FORMULATION AND INVITRO EVALUATION OF ETHOSOMAL …

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Chistia et al. World Journal of Pharmacy and Pharmaceutical Sciences

FORMULATION AND INVITRO EVALUATION OF ETHOSOMAL

GELS CONTAINING ANTIDIABETIC DRUGS

Sd. Riyaz Hussain Chistia*1, Dr. Pawan kumar

2 and Dr. Parwez Alam

3

1Research Scholar,

2Professor,

3Professor

Department of Pharmaceutical Sciences, Singhania University, Pacheri Bari, Jhunjhunu,

Rajasthan – India.

ABSTRACT

The process Touitou et al (2000) defined was used with little change to

prepare various ethosomal formulations with different levels of IPA

(20% to 40%) and sonicity. It was simple and reproducible techniques.

The prepared and disacrete ethosomes have been developed. However

ethosomes are more uniform in size and small, necessary for skin

penetration by the process of sonication. When the efficiency of the

trap was compared, ethosomes containing 30 percent w/w IPA, which

were generated by sonication, displayed the highest value with respect

to all others; Therefore, with 30 percent w/w IPA as the best formula

for all other aspects, ethosomally provided by sonicity was completed.

In all formulations, GF6 demonstrated full release of the drugs in 1,440

min compared to other formulations. The in vitro release decreased

with increased concentrations of polymer and copolymer. Centered on the research findings

of the drug release process the drug discharge followed the non-fickian diffusion mechanism

by formulations and followed the first order kinetics.

KEYWORDS: Ethosomes, Sonication, Transdermal, Entrapment, Stability.

INTRODUCTION

Controlled Drug Delivery Systems

A managed drug supply system has been developed to track drug delivery speeds, maintain

therapeutic duration and/or tissue delivery. drug delivery systems were developed.

A convenient four types of controlled drug delivery or modified drug deliveries are divided.

WORLD JOURNAL OF PHARMACY AND PHARMACEUTICAL SCIENCES

SJIF Impact Factor 7.632

Volume 10, Issue 7, 2062-2099 Research Article ISSN 2278 – 4357

*Corresponding Author

Sd. Riyaz Hussain Chistia

Research Scholar,

Department of

Pharmaceutical Sciences,

Singhania University,

Pacheri Bari, Jhunjhunu,

Rajasthan – India.

Article Received on

21 May 2021,

Revised on 11 June 2021,

Accepted on 01 July 2021,

DOI: 10.20959/wjpps20217-19438


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1. Delayed release.

2. Sustained release.

3. Site-specific targeting.

4. Receptor targeting.

Controlled delivery can be described more precisely as.[1]

(1) Sustained drug action by sustaining a comparatively stable, efficient body level of

prescription drugs with concomitant reduction of undesirable side effects.

2) Spatial location of controlled discharge systems adjacent to or in the tissue of illness,

localized drug activity.

3) Targeted action of drugs to supply the medication to a specific target cell type using

carriers or chemical derivatives.

4) Have a prescription release mechanism based physiologically and therapically. In other

words, physiological and clinical requirements of the body decide the volume and the rate of

release of drugs".[2]

"Usually, a regulated method for the delivery of drugs is built in particular to supply the

medication. Blood levels are kept stable and efficient for a time when the machine is

distributing medication. Regulated drug entry typically leads to significantly constant active

ingredient blood levels compared to uncontrolled fluctuations when several doses of rapid

release are administered to patients for the traditional dosage types.

Currently, an oral route is the most common method of drug delivery. While the benefit of

ease of administration is noteworthy, it also has significant disadvantages, namely poor

bioavailability, due to metabolism in the first place and a propensity to generate high and low

blood spikes, leading to a need for high or regular dosing, which can be both prohibitive as

well as inconvenient".[2]

"The development of an emergent drug delivery system, enhancing the therapeutic

effectiveness and drug protection, by making spatial and temporary placements within the

body more reliable (i.e. precise site), reduces both size and number of doses, is required in

order to solve these problems. New drug delivery schemes are also necessary in order to

supply the site without any major immunosupply or biological inactivation with novel

genetically engineered medicinal products (i.e. peptides, proteins). In addition to these

benefits, pharmaceutical firms understand that the idea and technologies of a managed drug


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delivery system and the cost associated with getting new drug companies into the market are

feasible for repatenting successable drugs. Transdermal delivery of medicinal substances

through the skin for systemic effects was one of the most commonly used methods.[2]

"The present study is equipped with two different polymer combinations: E RS100 and

HPMC E 15, E RL 100 with HPMC E 15, to establish a suitable matry style transdermal drug

delivery systems for Ketorolac. The acrylic acid matrices E RL100 and E RS 100 are used to

manufacture drug polymer matrix films, which are stated to be compatible with several drugs.

Penetration changes that can help increase drug permeation partitioning.[3]

Various D-

Limonene[4]

penetration enhancers, Oleic[5]

and their effects on drug permeation, were used at

various concentrations in the study”.[4]

DRUG DELIVERY SYSTEMS OF TRANSDERMAL

Definition

“Transdermal drug delivery systems are topical medicines in the form of patches delivering

medicines for a predetermined systemic impact and controlled pace. - And controlled rate.

A transdermal drug delivery system, 'which can be of an active or passive nature, is an

alternate medication path. These devices can be administered through the skin barrier to

pharmaceutical products. A medicine is administered to the skin of a patch at a reasonably

high dose over a long period of time. The drug reaches the bloodstream directly through the

skin through a dissemination process. The substance is still diffused in the blood"4 for a long

period of time because the patch is high and the blood concentrations are low, and the drug

persists in blood.

Advantages[6]

The transdermal route is a "interesting option," since it is a realistic and secure transdermal

route. The beneficial characteristics of drug delivery through the skin are systemic

consequences.

1. “Avoidance of first pass metabolism.

2. Avoidance of gastro intestinal incompatibility.

3. Predictable and extended duration of activity.

4. Minimizing undesirable side effects.

5. Provides utilization of drugs with short biological half-lives.

6. Improving physiological and pharmacological response.


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7. Avoiding the fluctuation in drug levels.

8. Avoiding inter and intra patient variations.

9. Maintain plasma concentration of potent drugs.

10. Termination of therapy is easy at any point of time.

11. Greater patient compliance due to elimination of multiple dosing profile.

12. Ability to deliver drug more selectively to a specific site.

13. Provide suitability for self-administration and enhance”[4]

therapeutic efficacies.

Limitations

1. “The drug that requires “high blood levels cannot be administered.

2. Not suitable for drugs with high molecular weight.

3. Not suitable for drugs that undergo metabolism during the passage through the skin.

4. TDDS cannot deliver ionic drugs.

5. TDDS cannot deliver drugs in a pulsatile fashion.

6. Cannot develop TDDS, if drug or formulation causes irritation to skin.

7. Variation of absorption rate based on site of application skin type and patient age.

8. Along with these limitations the high cost of the product is also a major drawback for the

wide acceptance of this”[4]

products.

Table 01: Ideal properties of drug candidate for transdermal drug delivery.[7]

“Parameter Properties

Dose Should be low (<20 mg/day)

Half-life in hour 10 or less

Molecular weight <1000

Partition coefficient Log P (octanol-water) between-1.0 and 4

Skin permeability coefficient >0.5 × 10−3

cm/h

Skin reaction Non irritating and non-sensitizing

Oral bioavailability Low

Therapeutic index Low” 2

Mechanism of absorption

Drug delivery routes across human skin

“Three potential routes can penetrate drug "molecules in contact with the skin surface:

through sweats, via hair follicles and sebaceous glands or directly through the state

corneum (collectively called the shunt or Appendageal route)”.[2]

Over the years the scientific community has been discussing the value of the shunt or

appendage route against the transport through the stratum corneum and complicated


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further by the lack of an appropriate experimental model allowing for three routes to be

segregated.

Hydrated skin or epidermal membranes prefer to be used in in vitro studies, so that

hydration-related swelling closes the attachments. Schedule and other collaborators

suggested the viability of the polar molecules and the flow of large Polar molecules or

ions which diffuse difficultly through a follicular shuttle through the intact stratum

corneum.

However, because the annexes cover a fractional permeation region of about 0.1%, they

are widely agreed to make a marginal contribution to the steady flow of most drugs. This

concept led to most of the strategies for enhancing skin penetration focusing on increased

transport over the corneal stratum rather than through the appendices. Exceptions are the

provision of iontophoresis that uses electric charges for driving molecules into the skin,

mostly via shunt paths, as they provide less electrical strength and vesicular supply.

The stratum corneum consists of 10 to 15 coenocyte layers with a thickness of around 10

to 15 μm in dry condition and a hydration thickness of 40 μm. A "brick and mortar,"

multi layered in structure, consisting mainly of long chain ceramides, free fatties (free

fatty acids), triglycerides, cholesterol and sterol-wax esters and a multi-layered coenocyte

matrix, composed in the shape of a multi-layered structure. It is therefore important to

understand this model in the light of the fact that it is polygonal, elongated and flat (0.2-

1.5 μm, 34-46 μm diameter) coenocytes are not brick-formed.

The intercellular lipid matrix is formed by keratinocytes that discharge their lamellar

content into the intercellular space in the middle to upper part of the stratum granulosum.

This extruded material rearrange into massive intercellular lipid lamels in the initial layers

of the stratum corneum, which are connected with the aligned hydrocarbon chains and the

polar head groups dissolved into an aquatic layer.

The lipid phase behavior is different from that of other organic membranes due to the

stratum Corneum lipid composition. In regions of crystalline, lamellar gel and lamellate

liquid crystal phases, the hydrocarbon chains are structured producing different areas

within the fiber bilayer. Intrinsic and extrinsic proteins such as enzymes also can

influence the stratum corneum's lamellar structure. Water is an important component of a

stratum corneum that acts as a plastizer to prevent the stratum corneum cracking and also

helps produce a natural moisturizer factor (NMF). The prevailing route of medicamental

permeation within the cornea stratum needs to be identified in order to understand how


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the physicochemical properties of the diffusing drug and vehicle affect penetration

through the stratumcorneum and maximize distribution.

In the Aqueous areas beyond the intracellular filaments of keratin (intracellular or

transcellular) it has been historically assumed that hydrophilic chemical substances spread

through the fluid matrix, while lipophilic chemicals diffuse between filaments”.[3]

Transdermal permeation kinetics

“Awareness of skin permeation kinetics is important if transdermal therapeutic systems are to

succeed. Transdermal drug permeation comprises the following steps:

1. Sorption by stratum corneum.

2. Penetration of drug through epidermis.

3. Uptake of the drug by the capillary network in the dermal papillary layer. This

permeation can be possible only if the drug possesses certain physiochemical properties”.

The permeation rate is determined throughout the skin.[8]

dQ/dt =Ps (Cd – Cr)

“Where Cd and Cr are the concentration of the skin penetrating in the donor body, on the

stratum cornea surface and the receptor body respectively. Ps is the overall coefficient of

permeability of the skin tissue. The relationship decides this permeability coefficient”.[6]

Ps =Dss Ks / hs

Here Ks is the partition coefficient for separating the penetrating molecule from either the

solution medium or transdermal to the stratum corneum, Dss indicates an apparent diffusivity

of a skin tissue thickness to the permanent state diffusion of the penetrating molecule and hs

the total thickness of skin tissue. The permeability coefficient Ps for skin penetrators can be

regarded as being constant under some conditions as “Ks, Dss and hs are. From equation it is

clear that a constant rate of drug permeation can be obtained only when Cd >> Cr i.e. the

drug concentration at the surface of the stratum corneum Cd is consistently and substantially

greater than the drug concentration in the” 6

body Cr. The equation becomes.

dQ/dt = Ps Cd

The rate of “skin permeation is constant provided the magnitude of Cd remains fairly

constant throughout the course of skin permeation. For keeping Cd constant, the drug should

be released from the device at a rate Rr i.e. either constant or greater than the rate of skin

uptake Ra I.e. Rr>> Ra. Since Rr>>Ra , the drug concentration on the skin surface Cd is

maintained at a level equal to or greater than the equilibrium solubility of the drug in the


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stratum corneum Cs .i.e. Cd>>Cs. Therefore, a maximum rate of skin permeation is obtained

and is given by the” equation.[6]

(dQ/dt) m = PsCs

From “the above equation it can be seen that the maximum rate of skin permeation depends

upon the skin permeability coefficient Ps and is equilibrium solubility in the stratum corneum

Cs. Thus skin permeation appears to be stratum corneum”[7]

limited.

Mechanism of penetration

"While speculation still concerns the exact process of medicinal supply by ethosomes, a

combination of processes probably contributes to the improvement of the effect. The multi-

layer stratum corneum lipid is tightly packaged and highly conformational at physiological

temperature. The high ethanol content makes ethosomes special, since ethanol is known to

disrupt the organization of the bilayer of the skin lipid. It thus enables the vesicles to

penetrate a streatum corneum when integrated into a vesicles membrane. The lipid membrane

is packed less securely than traditional vesicles due to its high ethanol content, but has an

equal strength that allows a structure that can be more malt-free and allows more mobility,

squeezing through small squares, such as openings created to disrupt the lipid stratum

corneum".[9]

“Ethanol interacts with lipid molecules in the hard group polar region, which decreases the

stiffness of the corneal stratum lipids and increases their fluidity. The intercalation of ethanol

into the environment of the polar head group will lead to increased membrane permeability.

The ethosomes itself can interact with the stratum corneal barrier as well as the effect of

ethanol upon the layered corneal structure”.[10]


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Figure 01: Proposed model of skin delivery ethosomal system.

The “interdigital and malleable vesicle of ethosomes will trace paths in the distorted stratum.

The higher positive zeta-potential of the drugs can increase the skin attachment of the

vesicles in the case of drug encapsulation ethosomes. The ethosomal system was shown to be

highly efficient carriers for increased drug use in skin while encapsulated medication in

classic liposomes remained primarily at the skin's surface. This method is a promising

candidate for transdermal supply of the medication because of its efficient supply along with

the long-term stability of the ethosomes.[11]

"

Planning Preparation

“The Touitou4 et al. reports that the ethosomal system can be produced with soya been

phosphotidyl choline 2 – 5 percent, 20 – 50 percent w/w, and medications and waters, at 100

percent W/w. "Ethosomal formulation and preparation is the result of Touitou4 et al. Ethanol

dissolves 90 and medication for the preparation of ethosomes phospholipon. As a slender

stream with a steady mix at 700 rpm, double distilled water was added slowly into a well

settled bottle. For another 5 minutes, mixing has been continued. During the preparation, the

machine was kept at 30 °C and then stored cold”.[12]


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Different Additives Employed in Formulation of Ethosomes.

Table 02: Different additives used in ethosomal formulation.

Class Example Uses

Phospholipid

“Soya phosphatidyl choline

Egg phosphatidyl choline

Dipalmityl phosphatidyl choline

Distearyl phosphatidyl choline.”[11]

Vesicles forming

component.

Polyglycol Propylene glycol,

Transcutol RTM

As a skin penetration

enhancer.

Alcohol Ethanol,

Isopropyl alcohol

“For providing the softness

for vesicle membrane.

As a penetration enhancer.”

12

Cholesterol Cholesterol For providing the stability

to vesicle membrane.

Dye

“Rhodamine-123,

Rhodamine red,

FluoresceneIsothiocynate (FITC),

6- Carboxy fluorescence.”[12]

For characterization study.

Vehicle Carbopol 934 As a gel former.

GLIBENCLAMIDE

Description: Glibenclamide is a non-insulin-dependent diabetes mellitus ant hyperglycemic

oral agent (NIDDM). He is a member of the insulin secretagogue sulfonylurea class that

stimulates the release of insulin by β cells in the pancreas. The release of basal insulin and

meal-stimulated insulin by sulfonylurea increases.

Pharmacodynamics: Blood glucose decreases acutely by stimulation of the release of

Pancreas' insulin, an impact dependent on the active beta cells of pancreas. Glibenclamide is

an antidiabetic sulphonyl urea agent of the second century. The blood glucose reduction

effect continues in the chronic administration of type II diabetic patients despite a steady

decrease in the secretive insulin response to the medication. The mechanism of action of oral

hypoglycemic sulfonyl urea can be impaired by extra pancreas impact. Glibenclamide and

metformin combinations may have synergistic effects as both agents increase the tolerance of

glucose by different but complementary mechanisms. Glibenclamide causes mild diuresis by

enhanced renal free water clearance, besides its blood glucose reductions. Glibenclamide is

twice as powerful as the glipizide glipizide of second generation.


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SAXAGLIPTIN

Figure 02: Chemical structure of Saxagliptin.

IUPAC Name: 2-[(3R,6S)-2-hydroxy-3-[2-(thiophen-2-yl) acetamido]-1,2-oxaborinan-6-yl]

acetic acid.

Molecular Formula: C12H16BNO5S.

Molecular Weight: 297.352

Category: DPP-4 inhibition.

The new class of drugs that inhibit dipeptidyl peptidase-4 (DPP-4) orally active

hypoglycemic (anti-diabetic drug) Saxagliptin, an anti-diabetes inhibitor for the treatment of

type 2 diabetes is dipeptidyl peptidase-4 (DPP-4). The inhibitors of DPP-4 are a class of

compounds that influence the effect of so called incretins on natural hormones in the body.

Incretins lower blood sugar by increasing the body's intake of sugar, primarily by increasing

pancreas insulin output and reducing liver sugar production. DPP-4 is a membrane-associated

peptidase present in many tissues, lymphocytes, and plasma. [Bristol-Myers Squibb Press

Release]. The DPP-4 has two main pathways, an enzyme activity, and one where DPP-4

binds the adenosine deaminase, which, through dimerisation, conveys intracellular signals.

Saxagliptin forms the reversible, histidine-aided covalent bond of DPP-4 hydroxy oxygen

between its nitrile group and S630. DPP-4 inhaled raises active blood glucagon levels like

peptide 1 (GLP 1), which inhibits pancreatic alpha cell development of glucagon and

improves pancreatic beta cell insulin production.

AIM AND OBJECTIVE

AIM: The purpose of the present investigation is aimed at: To prepare and evaluate

Saxagliptin and Glibenclamide ethosomes containing different concentration of ethanol and

phospholipids by sonication for size reduction of vesicles.

The designated ethosomes of Saxagliptin and Glibenclamide are characterized by

Size and shape.

Entrapment effectiveness.


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Study of release.

The influence of sonication was also investigated on the characteristics of the ethosomes of

Saxagliptin and Glibenclamide.

PLAN OF WORK

To formulate ethosomal gel.

To characterize the prepared formulation using cold method.

To carry out different criteria of assessment, including vesicular form and surface

morphology, vesicular duration, drug quality, efficiency of trapping.

To carry out in vitro drug diffusion study of ethosomal gel.

Preformulation studies.

API Characterization.

Studies into solubility.

Compatibility reports of Drug Excipients.

Construction of Calibration curve.

Formulation Development.

Characterization of ethosomal gel.

o Size and shape analysis.

o Entrapment efficiency.

o Ph.

o Spreadability.

o Drug content and content uniformity.

o Drug release study.

o Invitro release kinetics

Stability studies (at 30ºC / 75 % RH and 40ºC / 75 % RH).

NEED FOR THE STUDY

The increased need to supply patients with less adverse reactions and better compliance with

medication effectively has intensified the pace at which new medicines have been developed.

The transdermal route is expanded to include, separately from oral, innovative drug delivery

technology. The ability to improve transdermal permeation can be useful when complications

are associated with oral administration of medicine.


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Therefore the route of administration needs to be changed for enhanced drug absorption. The

transdermal administration course may be more fitting.

Glibenclamide transdermal penetration cannot be increased by niosomes or liposomes due to

their dimensions and their rigid lipid layer characteristics. In 2004, the lipid vesicles were

used to increase the amount of substrate, thus increasing the penetration of the compound

from the drug. As a result of the innovation, the lipid vesicles were formed. Glibenclamide

was influenced to enhance skin penetration. The compound was stabilized in the vesicles thus

stabilizing the compound for an extended duration. In addition the overstated effect of the

vesicles on cGMP led to the reduction of dose. As a result, the drugs reduced the adverse

effects of the drugs. Instead of the instance that the vesicles released the chemical suddenly,

the compound was formulated slowly for prolonged action. In addition, the vesicles reduced

the fluctuation in the blood levels of the drugs. Since the size of the vesicles was suitable, the

overall results of treatment on the whole varied. The results caused less impact on the

breakouts. Therefore, the patients were able to adapt the changes in the administration.

MATERIALS AND METHODS

Table 03: Chemicals and Materials.

CHEMICALS MANUFACTURED BY

Glibenclamide Chandra labs, Hyderabad, India.

Saxagliptin Chandra labs, Hyderabad, India.

Soya lecithin Mylan Chemicals.

Propylene glycol Avantor chemicals.

Methanol Mark chemical reagents.

Cholesterol Virat lab (Mumbai).

Carbopol-940 Srini Chemicals

Triethanol amine Avantor chemicals

Ultrapure water Mark chemical reagents

Pre-Formulation Studies[13]

“Preformulation testing was an investigation of physical and chemical properties of a drug

substance alone and when combined with excipients. It was the first step in the rational

development of dosage forms.

Analytical Methods

Standard Curvescanning Of Glibenclamide

10 mg of pures medicines have been dissolved and diluted to give 10 μg/ml and scanned for

λmax determination between 200 nm and 400 nm. As a matter of fact, the wavelength of 229


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nm has been chosen. The same thing was done to further analyse the drug solution and

calculate the absorption of the final standard 229 nm solution.

Preparation of standard calibration curve of Glibenclamide

Principle: The Glibenclamide exhibits peak absorbance at 229 nm in methanol.

Instrument used: Shimadzu-1800 UV Spectrophotometer.

Procedure

Preparation of standard solution

Glibenclamide's normal stock solution was made of methanol. The volume was made up of

6.8pHPphosphate buffer to get a concentration of 1000μg/mL. 100 mg of Glibenclamide was

accurily weighted to 100 ml and dissolved in low methanol amounts (SS-I). A concentration

of 100 μg/ml from this 10ml solution was extracted and diluted to 100ml (SS-II).

Preparation of working standard solutions

Additionally, from (SS-II) aliquotide of 0.5ml, 1ml, 1.5ml, 2ml and 2.5ml pipetted into10ml

volumetric flasks. To final phosphates of 5, 10, 15, 20, and 25μg/mlpv, Volumetric flasks

with H6.8 phosphate buffer. At 229 nm, the absorption of each stage was measured.

Standard Curve Scanning of Saxagliptin: The pure drug 10 mg was dissolved in methanol

and diluted to 10 μg/ml concentration and scanned between 200 nm and 400 nm for λmax

determination. A 229 nm long wave has been chosen to equal μmax. The same thing was

done to further analyse the drug solution and calculate the absorption of the final standard

229 nm solution.

Preparation of standard calibration curve of Saxagliptin

Principle: The Saxagliptin exhibits peak absorbance at 278 nm in methanol.

Instrument used: Shimadzu-1800 UV Spectrophotometer.

Procedure

Preparation of standard solution

In methanol, the normal stock solution of Saxagliptin was prepared. 100 mg of Saxagliptin

was correctly measured into a volumetric flask of 100 ml and dissolved in a small amount of

methanol. The volume was composed of a 6.8pHPhosphate buffer to achieve a 1000μg/ml

concentration (SS-I). It was extracted from this 10 ml solution and diluted to 100 ml to

achieve a concentration of 100μg/ml (SS-II).


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Preparation of working standard solutions

Furthermore, from (SS-II) aliquots of 0.5ml, 1ml, 1.5ml, 2ml and 2.5mlwere pipette into10ml

volumetric flasks. The volume was made up with pH6.8 Phosphate buffer to get the 5, 10, 15,

20, and 25μg/ml final concentrations. At 278 nm, the absorbance of each concentration was

measured.

Preparation of Saxagliptin and Glibenclamide Ethosomes (By Cold Method)

Preparation of Glibenclamide ethosomes was followed by method suggested by Touitou et al,

with little modification.[8]

The “ethosomal system of Saxagliptin and Glibenclamide comprised of 2-6 % phospholipids,

20-40 % isopropyl alcohol, 10 % of propylene glycol, 0.005g of cholesterol and aqueous

phase to 100 % w/w. Saxagliptin 0.05g, Glibenclamide 0.1 g was dissolved in IPA in a

covered vessel at room temperature by vigorous stirring. Propylene glycol was added during

stirring. This mixture was heated to 300

in a separate vessel and was added to the mixture

drop wise in the center of the vessel, which was stirred for 5min at 700rpm in a covered

vessel the vesicle size of ethosomal formulation can be decreased to desire extend using

sonication or extrusion method. Finally, the formulation is stored under refrigeration.

Ethosomes were formed spontaneously by the process”.[14]

Figure 03: Cold method for the preparation of ethosomes.


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Table 04: Composition of different ethosomal formulations.

Ethosomal

formulation

Lecithin

(Soya

lecithin%)

IPA(%)

Propylene

glycol

(%)

Saxagliptin

(g)

Glibenclamide

(g)

Cholesterol

(g) Water

GF1 2ml 20ml 5ml 0.05g 0.1g 0.005g 100ml

GF2 4ml 20ml 5ml 0.05g 0.1g 0.005g 100ml

GF3 6ml 20ml 5ml 0.05g 0.1g 0.005g 100ml

GF4 2ml 30ml 5ml 0.05g 0.1g 0.005g 100ml

GF5 4ml 30ml 5ml 0.05g 0.1g 0.005g 100ml

GF6 6ml 30ml 5ml 0.05g 0.1g 0.005g 100ml

GF7 2ml 40ml 5ml 0.05g 0.1g 0.005g 100ml

GF8 4ml 40ml 5ml 0.05g 0.1g 0.005g 100ml

GF9 6ml 40ml 5ml 0.05g 0.1g 0.005g 100ml

Preparation of Saxagliptin and Glibenclamide ethosomal gel

The best achieved suspension of ethosomal vesicles was introduced into carbopol gel (1%,

1.5%, 2% w/w). The specified volume of carbopol 934 powder was slowly applied to

ultrapure water and held for 20 minutes at 1000c. It was applied dropwise to triethanolamine.

Sufficient quantity of formula (GF-2) comprising Saxagliptin and glibenclamide 1.5 percent

w/w was then introduced into gel-base. Water q.s was added with other continuous stirring

formulations before homogeneous formulation was obtained (G-1, G-2 and G-3). Free

Saxagliptin and glibenclamide containing gel was prepared using 1.5 by similar process.

Table 05: Composition of different ethosomal gel formulation.

Saxagliptin &

Glibenclamide ethosomal

suspension(ml)

Carbopol

940(%)

Triethanolami

ne (ml)

Phosphat

e buffer

(pH 6.8)

G-1 20ml 0.5 0.5 q.s

G-2 20ml 1 0.5 q.s

G-3 20ml 1.5 0.5 q.s

Characterization of Ethosomes

Size and Shape Analysis

To determine the average size of the ethosomes, microscopic analysis was conducted. “In

order to observe individual vesicles, a sample of ethosomes was sufficiently diluted with

distilled water and a drop of diluted suspension was mounted on a glass slide covered with a

cover slip and examined under a microscope (magnification 15 Χ 45 X). The diameters of

150 vesicles were randomly calculated using a calibrated stage micrometer eyepiece

micrometer. The method used to measure the average diameter”.[15]

Average diameter = nd / n


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n = number of vesicles.

d = diameter of vesicles.

The vesicle size was diminished by sonication. Although the vesicular dimension of these

vesicles could not be measured with a magnification of 15 Χ 45 X using a microscopic

process. Sonicated vesicles were then examined under a special microscope linked to

software and photomicrographs were taken under magnifications of 400 and 800. For size

analysis, more selected photomicrographs were analyzed using special "particle size analysis"

software developed by BIOVIS. This particular programme functions on photomicrograph

images of normal dimensional measurements.

Scanning Electron Microscopy

Determination of surface morphology (roundness, smoothness and formation of aggregates)

of ethosomal gel with polymer was carried out by scanning electron microscopy (SEM).

In-Vitro Release Studies

Drug Release Study From Dialysis Membrane Of Glibenclamide

Glibenclamide “skin permeation from ethosomal formulation was tested using open-ended

diffusion cells precisely developed according to literacy in our laboratory”.[16]

The successful

permeation area was 2.4 cm and 200 ml for the diffusion cell and receptor cell number,

respectively. At 37 ± 0.5 °C.2 the temperature was preserved.

“The receptor compartment held 200 ml of pH 6.8 buffer and was continuously stirring at 100

rpm by means of a magnetic stirrer. Between the donor and the receptor compartments, ready

dialysis was placed. Ethosomal formulation was added to the dialysis membrane and the

material of the diffusion cell was placed under continuous stirring, during which 5 ml of

samples were extracted at fixed time intervals from the receptor compartment of the diffusion

cell and analyzed by spectrometric approach at 229 nm following sufficient dilution. The

receptor process was replenished directly with the same fresh pH6.8 buffer volume. For

opioid release trials, triplicate experiments were performed”.[2]

Drug Release Study From Dialysis Membrane of Saxagliptin

Saxagliptin's skin permeation from ethosomal formulation was tested using open-ended

diffusion cells primarily developed for literacy in our laboratory. The successful permeation


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area was 2.4 cm and 200 ml for the diffusion cell and receptor cell number, respectively. At

37 ± 0.5 °C.2 the temperature was preserved.

The “receptor compartment held 200 ml of pH 6.8 buffer and was continuously stirring at 100

rpm by means of a magnetic stirrer. Between the donor and the receptor compartments, ready

dialysis was placed. The ethosomal formulation was added to the dialysis membrane and the

material of the diffusion cell was placed under continuous stirring, during which 5 ml of

samples were extracted at fixed time intervals from the receptor compartment of the diffusion

cell and examined by spectrometric approach at 278 nm following sufficient dilution. The

receptor process was instantly replenished with a fresh pH 6.8 buffer of equivalent length.

For opioid release trials, triplicate experiments were performed”.[17]

In-vitro release kinetics: (Harris shoaib et al., 2006)[18]

A series of “kinetic models were used to describe the in vitro release effects and analyse the

way that they worked out. Eq. (2) determines the mechanisms by which the release rate of the

pharmaceuticals depends on its concentration. Eq. (3) determines the exhaust from the unit

where the exhaust rate depends on the concentration. Higuchi (1963) explains the release of

time dependent drugs and the distribution of insoluble matrix from a square root based on

Ficki”.

The “results of the in vitro release profile obtained for all the formulations are defined in the

following three data treatment modes: all release regulation, low and high release scenarios:

A zero-order kinetic model of cumulative drug release percentage versus time.

We plotted the course of first-order-Log cumulative percent for our therapy versus time.

Cumulative percent drug release d versus square root of time, Hikuichi's model.

The Korsmeyer Equation/Peppa Model- Log cumulative drug release percentage versus log

time”.[19]

Table 06: Developing a new diffusion model for cylindrical form.

S.No Diffusion Exponent (n) Overall solute

Diffusion mechanism

1. 0.45 Fickian diffusion

2. 0.45<n<0.89 Anomalous (non-Fickian) diffusion

3. 0.89 Case-II transport

4. n>0.89 Super case-II transport” 20


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Stability Studies: The stability analysis for Glibenclamide ethosomal preparation was

performed at two separate temperatures, i.e. the cooling temperature (4 ± 2 °C) at room

temperature (27 ± 2 °C) for 8 weeks (as per ICH guidelines). The formulation was subjected

to a stability analysis and placed in a borosilicate bottle to prevent any contact between the

ethosomal preparation and the container glass that could impact the findings.

In-vitro stability release study

Drug stability and vesicle stability are the key determinants of formulation stability,

experiments have been carried out to determine the overall drug content at room temperature

(27 ± 2 ° C) and cooling temperature (4 ± 2 ° C). Samples were collected every 2 weeks and

absorption was seen at 229 nm in the U.V spectrometer.

RESULTS AND DISCUSSION

Preformulation Studies

Description

These tests were performed as per the procedure and the results were illustrated in the

following table.

Table 07: Table showing the description of Saxagliptin (API).

Test Description

Colour White crystalline powder

Table 08: Table showing the description of Glibenclamide (API).

Test Description

Colour A white crystalline odorless powder

Result: The results were decided according to requirements.

Solubility

These studies were carried out carried out in conjunction with the protocol and the findings

are shown in the following table.

Table 09: Table showing Saxagliptin Solubility (API) in different solvents.

Solvents Solubility

Water Sparingly soluble

pH6.8 Phosphate buffer Soluble

Methanol Freely Soluble

Chloroform Slightly Soluble

Acetonitrile Freely Soluble” 12


2080


Table 10: Solubility of Glibenclamide (API) in various solvents.

Solvents Solubility

Water Insoluble

Methanol Soluble

Alcohol Soluble

Acetonitrile soluble

Melting Point

This test is performed as per procedure and the result was illustrated in the following table.

Table 11: Table showing the melting point of API’s.

Material Melting Point

Saxagliptin 103-1070c

Table 12: Showing the point of melting API’s.

Material Melting Point

Glibenclamide 1690c

Result: The result has been found to be minimal.

Scientific Research

Saxagliptin Linearization Order In Ph 6.8 Phosphate Buffer

Table 13: Calibration Curve Data of Saxagliptin.

Concentration (µg /ml) Absorbance

0 0

5 0.145

10 0.326

15 0.507

20 0.667

25 0.816

Figure 04: Calibration curve plot of Saxagliptin.


2081


Figure 05: Spectrum of Saxagliptin at 278nm by UV.

Glibenclamide calibration curve in ph 6.8 phosphate buffer

Table 14: Concentration and absorbance.

S.NO Concentration Absorbance

1 0 0

2 5 0.15

3 10 0.291

4 15 0.462

5 20 0.623

6 25 0.765

Figure 06: Calibration curve of Glibenclamide graph.


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Figure 07: UV spectrum for Glibenclamide at 229nm.

FTIR STUDIES

In functional peaks and in functional classes, IR spectra were compared and checked for

improvement. It is evident from the continuum that there is no correlation between the selected

carriers, drugs and mixtures. Consequently, without any mutual interaction, the chosen carrier

was found to be compatible with those selected carriers.

Figure 08: FTIR Spectra of SAXAGLIPTIN pure drug.


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Figure 09: FTIR Spectra of optimized formulation.

Table 15: FTIR Interpretation.

Charecteristic

Peak

Literature

Values

Observed Values

Pure Drug Optimized

Formulation

N-H 3000-2800 2983.16 2921.60

C ═ O 1685-1666 1675.50 1675.53

FTIR experiments were performed to illustrate the safety of the drug with several excipients.

The following figures indicate that functional groups such as N-H with the 3000-2800

observation range have peaks in pure drugs at 2983.16 and streamlined formulation at

2921.60. Likewise, the functional group C ⁇ O has a peak range of 1685-1666 with pure drug

peaks at 1675.50 and integrated formulation at 1675.53. The functional groups are present

both in the pure drug and in the refined formulation. The pure substance can then be inferred

to be consistent with the excipients used in the analysis.

Prepartion of Combination of Saxagliptin and Glibenclamid Eethosomes.

Ethosomal formulations consisting of phospholipid, two different medicines and ethanol have

been prepared using the procedure detailed in the last chapter materials and methods and also

with a slight literature modification. The ethosomal suspensions obtained by sonication were

mildly yellowish in color and hazy in nature. Further examination of different features and

effects of sonication has been performed, and characterization results are reported.


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Ethosomes Characterization

Although the physical characterization of the dose form is directed at physical honesty, the

data have been clustered together at one location. Discussion of the results listed under the

same heading for the formulation of ethosomes.

Size And Shape Analysis

Microscopical inspection was conducted under distinct magnification to imagine the vesicular

shape, lamellarity and the size of ethosomal preparations.

Scanning Electron Microscope (Sem).

Figure 10: Scanning electron microscope image.

Ethosomal Gel Entrapment Efficiency

Ultracentrifugation explored the potential of vesicles for drug trapping until the existence of

bilayer vesicles was established in the ethosomal method. The method used to extract drug-

containing vesicles and the untapped or free drug used for the evaluation of trapping

performance was ultra-centrifugation.

Table 16: Drug entrapment efficiency of Saxagliptin.

Formulation code Entrapment

efficiency(%)

GF1 71.8

GF2 77.6

GF3 78.8

GF4 77.9

GF5 81.27

GF6 85.42


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GF7 82.46

GF8 82.17

GF9 86.67

Table 17: Drug entrapment efficiency of Glibenclamide.

Formulation code Entrapment

efficiency (%)

GF1 72.1

GF2 76.5

GF3 78.2

GF4 77.1

GF5 81.4

GF6 84.8

GF7 81.9

GF8 82.7

GF9 86.8

As calculated by ultracentrifugation, the overall capture efficiency of ethosomal vesicles was

86.67 percent for Saxagliptin and 86.8 percent for Glibenclamide ethosomal formulations

containing 40 percent IPA (GF9). As the IPA concentration increased from 20 percent to 40

percent w/w, the capture efficiency increased and the ethanol concentration increased more

(> 40 percent w/w). Four percent phospholipid trapping results also indicate optimum

trapping efficiency concentrations and, subsequently, higher or decreased phospholipid

concentrations lower trapping efficiency. The findings further validate these findings of Jain

NK et al.

The efficacy of ethosomal formulations in capture is substantially different and is stated in

the table.

The potential decrease in vesicle size may be due to an improvement in entrapment

performance. The negative effect on the vesicle, which is greater in size during ultra-

centrifugation. The more uniform lamellae, smaller vesicles and uniform scale are given by

Sonication which can also contribute to improved stabilization during ultracentrifuge and

lower vesicular disruption.


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Evaluation of Ethosomal Gel

Table 18: Organoleptic characteristics of ethosomal gel.

Organoleptic Characteristics:

Color: brownish-yellow.

Greasiness: Non greasy.

Grittiness: Free from grittiness.

Ease of application: Easily/smoothly applied.

Skin irritation: No skin irritation.

Washability: Easily washable without leaving any residue on

the surface of the skin.

Table 19: Physical appearance.

Formulation

code Color Homogeneity Consistency

Phase

separation

GF1 Creamy white Homogenous Smooth -









Table 20: Spreadability studies.

S.no Formulation code Spreadability (g.cm/sec)

1 GF1 14.28

2 GF2 16.42

3 GF3 13.74

4 GF4 15.26

5 GF5 16.67

6 GF6 17.81

7 GF7 18.54

8 GF8 15.49

9 GF9 18.76

Figure 11: Spreadability graph for GF1-GF9.


2087


Table 20: Rheological studies (for 10rpm spindle 6)

S.no Formulation code Viscosity (cps)

1 GF1 1815

2 GF2 1824

3 GF3 2046

4 GF4 2132

5 GF5 1674

6 GF6 1679

7 GF7 1670

8 GF8 1674

9 GF9 1672

Figure 12: Viscosity Graph for GF1-GF9.

Figure 13: PH

measurements.

The pH of gels was measured by using electrode based digital pH meter.


2088


Table 20: PH

measurements of Ethosomal gel.

The pH values for all formulations were in the range of 5.9 to 6.6

Formulation code pH

GF1 5.9

GF2 6.3

GF3 6.8

GF4 5.9

GF5 6.4

GF6 6.6

GF7 6.5

GF8 6.6

GF9 6.8

Figure 14: Surface PH graph for GF1-GF9.

Drug content and content uniformity

Table 21: Drug content for Saxagliptin.

Formulation code Drug content(%)

GF1 95.4

GF2 97.8

GF3 98.1

GF4 96.9

GF5 97.7

GF6 98.6

GF7 94.7

GF8 96.4

GF9 98.2


2089


Figure 15: Drug Content Graph For GF1-GF9.

Table 22: Drug content for Glibenclamide.

Formulation code Drug content (%)

GF1 95.2

GF2 97.4

GF3 98.6

GF4 96.5

GF5 97.1

GF6 98.4

GF7 94.8

GF8 96.9

GF9 98.4

Figure 16: Drug Content Graph For GF1-GF9.

Vesicle size

Vesicle size of the ethosomes was measured by zeta sizer. The size of the vesicle was found

to be 430nm with a pdi of 0.168.


2090


Results size(d.nm): %Intensity Width(d.nm)

Z- AVERAGE (D.NM): 430 Peak1: 426.5 100 104.2

PdI: 0.168 Peak2: -- -- --

Intercept: 0.96 Peak3: -- -- --

Result quality: good

Figure 17: Vesicle size.

Table 23: Ethosomes In-vitro cumulative % drug release profile for Saxagliptin.

Time

(hrs) GF1 GF2 GF3 GF4 GF5 GF6 GF7 GF8 GF9

0.08 32.6 12.6 6.57 2.15 6.28 7.26 6.6 5.31 4.46

0.16 48.3 29.08 16.29 8.26 13.5 16.84 11.08 12.42 9.4

0.25 56.41 32.2 29.04 20.68 26.15 31.29 27.2 24.4 20.42

0.5 68.27 51.04 36.02 26.48 37.62 44.61 34.04 31.24 28.45

1 74.42 67.5 62.1 37.02 43.67 55.29 39.7 37.02 34.72

2 86.3 82.11 74.3 44.6 55.72 60.46 52.4 49.7 46.27

4 100.2 90.6 89.3 55.7 64.28 68.4 60.6 55.5 58.37

6 100.8 100.9 60.77 69.5 74.6 67.7 64.4 60.22

12 69.1 77.2 83.6 79.5 73.3 68.12

24 77.62 86.12 96.7 88.11 78.2 72.7


2091


Figure 18: Ethosomes containing Saxagliptin dissolution profile for formulations GF1-

GF3.

Figure 19: Ethosomes containing Saxagliptindissolution profile for formulations GF4-

GF6.


2092


Figure 20: Ethosomes containing Saxagliptindissolution profile for formulations GF7-

GF9.

Table 24: Cumulative proportion of cumulative Glibenclamide in vitro ethosomes.

Time

(hrs) GF1 GF2 GF3 GF4 GF5 GF6 GF7 GF8 GF9

0.08 32.4 12.8 6.57 2.16 6.24 7.24 6.6 5.12 4.42

0.16 47.2 29.14 16.29 8.24 13.59 15.85 11.08 12.46 9.18

0.25 57.40 32.21 29.04 20.67 26.14 30.22 27.2 24.25 20.35

0.5 69.24 51.24 36.02 26.42 37.61 42.64 34.04 31.24 28.62

1 72.46 67.16 62.1 37.04 43.66 54.26 39.7 35.14 34.59

2 84.5 82.17 74.3 44.8 55.72 61.47 52.4 48.75 46.24

4 100.4 90.52 89.3 55.6 64.27 67.45 60.6 56.52 57.64

6 100.6 100.9 60.74 69.54 74.40 67.7 65.14 61.22

12 69.12 77.25 83.12 79.5 72.26 67.12

24 77.69 86.14 96.48 88.11 78.4 73.7

Figure 21: Ethosomes containing Glibenclamide dissolution profile for formulations

GF1-GF3.


2093



GF4-GF6.


GF7-GF9.

Discussion For In-Vitro Release of Combination of Saxagliptin and Glibenclamide

Ethosomal Gel .

From the above two tables, it was verified the ethosomal gel release theory of GF4, GF5,

GF6, GF7, GF8, GF9 up to 24 hours in two products. And it was also confirmed from the

tables that the formulation (GF6) showed maximum drug release of up to 24 hours in both

products.


2094


Pharmacokinetic Profiles Forgf6 Ethosomal Gel.

Table 25: Release kinetics for optimized formulation.

ZERO FIRST HIGUCHI PEPPAS

% CDR Vs

T

Log %

Remain Vs T

%CDR Vs

√T

Log C Vs

Log T

Slope 2.957925303 -0.05439445 17.25120161 0.38390172

Intercept 37.11954601 1.806980214 24.37709307 1.58555375

R 2 0.611771069 0.949144503 0.842450504 0.82013441

Figure 24: Zero Order Kinetics ForGF6 Ethosomal gel.

Figure 25: First Order Kinetics For GF6 Ethosomal gel.


2095


Figure 26: Higuchis model For GF6 Ethosomal gel.

Figure 27: Peppas model For GF6 Ethosomal gel.

Stability Studies

The stability experiments were performed in conjunction with the protocol mentioned in the

section of the chapter. The findings are seen in the table below.


2096


Table 26: % Entrapment efficiency and % Drug content after stability studies.

Number of

Days

% Entrapment Efficiency

at temperatures

% Drug Content

at temperatures

4±2oC 25±2

oC 37±2

oC 4±2

oC 25±2

oC 37±2

oC

15 84.8 84.8 84.8 98.6 98.47 98.60

30 84.79 84.48 83.49 98.12 98.92 98.71

45 84.26 83.28 83.27 97.11 98.42 98.50

90 83.47 83.22 82.78 97.42 98.34 98.02

Table 27: Cumulative in-vitro percent drug release profile for Glibenclamide

Ethosomes Optimized GF6 formulation at various temperatures.

S.No Time

in hrs 4±2

oC 25±2

oC 37±2

oC

1 0 0 0 0

2 0.08 5.942 6.4476 6.134

3 0.16 15.1530 16.428 15.630

4 0.25 28.7 30.91 29.42

5 0.5 39.59 42.930 40.845

6 1 50.634 54.897 52.236

7 2 54.815 59.431 56.546

8 4 61.18 66.332 63.112

9 6 67.07 72.718 69.188

10 12 78.47 85.078 80.948

11 24 89.015 96.511 91.826

Figure 28: Graph showing dissolution profile for formulations GF6 after storage at

different temperatures.


2097


SUMMARY

There are some possible benefits of the Transdermal path over traditional routes. These

benefits include the prevention of first-pass digestion, the predictable and sustained duration

of action, the minimization of adverse side effects, the effectiveness of short half-life

medications, the enhancement of physiological and pharmacological reactions, and, most

notably, the avoidance of changes in blood levels. It offers comfort for patients. Yet poor

penetration rates are one of the big issues with successful drug delivery.

The vesicular mechanism (liposomes and niosomes) is emerging as the next step in

enhancing the topical delivery of drugs. Liposome technology has improved, so much so that

they can now be called "Ehosomes", along with the "system" which has greater transdermal

flux as it produces little skin inflammation and produces excellent skin depositing capability.

With little change, the mixture of Saxagliptin and Glibenclamide ethosomes was prepared

using the method stated by Touitou et al., (2000). Ethosomes containing 20 percent, 30

percent, & 40 percent IPA with sonication were tested.

After verifying the presence of vesicles and their duration, ultra-centrifugation was tested for

drugs trapped by the vesicular system. Sonicated ethosomes showed higher value i.e. 96.7

percent containing 40 percent w/w IPA. The dialysis membrane was used to conduct in-vitro

liberation. The first order was found to be the order of drug release for optimized gel

formulation. The ethosomes containing 30 percent w/w IPAA have considered the percentage

of opioid concentration in the skin to be maximum.

CONCLUSION

In the present work, the mixture of ethosomal gels of Saxagliptin and glibenclamide has been

formulated.

The findings of this research suggest that the process of ion gelation can be successfully

used to generate ethosomal gels of Saxagliptin and glibenclamide.

The physical mixture's FT-IR spectrum showed that the drug is consistent with the

polymers and copolymers used.

Carbopol and IPA comprising ethosomal gels and phospholipids had a minimum size

length of 613μm.

Increased polymer concentration contributed to an improvement in the efficacy of drug

entrapment, particle size, percentage.


2098


As the polymer and copolymer concentration increased, the in vitro drug release

decreased.

In comparison with other formulations, GF6 displays maximal drug release in 1440 min,

across all formulations.

Drug release mechanism review found that the drug release from the formulations

followed the mechanism of non-fickian diffusion and followed first-order kinetics.

Based on the results of evaluation tests, GF6 coded formulation was considered to be the

best formulation.

REFERENCES

1. Jain N, Talegonkar S, Jain NK. Modern methods of accessing the blood stream: Evolving

transdermal drug delivery techniques. The Study in Pharma, Sep - October 2004; 41-60.

2. Jain NK. Advances in the distribution of controlled and innovative medications. 1ed,

Publishing of New Delhi:stCBS, 2001; 428-451.

3. Jain S, Bhandra D, Jain S, and Jain NK. Transfersomes, a novel transdermal drug delivery

carrier. 1st Version Controlled and Novel Drug Distribution. CBS New Delhi Publishers

and Distributors, 1997; 426-451.

4. Vyas SP, Khar RK. Managed principles and developments in drug distribution. Fresh

Delhi's Vallabhprakashan. The, 2002 First Edition: 173 - 243.

5. Tuitou E, Godin B, and Weiss C. Improved transmission by Ethosomal carriers into and

through the skin. Study on Substance Dev, 2000; 50; 406 - 445.

6. Meera C Singh, Ajinkya S Naik, Sawant SD. Method of delivery of transdermal

medications with a big focus on transdermal patches, 2010 J Pharm Res; 3(10):

2537-2543.

7. Lachman L, Lieberman HA, Kanig JL. Industrial Pharmacy Theory and Practice.

Philadelphia, PA: Lea and Febiger, respectively. From, 1987; 317-318.

8. Leon Shargel, Susanna Wu-Pong, Andrew BC Yu.Drug products with modified-release.

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Two-nd ed. New Delhi: Editors & Distributors of CBS, 2005; 29: 523-552.

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formulation and assessment. Clinical Res of Asian J Pharm, 2011; 4(2): 93-98.

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12. Kaplun-Frischoff Y and Touitou E. Skin permeation testosterone improvement through

technique by eutectic drug development and association with skin lipids, 1997 J Pharm

Sci; 86: 1394 - 9.

13. Bhavsar JD, Brahambhatt VG, Patel MR, Patel KR, Patel NM. Article of Review: Novel

Semisolid Approaches. World Res, 2011 Int J Pharm, 2(1): 1-22.

14. Khandare JN, Jiwandad BH and Uppal RR. Nimesulideniosomes for topical distribution

planning and assessment. Int Drug, 2001 April; 38(4): 197-202 respectively.

15. Facts on Medications and Reference. Facts and reference publication, edition of Missouri,

® 1999; 25855.

16. Bhavana Vora, Khopade AJ and Jain NK. Transdermal device dependent on niosomes for

distribution of contraceptives. Abstract of the, 1996 IPC: AP59; 41.

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planning and assessment. Int Drug, 2001 April; 38(4): 197-202 respectively.

18. Satturwar PM, Khandare JN and Nanda VS. Ketoconazole Niosomaldelivery. Ind Drugs,

2001 April: 38 (12): 620-624, respectively.

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FORMULATION AND INVITRO EVALUATION OF ETHOSOMAL …