INTRODUCTION:

Ginger (Zingiber Officinale Roscoe), a rhizome of the plant, is one of the world's most precious herbs as medicine. It belongs to the Zingiberaceae family. It is also used as a spice to enhance taste and flavour making food products delicious. In ancient times, fresh ginger has been used as food, medicine, and dietary supplements. Ginger has many functional health benefits such as food antioxidants, and good food for curing different diseases.

The rhizome ginger is a rich source of vitamins, minerals, carbohydrates, other proximate compositions, and bioactive compounds which have to be essential to care for different diseases such as inflammation.^1,2

Ginger root and its main phenolic compounds such as gingerols and zerumbone have anticarcinogenic activity, antioxidant, and anti-inflammatory activity.³

Gingerol is an active component of ginger. It is further divided into three categories i.e., 6-gingerol, 8-gingerol, and 10-gingerol. They have many pharmacological effects. In the Series of gingerol, 6-gingerol has a more convertible mechanism of kinetics in extraction by using ethanol. In fresh ginger, there is the percentage of bioactive compounds like gingerol is 96% - 97%, and shogaol is 3% - 4%. However, the gingerol is changed over into shogaol [pungent constituents of ginger similar in chemical structure to gingerol] when applying heat. It is because of thermal degradation as at high temperatures they are not stable. Different extraction methods can be used to get oils or extract from the rhizome of ginger. The old extraction technique includes steam distillation, hydro-distillation, maceration, Soxhlet extraction, and empyreumatic distillation. The latest and new technologies are environmentally friendly i.e., supercritical fluid extraction and subcritical extraction methods although these extraction methods can be performed by use of water for extraction purposes.⁴

Figure 1. Molecular targets of ginger and its active constituents against gastrointestinal cancer⁵

Chewing Gum also has proven great value as a delivery vehicle for pharmaceutical and nutraceutical ingredients. It can be taken discreetly without water and the drug is allowed for local and systemic delivery. It can be employed for the treatment of diseases of the oral cavity and throat e.g., for caries prevention, or it can release drugs that can be absorbed through oral mucosa directly into the systemic circulation. In addition, drug that is not absorbed by the oral cavity membranes can be dissolved in the saliva before swallowing, thus leading to a more rapid onset of action.

Figure 2. Medicated chewing gum

The Medicated Chewing Gums (MCG) can also be utilized for site-specific activity in case of oral cancer. Chewing gum is going to advance more and more in nowadays research and it seems to get more standardized in the future industry because it can deliver either pharmaceuticals or nutrients which are known as medicated chewing gum (MCG) and NonMCG. MCG is supposed to act as an extended-release dosage form that provides a continuous release of medicine contained.⁶

Drug Profile:

Below table 1 gives detailed information about ginger.⁷

Table 1. Drug profile of Zingiber officinale (ginger)

Drug name	Zingiber officinale (ginger)
Structure
IUPAC Name	(5S)-5-Hydroxy-1-(4-hydroxy-3-methoxyphenyl) decan-3-one
Mol. formula	C₃₅H₅₂O₆
Mol. weight	568.8 g/mol
Melting Point	30-32 ℃
Description	White Powder
Solubility in water	The solubilities of the ginger bioactives are lower than 0.0001 at temperatures lower than 130 °C
Pharmacognosy	Ginger is used as an antiemetic, positive inotropic, spasmolytic, aromatic stimulant, carminative, condiment, and flavoring agent. It is prescribed for dyspepsia, flatulent colic, and vomiting spasms, as an adjunct to many tonics and stimulating remedies, for painful affections of the stomach, cold, cough, and asthma.
Molar Mass	294.38 g/mol
Adverse Effect	Heartburn, Diarrhoea, Burping, etc

Table 2. Active Chemical Constituents of Ginger⁸

Phenols

Volatile Sesquiterpenes

Others

Gingerols and Shogoals

Bisapolene

Zingiberene

Zingiberol

Sesquiphellandrene

Curcumin

6-dehydrogingerdione

Galanolactone

Zingerone

Geraniol

Neral

Gingerglycolipids

Table 3. Anticancer Activity of Ginger and its Compounds⁹

Sr. No.	Compound Name	Type of cancer	Mechanism	Cell System
1	Beta Elemene	Nonsmall cell lung cancer cells	Release of cytochrome	In vitro
2	6-gingerol	Ovarian cell	Inhibition NF-kB	In vitro
3	Ginger extract	Liver cancer	Reduced the elevated expression of TNF alpha and NF-kB	Rats
4	Zerumbone	Lung and Colon cancer	Suppresses modulatory mechanism of growth and reduces the expression of NF-kB	Mouse

MATERIALS AND METHODS:

Chemicals and Reagents:

Ginger extract (Gingerol), Ascorbic Acid, Beeswax, PVP, Dextrose, Peppermint oil, PEG -400, Calcium carbonate

Instruments and Apparatus:

Calibrated weighing balance, Burner, Mortar and Pestle, Measuring Cylinder, Spatula, and Pippete.

Method of Preparation:

Isolation of Gingerol from Ginger:

Dry ginger was crushed to a coarse powder and extracted with 95% ethanol by a simple maceration process. The solvent was evaporated by distillation to obtain a thick pasty mass. The thick pasty mass was suspended in water. The Ginger resin precipitates in water which was removed by filtration and the residue obtained was dried under a vacuum.¹⁰

Preformulation Study:

Evaluation of Ginger extract (Gingerol)

Determination of Solubility:

Solubility is made by adding solute in small incremental amounts to a fixed volume of solvents such as distilled water, ethanol, chloroform, acetone, and ether. Then it is examined for undissolved particles.

UV-VIS Spectrophotometric method:

Methanol was selected for the preparation of the calibration curve. 100mg of crude extract was dissolved in methanol and diluted up to 100ml to get the concentration of 1000ppm which is treated as stock solution. This stock solution was diluted further to get different concentrations. Resultant solutions were scanned for λmax in the range of 200‐400 nm using a UV spectrophotometer.

IR spectrum interpretation:

Fourier transform infrared (FTIR) was used to identify the characteristic functional groups in the extract. A small quantity (5 mg) of the extract was dispersed in dry potassium bromide (KBr). The mixture was thoroughly mixed in a mortar and pressed at a pressure of 6 bars within 2 min to form a KBr thin disc. Then the disc was placed in a sample cup of a diffuse reflectance accessory. The sample was scanned from 4000 - 400 cm^-1.

Formulation of Chewing Gum^11:

Table 4. Table for the formulation of chewing gum

Sr. No.	Ingredients	F1	F2	F3	F4	F5
1	Gingerol	4.0mg	4.0mg	4.0mg	4.0mg	4.0 mg
2	Ascorbic Acid	0.2g	0.2g	0.2g	0.2g	0.2g
3	Beeswax	1.0g	1.0g	1.0g	1.0g	1.0g
4	PVP	4.5g	5.0g	5.5g	6.0g	5.4g
5	Dextrose	0.7g	0.8g	0.9g	1.0g	0.6g
6	Peppermint oil	0.5ml	0.5ml	0.5ml	0.5ml	0.5ml
7	PEG-400	0.8g	1.0g	1.2g	1.2g	0.7g
8	Calcium Carbonate	0.5g	0.5g	0.5g	0.5g	0.5g
9	Methyl Paraben	0.5mg	0.5mg	0.5mg	0.5mg	0.5mg
10	Propyl Paraben	0.5mg	0.5mg	0.5mg	0.5mg	0.5mg

PEG: Polyethylene glycol, HCL: Hydrochloride, PVP: Polyvinyl pyrrolidine

Preparation of Chewing Gum¹²:

Chewing gum was prepared by direct compression mold method.

1. In this method, each ingredient was weighed accurately and separately.

2. Gingerol PVP, beeswax, dextrose, calcium carbonate, peppermint oil, and ascorbic acid, all ingredients were thoroughly mixed in ascending order of their weights in a mortar.

3. After proper mixing, ingredients were smoothly grounded in a mortar pestle, and then a previously weighed quantity of PEG-400 was added.

4. Then, the whole mixture was again mixed thoroughly in a pestle mortar.

5. After mixing and grinding, the mixture was subjected to compression into the desired molds and pressed to form medicated chewing gum.

6. From mold, formulated chewing gums were weighed and wrapped properly.

Figure 3. Preparation of chewing gum

Evaluation Parameter of Chewing Gum:

Physical Appearance:

All the batches were visually evaluated for physical appearance, color, odour, and taste. The texture study was performed manually by pressing the gum between the thumb and the finger. The texture feel was characterized by sticky, good, or solid mass.¹³

Weight uniformity test:

For single dosage units, 20 samples of chewing gum were selected randomly and the average weight was calculated, not more than 2 samples should differ from the average weight. The maximum deviation from the average weight should be 5%.¹⁴

Uniformity of content:

10gums from each formulation F1 to F5 were randomly selected and weighed. Each gum separately was dissolved in 50ml of chloroform. After they were dissolved completely, 100 ml of phosphate buffer with adjusted pH of 6.8 were added to extract the gingerol in the aqueous Phase.¹⁵

Elasticity Study:

The elasticity of chewing gum is one of the important parameters. Appropriate elasticity of chewing gum contributes to increasing patient compliance as well as the proper release of the drug. The elasticity of F1 to F5 was determined using a CT3 texture analyzer (Brookfield Engineering Inc., USA). To determine the elasticity, the TA3/100 probe and fixture of TA-RTKI were used for the study. The tension was selected as a test mode at the target of 80mm. Trigger load and load cell were kept at 10 kg and 10,000g, respectively. Chewing gum of size1×1 mm² was randomly selected and was fixed between the two clamps of probe TA3/100. The lower clamp was held stationary and chewing gum was pulled apart by the upper clamp. It was pulled and stretched till the chewing gum broke, losing its elasticity. The force required at this point was recorded. Data collection and calculations were performed using texture-pro CT V1.3build 14 software.

Stickiness:

On a plain surface, chewing gum was placed; it is subjected to collide with a Teflon hammer with a mass of 250g for a period of 10min. hammering frequency was 30/min. After a specified time, the amount of mass stuck to the hammer was observed and reported.

Chewing Study of Gum Formulation:

The chewing gum should provide a good mouth feel and comfort during chewing without sticking to the teeth. However, the amount of saliva secreted contributes mainly to chewing the formulation. Three male and three female volunteers were asked to chew the placebo chewing gums (without drug) F1 to F5 for 1 min without swallowing saliva. The outcome of the chewing process provided by volunteers has been categorized accordingly as good, bad, and sticky.

In vitro drug release studies:

A chewing gum was immersed in a pH solution maintained at pH 6.8 (buccal cavity) then placed on a magnetic stirrer and subjected to stirring every 5 min interval 2ml of solution was taken out and replaced with fresh buffer solution. The sample was withdrawn at regular intervals of 5, 10, 15, 20, 25, and 30 min. On completion of the process, all the collected samples were UV spectrophotometrically analyzed on a maximum wavelength of 310nm.

Drug release study in saliva:

The release of drug from chewing gum in saliva was studied by recruiting a panel of six members of volunteers and designed chewout studies. One sample was given to each volunteer for chewing for a particular time interval period, i.e., 0.5, 1, 2, 5, 10, and 15min. After chewing, chewed out chewing gum samples were collected from volunteers, stretched maximum, and cut into small pieces after that dispersed in a 100ml volumetric flask having phosphate buffer pH 6.8, which was then heated and sonicated for 10 min. These samples were then analyzed by UV spectrophotometer at absorption maxima 310nm for residual drug content in chewing gum.

Amount of drug release during mastication = The total drug content –Residual drug after chewing.

Stability Study:

The stability study of chewing gum was studied to obtain a stable product that assures safety and efficacy, till shelf life, at defined storage and package conditions. A stability study was done according to ICH guidelines to assess the combined effect of the drug, gum base, and excipients on the stability of the formulation. Optimized formulation was placed in vials and stored in a stability chamber (Thermolab, India) at 30°C±2°C/65% RH±5% RH. The samples were evaluated for the color, taste, drug content, in vitro drug release study, and growth of microorganisms after 7, 15, and 30days and 3months.

RESULTS AND DISCUSSION:

Pre-formulation Study:

Organoleptic Parameter:

The below table shows the results of the organoleptic study.

Table 5. Table showing organoleptic study results

Parameters	Results
Color	Light Brown
Taste	Pungent and Sweet
Odour	Characteristic
State	Solid

Solubility study:

The below table shows the results of the solubility study

Table 6. Table showing solubility study results

Solvent	Solubility behavior
Water	Insoluble
Acetone	Insoluble
Ethanol	Soluble
Methanol	Soluble
Chloroform	Soluble

UV Analysis:

After studying the UV- spectra of ginger rhizome extract, it was found that it shows maximum absorbance at 249.4 nm. So, absorbance at 249.4 nm was considered as λmax for Ginger extract.

IR analysis:

The identity of the drug was confirmed by comparing the IR spectrum of the drug with the reported spectrum of Gingerol. The characteristic absorption bands in the infrared absorption spectrum of Gingerols and a summary of the description of the characteristic IR bands for gingerol are described.

Table 7. Table showing IR interpretation of ginger extract

Sr. No.	Functional Group	Theoretical Peaks (cm^-1)	Practical Peaks (cm^-1)
1	C-H	2700-3300	2900, 282.5
2	C-O	900-1300	1009.5
3	N-H	3100-3500	3390.5
4	O-H	3000-3700	3599.5
5	C=O	1600-1900	1643.5
6	C=C	1475-1610	1418.5

Evaluation parameter of finished products:

Physical appearance:

Table 8. Table showing organoleptic parameter

Batch Code	Color	Odour	Texture
F1	Light brown	Characteristic	Solid mass
F2	Light brown	Characteristic	Very Good
F3	Light brown	Characteristic	Good
F4	Light brown	Characteristic	Sticky
F5	Light brown	Characteristic	Sticky

Table 09. Physical Evaluation for Finished Chewing Gum

Formulation Code	F1	F2	F3	F4	F5
Weight uniformity test(gm)	6.22	6.56	6.67	6.90	6.79
Uniformity of contents	93.1± 5.7%	95.5 ± 4.5%	92.6 ± 4.4%	94.6 ± 3.8%	93.7± 4.3%
111Elasticity Study(mm)	10.7	12.55	7.6	11.6	10.5
Stickiness	Solid mass	Very Good	Good	Sticky	Sticky
Chewing study of gum formulation	+	++	11+	-	-
In Vitro drug release test	96.2%	96.7%	96.5%	96.3%	96.1%
Drug release study in saliva	52.4±1.23	53.66± 1.62%	52.20± 1.73%	52.11± 1.65%	52.60± 1.45%

++:- Very Good, +:- Good, - :- Sticky

Weight Uniformity Test:

Weight variation determination is a pharmacopoeia test for the evaluation of weight uniformity of all chewing gums but as described in European Pharmacopoeia (EP) this experiment was performed on 20 randomly selected gums from formulations with the best volume. The average weight was calculated and the range of their weights was 8.204gm to 10.404gm.

Uniformity of Content:

The mean drug content of 10gums that were randomly selected from each series of formulations F1 to F5 was 93.1±5.7%, 95.5±4.5%, 92.6±4.4%, 94.6±3.8%, and 93.7± 4.3% of the claim. All results satisfied the criteria which are commonly required by USP for solid dosage forms.

Elasticity Study:

The elasticity of F1 to F5 was determined by using a CT3 texture analyzer. F2 showed an increase in elasticity.

Stickiness:

All formulations showed negligible stickiness.

Chewing Study of Gum Formulation:

The outcomes of chewing of placebo formulations (F1 toF5) expressed by three male and three female volunteers areas. All six volunteers’ highlighted the F2 formulation as possessing a very good texture feel, good mouthfeel, and chewing easiness.

In Vitro Drug Release Study:

In vitro drug release, it was found that almost all formulations release more than 96% drug after 30 min. However, the optimized formulation F2 has shown the best release in comparison to other formulations. Hence, it was selected for further study.

Drug Release Study in Saliva:

It was observed that within 15 min; more than 50% of the drug was released from the optimized formulation F2. This study revealed that the drug release depended on the chewing frequency of the volunteer.

Stability Study:

The formulation was found to be stable for 3 months at accelerated conditions. There was no change in colour and taste after the periodic removal of samples (7, 15, and 30 days and 3 months). The drug content of F2 was found to be reduced from 97.2% to 96.6%, which was within the acceptable range. Drug release did not get much affected and reduced from 94% to 93.4% within 16 min and was found to be within the acceptable range. No growth of microorganisms was observed after the completion of an accelerated stability study of 3 months.

CONCLUSION:

Herbal Chewing Gum was successfully prepared. Gingerol is potent and reduces the expression of NF-kB having important anticancer activity and good tolerability. It is a cost-effective formulation and has better patient compliance and bioavailability. Gingerol is completely absorbed by GIT which makes this a choice of drug in preparing Herbal chewing gum Ginger.¹⁶ By considering the results of our study, the ginger hydroalcoholic extract obtained from maceration has beneficial active compounds that help in various types of cancer. Ginger chewing gum with efficient active ingredients can be formulated based on this study. The release of the active agent is acceptable as the gum nearly passed the EP protocols. Best formulations contain no extra flavoring agents. The mixture of different sweeteners provides a better taste and sense. According to organoleptic properties, formulation F2 is considered to be the best among others.

In vitro drug release study indicated that drug release from formulation F2 is best among others so it is the best formulation than others. All the parameters were found to be satisfactory; hence, the therapeutic dose of Gingerols can be given in Herbal chewing gum with optimized formula, i.e., F2. This study concluded that it is possible to make herbal chewing of Gingerol for the prevention and treatment of Cancer.

List of Abbreviations:

MCG: - Medicated Chewing Gums

MTT: - (3-[4, 5-dimethylthiazol-2-yl]-2, 5 diphenyl tetrazolium bromide) assay

PEG: Polyethylene glycol

HCL: Hydrochloride

PVP: Polyvinyl pyrrolidine

N-f-K B: - Nuclear factor kappa B

STAT 3:- Signal transducer and activator of transcription 3

MАРК: - Mitogen-activated protein kinase

PT 3K: - Phosphoinositide 3-kinase

ERK: - Extracellular Signal regulated kinase

AKt: - AK strain transforming.

TNF alpha: - Tumor Necrosis factor alpha

Cox-2:- Cyclooxygenase -2

Cdk: - Cyclin-dependent kinases

MMP-9:- Matrix Melka proteinase

CIAP-1:-Cellular inhibitor of apoptosis protein.

XTAP: - Expandable Transient Analysis program

BCL 2:- B-cell leukemia/lymphoma 2

HPLC:-High performance liquid chromatography

FTIR: - Fourier Transform infrared.

UV-Spectra: - Ultraviolet Spectroscopy

IR: - Infrared

KBr: - Potassium Bromide

CT 3 ·Texture analyzer: - Compression and tension testing analyzer

TA3/100 probe: - Temperature analyzer probe

TA RTKI:-Rotary Base table

ACKNOWLEDGMENT:

The authors were thankful to Vidyabharati College of Pharmacy, Amravati for providing the necessary infrastructural facilities to carry out research work.

DISCLOSURE STATEMENT:

No potential conflict of interest was reported by the authors.

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Received on 01.03.2023 Modified on 08.05.2023

Asian J. Pharm. Ana. 2023; 13(4):278-284.

DOI: 10.52711/2231-5675.2023.00046