INTRODUCTION:

Zingiber officinale Rosc have a place with the family Zingiberaceae, It has a long history of therapeutic use going back 2500 years in China and India , for example, cerebral pains, sickness, ailment, anticancer and antioxidant agent impacts, and colds¹. The English botanist William Roscoe named the plant 'Zingiber officinale' in 1807. The name 'Zingiber' is by means of the Greek word 'zingiberis' got from the Sanskrit word 'shringavera', which signifies 'molded like a deer's tusks'; 'officinale' shows therapeutic properties of the plant Zingiber officinale². Ginger is a typical spice for various nourishments and drinks³. Zingiber officinale Rosc is usually pharmaceutical items yet can likewise be utilized in beauty care products and everyday requirements⁴.

umerous reports are accessible on the compound synthesis of ginger oil, The chemical composition of the ginger oils was investigated by GC and GC-MS the major components of ginger oil were zingiberene (46.71%), citronellyl n-butyrate (19.34%), valencene (7.61%) β-phellandrene (3.70%), β-funebrene (3.09%), camphene (2.59%), α-pinene (1.09%) and selina-4(14), 7(11)-diene (1.03%)³. ethanolic extract from the rhizome of Zingiber officinale were more sensitive to Gram-positive bacteria: Staphylococcus aureus and Staphylococcus aureus ATCC 25923.⁵ Our study was conducted to identify the compounds of ginger (Zingiber officinale Rosc) by preparing an ethanolic extract by sonication method that subjected for separation and identification of these compounds by using GC-Q-TOF technique and focus on the existed studies on their biological activity. This study will assist to distinguishing the compounds of remedial importance.

MATERIALS AND METHODS:

Reagents and chemicals:

Chemicals used in this research with the highest purity were Ethanol absolute purchased form (ROMIL-SpS™ Super Purity Solvents) and N-Hexane purchased from (AVONCHEM).

Plant material:

The rhizome of Ginger was purchased from the local vendor and dried at 37℃ for 24 h where exposure to sunlight was avoided to prevent the loss of active components, the plant material was ground in a grinding machine (IKA, Staufen, laboratory mill, Type A 10, screen size 0.5mm Speed max. [rpm] 2000, Germany).

Preparation of plant extract:

5gm of Ginger (Zingiber officinale Rosc) were powdered, placed in glass container then mixed with ethanol with ratio 1:10 (w/v) of dry weight to solvent⁶, labelled and extracted by ultrasound assisted technique (Sonication Method) at 60°C for 30 mints⁷ using a 20 kHz ultrasonic generator (Misonix S3000; Misonix Inc., New York, NY, USA). The mixture was filtered through a Whatman No. 4 filter paper where the obtained ethanolic extract was centrifuged (P SELECTA) at 5000rpm for 10 min, the supernatant was collected, filtered through 0.2μm filters (Sarstedt, Germany)and stored at -30°C until use⁸.

GC-Q-TOF Analysis:

Taken 2ml of filtrate of ethanol plant extracts, the filtrates were evaporated to dryness under gentle stream of nitrogen at 40°C (Alliance Nitrogen evaporator) then reconstitute the dryness filtrate by 2ml of hexane then stirring by vortex mixer to dissolve the dried parts of plant extract (vortex mixer, KMC -1300V) and 1μl of Sample injected to vials of GC -Q -TOF finally.

The GC-Q-TOF analysis using HP-5MS ultra inert column with: 30m × 250µm × 0.25µm film thickness, Helium was used as carrier gas. Injector and detector temperatures were 325°C. An Agilent Technology gas chromatograph type 7250 Accurate-Mass Quadrupole Time-of-Flight GC/MS System. The 7250 Q-TOF GC/MS is equipped with three ion vacuum gauges: Vacuum manifold chamber, TOF vacuum manifold chamber and Turbomolecular vacuum pumps exhaust. To Condition a Capillary Column: Materials needed: Carrier gas, (99.9995% pure or better), Wrench, open-end, 1/4-inch and 5/16-inch (8710-0510). The 7250 Accurate-Mass Q-TOF GC/MS can operate the LE-EI ion source in standard or low energy modes. The column oven temperature was programmed as follows: start temperature at 60ºC hold for one minute; increased to 120ºC with a ramp of 40ºC/min with run time 2.5 minutes, increased to 310ºC with a ramp of 5ºC/min with run time 40.5 minutes. After 15 s the split valves were opened for 3 min to purge the injector.

Identification of components:

Components were identified via Mass Hunter Workstation Software, Qualitative Analysis Navigator, version B.08.00 service pack 1build 8.0.8208.38. Interpretation of mass spectrum of GC-Q-TOF was done using database of National Institute Standard and Technology (NIST). The mass spectrum of unknown component was compared with the spectrum of the known component stored in the NIST library.

RESULTS AND DISCUSSION:

forty-eight compounds were identified in Zingiber officinale Rosc by GC-Q-TOF analysis. The active principles are presented in Table 1 and Figure 1. They were identified by computerized library (NIST) using the GC-Q-TOF spectra at different retention times where the name, probability, molecular formula, molecular weight, CAS and peak area for each peak were investigated. The mass spectrometer analyzes the compounds eluted at diverse times to distinguish the nature and structure of the compounds. The large compound parts into small compounds giving rise to the appearance of peaks at diverse m/z proportions. These mass spectra are unique mark of that compound which can be recognized from the data library⁹.

Nineteen of these compounds has been reported as biological activity as show in Table 2. the most important identified compounds due to their biological activity were 3-Allyl-6-methoxyphenyl acetate(0.82%), Clovene(0.19%), 10S, 11S-Himachala-3(12),4-diene(3.57%), Cubebol(2.01%), Zingiberone(4.12%), α-Isonootkatol(0.23%), α-Vetivol(0.53%), Cyclodecasiloxane, eicosamethyl-(0.63%), (±)-[6]-Gingerol(1.03%), [6]-Gingerdiol 3,5-diacetate(7.77%), Methyl diacetoxy-[6]gingerdiol(4.30%), Barringenol R1(1.12%),[10]-Shogaol (5.80%),1.1,1,3,3,5,5,7,7,9,9,11,11,13,13,15,15-Hexadecamethyloctasiloxane # (0.26%),[6]-Gingerdiol (2E)-geranial acetal(2.76%), Vitamin E acetate(0.53%), Campesterol (1.52%), Stigmasterol (1.71%) and γ-Sitosterol(4.46%). Fig (2) shows the mass spectrum and structures of therapeutic important phytochemical constituents of ethanolic ginger extract.

Comparative to this study, Cloven were characterized through Column chromatography (CC) analysis total ethanolic extract of G. parvifl ora¹⁰ and characterized using GC and GC-MS analysis of the seeds of Arnebia linearifolia, gingerols and shogaols were characterized through HPLC analysis of ethanolic extract of Ginger¹¹, (6)-Shogaol were characterized through HPLC analysis of Extract from Ginger (Zingiber officinale Roscoe)¹², Campesterol and β-Sitosterol were characterized through GC-MS analysis of Rice Bran Extract¹³.

Figure (1): GC-Q-TOF Analysis of Ethanolic Extract of Ginger (Zingiber officinale Rosc)

Table 1: Identified Compounds of Ethanoic Extract Ginger by Gc-Q-Tof Analysis

Peak No	RT	Compound name	Prob	Molecular formula	CAS	MWT	Area%
1.	6.25	Cholestan-26-oic acid, 3,7,12-trihydroxy-, (3α,5β,7α,12α)-	28.7%	C₂₇H₄₆O₅	547-98-8	450	0.15%
2.	7.79	3-Allyl-6-methoxyphenyl acetate	22.3%	C₁₂H₁₄O3	1941-09-9	206	0.82%
3.	8.44	Clovene	31.5%	C₁₅H₂₄	469-92-1	204	0.19%
4.	9.17	Clovene	30.2%	C₁₅H₂₄	469-92-1	204	0.34%
5.	10.04	(E)-Eremophila-1(10),7(11)-dien-12-yl acetate	31.3%	C₁₇H₂₆O₂	352461-71-3	262	2.63%
6.	10.86	10S,11S-Himachala-3(12),4-diene	8.61%	C₁₅H₂₄	60909-28-6	204	3.57%
7.	11.36	1R,4R,7R,11R-1,3,4,7-Tetramethyltricyclo[5.3.1.0(4,11)]undec-2-ene	7.93%	C₁₅H₂₄	137235-59-7	204	1.38%
8.	12.12	Cubebol	9.39%	C₁₅H₂₆O	23445-02-5	222	1.11%
9.	12.59	Cubebol	9.71%	C₁₅H₂₆O	23445-02-5	222	2.01%
10.	13.45	Zingiberone	21.4%	C₁₁H₁₄O₃	122-48-5	194	4.12%
11.	14.14	1,3,5-Trimethoxy-2-propenylbenzene	6.23%	C₁₂H₁₆O₃	5273-94-9	208	1.70%
12.	15.28	Phenol, 2-methyl-5-(1,2,2-trimethylcyclopentyl)-, (S)-	31.1%	C₁₅H₂₂O	38412-84-9	218	0.92%
13.	15.71	α-Isonootkatol	36.2%	C₁₅H₂₄O	1380573-94-3	220	0.23%
14.	16.48	3α,9β-Dihydroxy-3,5α,8-trimethyltricyclo[6.3.1.0(1,5)]dodecane	9.53%	C₁₅H₂₆O₂		238	2.24%
15.	16.83	α-Vetivol	11.8%	C₁₅H₂₄O	57422-86-3	220	0.53%
16.	17.70	8-Amino-2,6-dimethoxylepidine	14.8%	C₁₂H₁₄N₂O₂	74509-65-2	218	2.13%
17.	18.09	Ethyl 3,4-dimethoxy-N-(3-phthalimidopropionyl)-dl-phenylalaninate	26.9%	C₂₄H₂₆N₂O₇		454	0.77%
18.	19.28	Bis[Methyl(pentamethylene)]disiloxane	46.5%	C₁₂H₂₆OSi₂	18035-52-4	242	0.77%
19.	20.68	Verticillol	16.7%	C₂₀H₃₄O	70000-19-0	290	0.49%
20.	21.64	1,4-Naphthalenedione, 3-acetyl-2,5,7-trihydroxy-	40.0%	C₁₂H₈O₆	54725-01-8	248	0.99%
21.	22.16	17α-Ethynyl-17β-hydroxy-5β-estran-3-one	15.3%	C₂₀H₂₈O₂	28044-91-9	300	2.50%
22.	22.55	Cyclodecasiloxane, eicosamethyl-	58.9%	C₂₀H₆₀O₁₀Si₁₀	18772-36-6	740	0.63%
23.	23.05	Tributyl aconitate	94.1%	C₁₈H₃₀O₆	7568-58-3	342	2.34%
24.	23.73	Benzo[a,i]quinolisin-5-one, 1,2,3,5-tetrahydro-6-cyclohexyl-7-hydroxy-	51.7%	C₁₈H₂₁NO₂	59315-08-1	283	1.11%
25.	24.64	Ethyl 3,4-dimethoxy-N-(3-phthalimidopropionyl)-dl-phenylalaninate	26.9%	C₂₄H₂₆N₂O₇		454	3.90%
26.	25.39	Podocarpa-1,8,11,13-tetraen-3-one, 14-isopropyl-1,13-dimethoxy-	11.1%	C₂₂H₃₀O₃	18326-20-0	342	12.51%
27.	26.25	Podocarpa-1,8,11,13-tetraen-3-one, 14-isopropyl-1,13-dimethoxy-	20.1%	C₂₂H₃₀O₃	18326-20-0	342	1.54%
28.	26.81	(±)-[6]-Gingerol	69.0%	C₁₇H₂₆O₄	39886-76-5	294	1.03%
29.	27.46	2,4-Diamino-5,6-decamrthylenethieno[2,3-d]pyrimidine	13.1%	C₁₆H₂₄N₄S	42159-82-0	304	3.65%
30.	28.26	6-Hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid, ethyl ester	16.4%	C₁₆H₂₂O₄		278	1.41%
31.	28.64	trans-1-(p-(Methylthio)phenyl)-1-dodecen-3-one	21.1%	C₁₉H₂₈OS	123044-46-2	304	6.68%
32.	28.98	[6]-Gingerdiol 3,5-diacetate	89.5%	C₂₁H₃₂O₆	143615-75-2	380	7.77%
33.	29.34	Methyl diacetoxy-[6]gingerdiol	88.4%	C₂₂H₃₄O₆	53254-52-7	394	4.30%
34.	29.93	Bauer-7-en-3-one	13.1%	C₃₀H₄₈O	6895-55-2	424	0.45%
35.	30.66	[10]-Shogaol	55.9%	C₂₁H₃₂O₃	36752-54-2	332	2.47%
36.	31.42	Barringenol R1	36.8%	C₃₀H₅₀O₆	15399-43-6	506	0.42%
37.	31.78	[10]-Shogaol	34.9%	C₂₁H₃₂O₃	36752-54-2	332	5.80%
38.	32.38	9-Ethyl-3-hydroxy-10,11b-dimethyl-1,2,3,4,6,6a,6b,7,8,9,11a,11b-dodecahydrobenzo[a]fluoren-11-one	17.4%	C₂₁H₃₀O₂		314	0.47%
39.	32.25	Barringenol R1	15.9%	C₃₀H₅₀O₆	15399-43-6	506	1.12%
40.	33.72	1,1,3,3,5,5,7,7,9,9,11,11,13,13,15,15-Hexadecamethyloctasiloxane #	33.2%	C₁₆H₅₀O₇Si₈	19095-24-0	578	0.26%
41.	34.66	1H-Cyclopropa[3,4]benz[1,2-E]azulene-3-carboxaldehyde, 9a-(acetyloxy)-1a,1b,4,4a,5,7a,7b,8,9,9a-decahydro-4a,7b-dihydroxy-1,1,6,8-	20.4%	C₂₂H₂₆O₇	77573-49-0	402	0.30%
42.	35.47	1.1,1,3,3,5,5,7,7,9,9,11,11,13,13,15,15-Hexadecamethyloctasiloxane #	23.5%	C₁₆H₅₀O₇Si₈	19095-24-0	578	0.45%
43.	36.34	Alizarin Yellow GG, O,O'-di(trimethylsilyl)-	10.9%	C₁₉H₂₅N₃O₅Si₂		431	0.82%
44.	36.84	[6]-Gingerdiol (2E)-geranial acetal	69.3%	C₂₇H₄₂O₄	863913-68-2	430	2.76%
45.	37.37	Vitamin E acetate	19.6%	C₃₁H₅₂O₃	58-95-7	472	0.53%
46.	38.56	Campesterol	65.7%	C₂₈H₄₈O	474-62-4	400	1.52%
47.	39.96	Stigmasterol	22.1%	C₂₉H₄₈O	83-48-7	412	1.71%
48.	39.67	γ-Sitosterol	90.7%	C₂₉H₅₀O	83-47-6	414	4.46%

Table 2: Biological Activates of Constituents of The Ethanoic Extract of Ginger

S. No	Compound name	Biological Activities
1.	3-Allyl-6-methoxyphenyl acetate	Anticancer action ¹⁴
2.	Clovene	Antibacterial and antifungal activity.¹⁵, insecticide¹⁶
3.	10S,11S-Himachala-3(12),4-diene	Antibacterial activity¹⁷
4.	Cubebol	Dietary supplement and flavoring agent¹⁸
5.	Zingiberone	Strong anti-inflammatory, antidiabetic, antilipolytic, antidiarrhoeic, antispasmodic, and so forth properties. Other than, it shows the property of enhancing growth and immune stimulation¹⁹
6.	α-Isonootkatol	Insecticide ¹⁶
7.	α-Vetivol	Insecticidal ,herbicidal and antioxidant activities¹⁶
8.	Cyclodecasiloxane, eicosamethyl-	Antibacterial²⁰
9.	(±)-[6]-Gingerol	Antioxidant,anti-inflammatory, anticancer, antiemetic and other bioactivities²¹
10.	[6]-Gingerdiol 3,5-diacetate	The antioxidant activity.²²
11.	Methyl diacetoxy-[6]gingerdiol	Antioxidant activity.²²
12.	Barringenol R1	Anti-tumour, anti-Alzheimer's disease, anti-inflammatory,anti-microbial, anti-obesity and anti-allergic activities²³
13.	[10]-Shogaol	Antioxidant activity.²²
14.	1.1,1,3,3,5,5,7,7,9,9,11,11,13,13,15,15-Hexadecamethyloctasiloxane #	Anti-inflammatory ²⁴
15.	[6]-Gingerdiol (2E)-geranial acetal	anti-inflammatory activity²⁵
16.	Vitamin E acetate	Antioxidant ²⁶
17.	Campesterol	Anti-inflammatory impact and cytotoxic against a few cancer cell line²⁷
18.	Stigmasterol	Anti-inflammatory impact and cytotoxic against a few cancer cell line²⁷
19.	γ-Sitosterol	Antidiabetic medicate ²⁸&²⁹

Figure 2: Mass Spectrum and Structure of Constituents of The Ethanoic Extract of Ginger Have Biological Activates

CONCLUSION:

This study has helped to identify the compounds of Ethanoic extract of ginger. The presence of bioactive compounds legitimizes the utilize of the ginger (Zingiber officinale) in remedy for nausea, stomach pain, and other health issues. The present study aimed at identifying the nature of the components responsible for their antioxidant, Anti-inflammatory and antibacterial activity by characterization of active compounds in ginger (Zingiber officinale) using GC-Q-TOF technique.

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Received on 17.07.2020 Modified on 29.08.2020

Research J. Pharm. and Tech. 2021; 14(8):4307-4313.

DOI: 10.52711/0974-360X.2021.00748