book, pp. 107- 120. Oyebode O, Kandala NB, Lilford RJ. (2016). Use of tradi- tional medicine in middle-income countries: a WHO- SAGE study. Health Policy Plan 38(8): 984-991. Perroni A, Masullo MA, Basarello C, Hamed AI, Belisa- rio MA, Pizza C, Piacente S. (2007). Journal of Natu- ral Products 70(4): 584-588. Pormohammad A, Nasiri MJ, Azimi T. (2019). Prev- alence of antibiotic resistance in Escherichia coli strains simultaneously isolated from humans, ani- mals, food, and the environment: a systematic re- view and meta-analysis. Infect. Drug Resistance 12: 1181–1197. Raqiya MSM, Hossain MA. (2017). Evaluation of anti- oxidant and cytotoxic activities of different extracts of folk medicinal plant Hapllophyllum tuberculatum. Egyptian Journal of Basic Applied Science 4: 101-106. Said MA, Hossain MA, Ahmed AA. (2018). Antimicro- bial and cytotoxic comparative study of different ex- tracts of Omani and Sudanese Gum acacia. Beni-Suef University Journal of Basic and Applied Science 7: 22-26. Sajid B, Alia E, Rizwana K, Uzma S, Hafiz MI. (2011). Phytochemical screening and antimicrobial activi- ty of Fagonia cretica plant extracts against selected microbes. Journal of Pharmacology Research 4:962– 963. Satpute RM, Kashyap RS, Deopujiari JY, Taori GM, Dag- inawala HF. (2009). Protection of PC12 cells from chemical ischemia induced oxidative stress by Fag- onia arabica. Food and Chemical Toxicology 47(11): 2689-2695. Sharawy SM, Alshammari AM. (2009). Checklist of poi- 66 SQU Journal of Agricultural and Marine Sciences, 2020, Volume 25, Issue 2 Antibacterial Activity and Chemical Composition of Crude Extract and Oil of Zygophyllum (Fagonia) luntii (Baker) 1894 (Family Zygophyllace- ae) sonous plants and animals in Aja Mountain, Ha’il re- gion, Saudi Arabia. Australian Journal of Basic and Applied Science 3(3): 2217-2225. Sharrma S, Gupta V, Sharma G. (2010). Phytopharma- cology of Fagonia Indica (L): A Review. Journal of Natural Conscience 1(1): 143-147. Shoeb HA, Sharada MM, El-Sayed LAR, El-Wakeel E. (1994). Triterpenoid and sterol glycosides from Fag- onia arabica L. Al-Azhar Journal of Pharmaceutical Science 13: 41-48. Soad MA. (1994). Chemical and Biological studies of some Fagonia species (Family Zygophylllaceae). PhD Thesis 1994, Department of Pharmaceutical Scienc- es, Faculty of Pharmacy, Cairo University, Egypt. p 16-47. Troy F, Lerma-García MJ, Herrero-Martínez JM, Simó-Alfonso EF. (2015). Classification of vegetable oils according to their botanical origin using N-al- kane profiles established by GC–MS. Food Chemis- try 167: 36–39. Zhang W, Krohn K, Draeger S, Schulz B. (2008). Bioac- tive isocoumarins isolated from the endophytic fun- gus Microdochium bolleyi. Journal of Natural Prod- ucts 71(6): 1078-1081.