INTRODUCTION:

As we know that our GI tract absorbed molecules by diffusion process. Orally administered drugs move across the membrane in the cells by facilitated diffusion or by selectively permeable method. Two main factors by which the enhancement of drug dissolution is achieves i:e,

1) Drug profile

2) Aqueous nature of GI surrounding

Most of the drugs are water insoluble¹. Poor water solubility of drugs and its active pharmaceutical ingredients shows wide range of problems in industrial technology. Higher percentile of drugs exhibits formulation problems due to very high lipophillicity and most of newly established drugs are insoluble in water^2-3. Absorption studies over part 10 years reveal that broader range of drugs that show low solubility and high permeability belongs to class II BCS category⁴.

All these problems related to drug dissolution can be refined by solubility enhancement techniques of drug development.

Importance of Solubility in Oral Administration:

Solubility is major factor for the effectiveness of drug. Oral ingestion is convenient and frequent route of drug delivery, as most of the drugs are administered through it, because it produces systemic effect by reaching their target site via blood stream. To maintain efficient delivery of various therapeutic agent, bioequivalent oral dosage form are necessary so that to enhance the systemic effect. For this intend, different approaches are being used in drug development process.

About 70% of New Chemical Entity (NCE) discovered during the drug discovery programs have poor bioavailability. Therefore, it is a challenging task for scientist to make the drug more bioavailable for showing targeted therapeutic action.

Enhancing solubility using pharmaceutical awareness will assist both medicinal chemist and druggist for making molecule useful as therapeutic agent rather than throwing it away for its poor solubility.

An improved thoughtfulness of types of polymers, different preparation methods, with molecular contacts of drug and polymer is found in favor of scheming proficient and stable drug delivery system. An enhanced solid-form stability and supersaturation creation with continuation will guide us to deliver a system conferring advantageous plus expected properties and a realistic selection for solubilizing the “difficult to solubilize” drugs^5-6.

Solubility Enhancement Techniques:

Various Techniques are used to enhance the solubility^7-9.

A. Chemical Modifications

1) Salt Formation

2) Co-crystallization

3) Co-solvency

4) Hydrotropic

5) Solubilising agent

6) Nanotechnology.

B. Physical modifications:

1) Particle size reduction (micro ionization, nanosuspension)

2) Modification of crystal habit like polymorphism and pseudopolymorphism

C. Nanotechnology approaches:

1) Nanocrystals

2) Nanocomposites

3) Bionanocomposites

Bionanocomposites:

Bio-Nanocomposites are a subject of current interest to scientists as they offer exciting platforms and serve as an interface between materials science, biology, and nanotechnology. Existing bio-nanocomposites are a result of biopolymers, including polysaccharides, polypeptides and proteins, aliphatic polyesters and polynucleic acids; whereas fillers include clays, hydroxyapatite, and metal nanoparticles¹⁰. Different properties shown by bionanocomposites are thermal stability, solubility in water, biocompatibility and biodegradability. Bionanocomposites are biodegradable polymer or hybrid materials that are produced from plants or microorganisms or either from inorganic solids and exhibit range having less than 100nm scale. Nanocomposites can be distinguished from the composite in terms of petroleum-based polymer as an organic part in nanocomposites, and on the other hand composites are constituted of inorganic additives at nanosize scale. Additionally, one major difference between bio-nanocomposites and bio-composites is that the latter may be constituted of biopolymers, but they do not have the nanosized additives^11-12. These biopolymers show solubility in polar solvents (e.g., water), and petroleum derived polymers are soluble in organic solvents.

Itraconazole^13-14

Itraconazole is a small molecule having molecular formula C₃₅H₃₈C₁₂N₈O₄ and a member of triazole and dioxolane, a N – aryl piperazine (dichlorobenzene, cyclic ketal, conazole anti mycotic drug) shown anti mycotic action.

Important features:

· Interact with 14- α demethylase (cytochrome P-450 enzyme) necessary to convert lanosterol to ergosterol.

· Orally administered.

· Treatment of various skin infections.

· Used to treat yeast infection of mouth, throat, oesophagus

It is potent inhibitor of cytochrome P-450 3A4 isoenzyme, so to inhibit synthesis of ergosterol the important constituent of fungal cell membrane. Itraconazole also interact with phospholipid membrane of the fungus to inhibit its endogenous respiration and impair triglyceride biosynthesis.

MATERIAL AND METHOD:

Materials:

Itraconazole was gifted by Lifecare Neuro Products ltd, baddi, solan. And PVP, chitosan and β –cyclodextrin was obtained from Sri Sai College of pharmacy, Badhani, Punjab.

Technique used in the preparation of Bionanocomposites of itraconazole:

The Bio-nanocomposites of Itraconazole were prepared by Microwave induced technique (MIND). The BNCs were prepared by adding accurately weighed quantity of drug Itraconazole and carrier such Polyvinyl Pyrrolidone, β-Cyclodextrin and Chitosanwere taken in 1:1 to 1:4 w/w proportions as shown in table 1. Homogeneous physical mixture of drug and carrier was prepared using mortar and pestle. Slurry was prepared by adding 5ml ofdistilled water in each gram of drug-carrier physical mixture. A fixed amount of slurry (6g) was placed in a glass beaker and irradiated with microwave radiations at power 700 W with continuous stirring ¹⁵.

Evaluation of bionanocomposites of itraconazole:

A. Preformulation studies:

a. Organoleptic properties: Appearance, color and odor¹⁶.

b. UV spectrum of Itraconazole: Double beam UV-visible spectrophotometer was used to know the λ_max of drug. A 12µg/ml solution of Itraconazole in methanol was scanned in the range of 200-800nm¹⁷.

Estimation of Itraconazole:

· Estimation of Itraconazole by UV-visible spectrophotometer:

The standard stock solution of Itraconazole (1mg/ml) was prepared in methanol. This solution was diluted with methanol, to obtain various dilutions from 3-24µg/ml. Absorbance of these solutions was recorded at 262nm against methanol as blank using UV-visible spectrophotometer and standard curve was plotted against concentration.

c. Solubility Studies:

For quantitative solubility study, surplus amount of drug was taken in washed test tubes containing 1ml of different solvents (Methanol, Ethanol, Acetone, Chloroform, 0.1N HCl, water, phosphate buffer saline of pH 6.8 and 7.4) and test tubes were tightly closed. These test tubes were shake on water bath shaker for 24h at room temperature. After 24h each sample was centrifuged at 15,000rpm and then the supernatant was taken out. After that supernatant was filtered and filtrates was suitably diluted and determined spectrophotometrically^18-19.

d. FTIR of Itraconazole:

FT-IR Spectroscopy was used for structure analysis. The potassium bromide (KBr) disc technique was applied. KBr has transmittance window of 100% (wave number range 4000-4001/cm) and thus does not show any absorption in IR spectrum.

The KBr disc was prepared using 1 mg of Itraconazole in 100mg of spectroscopic grade KBr which has been dried using IR lamp. Both KBr and Itraconazole was mixed and subjected to hydraulic pressure to form disc. This disc was placed in FT-IR chamber. Infrared spectrum was recorded in the 4000 - 400cm^-1 region²⁰.

B. Post formulation evaluation of bionanocomposites of itraconazole:

a. Optical Microscopy:

In the process of preparation of Bio-nanocomposites, each batch were collected and stored at room temperature for further analysis. Pure Itraconazole and Bio-nanocomposite were observed using a Biolux-CTX optical microscope (OM), at magnification of 40X²¹.

b. Transmission Electron Microscopy:

With transmission electron microscopy the particle size and shape of pure drug crystal dispersed in polymer were examined. The morphology of the NCs was obtained by transmission electron microscope.

c. Solubility profile of Bio-nanocomposites of Itraconazole:

The solubility study of BNCs was carried out by adding excess amount of and BNCs (equivalent to 30mg) to 10 ml Phosphate Buffer pH=6.8 in a separate flask. The resultant mixture was stirred for 24 h at 25°C temperature by using orbital shaker incubator. The supernatant liquid was collected and filtered through 0.2 μ membrane filter and analyzed by UV-Visible spectrophotometer at 262nm wavelength.^22-23

d. Percentage Drug Content:

The incorporated itraconazole drug into the BNCs was calculated by dissolving BNCsmixture in the 25ml methanol. The resulting solution was filtered by 0.2μ membrane filter and analyzed by UV-visible spectrophotometer at the wavelength of 262nm against the methanol as a blank.²⁴

e. In-Vitro Drug Release Study:

The in-vitro powder dissolution test was carried out using USP XXIV apparatus II (Paddle) of developed optimized. Drug Release experiments were carried out using magnetic stirrer with controlled temperature. In this method pH 6.8 phosphate buffer was used as dissolution media. The rate of stirring was 100±2rpm. In all formulations the amount of Itraconazole was 200mg. The dosage forms were placed in 900ml of pH 6.8 phosphate buffer and maintained at 36±1˚C. For 60 minutes at suitable intervals (1, 2.5, 5, 10 minutes and so on), 5ml of sample were taken. The dissolution medium was substitutes by 5ml of fresh dissolution fluid to maintain a constant volume. The samples were filtered through a 0.45mm Millipore filter, diluted and analyzed at 262nm by UV/visible spectrophotometer.^25-26

f. XRD of Bionanocomposites: XRD patterns were recorded on a powder x-ray diffractometer using K-beta filter, CuKα-radiation, voltage of 45 kV and a current of 30 mA. The scanning employed was over the 10.00 to 50.00° diffraction angle (2θ) range.²⁶

RESULT AND DISCUSSION:

A. Results and discussion of pre formulation studies of itraconazole:

a. Organoleptic properties:

Table 2: Organoleptic Properties of Itraconazole

Sr. No.	Properties	Inferences
	Colour	White
	Odour	Odourless
	Form	Crystalline
	Taste	Bitter

b. UV Spectroscopy:

i. Determination of absorption maxima in Methanol:

A 12µg/ml solution of Itraconazole in methanol was scanned in the range of 200-400nm. The result of UV spectrum of Itraconazole is shown in Figure 1.

Figure 1: UV Spectrum of Itraconazole in Methanol

Table 3: Absorption maxima (λ _max) of Itraconazole in Methanol

Name of drug	Absorption maxima (λ _max)
Name of drug	Observed	Reference
Itraconazole	262	261

ii. Preparation of standard curve of Itraconazole in Methanol

Table 4: Calibration curve of Itraconazole in Methanol (λ_max= 262nm)

Sr. No.	Concentration (µg/ml)	Absorbance
01.	3	0.154±0.003
02.	6	0.273±0.004
03.	9	0.385±0.002
04.	12	0.505±0.003
05.	15	0.612±0.003
06.	18	0.748±0.003
07.	21	0.864±0.004
08.	24	1.000±0.002

Figure 2: Standard calibration curve of Itraconazole in Methanol

Table 5: Result of regression analysis of UV method

Statistical parameters	Results
λ_max	262nm
Regression equation (y =mx+ c)	y =0.0399x + 0.0284
Slope (m)	0.0399
Intercept (C)	0.0284
Correlation coefficient (R²)	0.9992

DISCUSSION:

The calibration curve for Itraconazole was obtained by using the 3 to 24µg/ml concentration of Itraconazolein methanol. The absorbance was measured at 262nm. The calibration curve of Itraconazole as shows in graph indicated the regression equation Y=0.0399 xs+0.0284 and R²value 0.9992, which shows good linearity as shown in table 5 and figure 2.

c. Solubility studies:

Table 6: Solubility studies of Itraconazole for different solvents

Sr. No	Solvent	Solubility in (mg/ml)
1	Water	0.001±0.001
2	Phosphate Buffer pH 6.8	0.006±0.001
3	Ethanol	0.474±0.005
4	Methanol	1.091±0.005
5	DMF (Dimethyl Formamide)	6.824±0.063
6	DMSO (Dimethyl Sulfoxide)	8.378±0.077
7	Acetone	12.070±0.050
8	Chloroform	14.000±0.025

Figure 3: Solubility study of drug in different solvents

DISCUSSION:

From the above data, it is clearly seen that Itraconazole is highly soluble in Dimethyl Formamide, Dimethyl Sulfoxide, Acetone and Chloroform (figure 3 and table 6).

d. FTIR Studies:

Figure 4: FTIR spectrum of Itraconazole

Table 7: FTIR interpretation of Itraconazole

Reported (cm^-1)	Observed (cm^-1)	Characteristics Peaks
2969	2973.21	Stretchingvibrations of aminogroup
1697	1696.97	C=O and C-O bonds
1425	1454.17	C-N bonds

DISCUSSION:

The FTIR spectra of Itraconazole were shown in the figure 4 and table 7. The principal IR absorption peaks of Itraconazole at 2973.21 cm^-1 (Stretchingvibrations of aminogroup), 1696.97cm^-1 (C=O and C-O bonds), 1454.17 cm^-1 (C-N bonds) were all observed in the spectra of Itraconazole. These observed principal peaks confirmed the purity and authenticity of the Itraconazole.

B. Result and discussion of post formulation evaluation of bionanocomposites of itraconazole:

a. Optical Microscopy:

Figure 5: Optical Microscopy of Bio-nanocomposite containing Itraconazole in Formulation F4.

b. Transmission Electron Microscopy:

Figure 6: TEM image of formulation F4

c. Solubility profile of Bio-nanocomposites of Itraconazole:

Table 8: Solubility Profile of all the Formulations

S.No.	Formulation Code	Solubility (mg/ml)
1	Pure Drug	0.006±0.001
2	F1	0.045±0.005
3	F2	0.094±0.003
4	F3	0.115±0.009
5	F4	0.145±0.005
6	F5	0.011±0.001
7	F6	0.019±0.001
8	F7	0.027±0.001
9	F8	0.024±0.000
10	F9	0.009±0.001
11	F10	0.034±0.001
12	F11	0.042±0.001
13	F12	0.045±0.002

Figure7: Solubility Profile of Bio-nanocomposite containing Itraconazole

DISCUSSION:

Solubility of prepared Bio-nanocomposite was calculated in phosphate buffer pH=6.8as pure drug has less solubility. The results showed in table 8 indicate that Bio-nanocomposite improved the solubility of Itraconazole. It was found that there was a maximum 23 fold increase in solubility when polyvinyl pyrrolidone was used as a carrier in the ratio of 1:4 (drug:carrier).

d. Percentage Drug Content:

Table 9: Percentage drug Content of Itraconazole loaded Bio-nanocomposite

S. No.	Formulation Code	Percentage Drug Content
1	F1	71.50±0.383
2	F2	79.10±0.251
3	F3	86.28±0.383
4	F4	91.80±0.145
5	F5	60.05±0.904
6	F6	64.06±0.663
7	F7	47.77±0.501
8	F8	74.42±0.383
9	F9	55.37±0.383
10	F10	65.31±0.501
11	F11	69.74±0.949
12	F12	74.00±0.631

Figure 8: Percentage drug content of Bio-nanocomposite of Itraconazole

DISCUSSION:

From the table 9, it was found that Percentage drug content of all formulation was found to be in a range 47.77±0.501to91.80±0.145. These results explain that there is a significant effect on percent drug content of bio-nanocomposite with respect to the carrier used as polyvinyl pyrrolidone was found to be the best fit for this study.

e. In-vitro Drug release study:

Table 10: Percentage drug release of Formulation F4 and Pure drug

Time (mins)	% Drug release of F4 Formulation	% Drug release of Pure Drug Suspension
0	0	0
1	41.45±0.830	5.69±0.271
2.5	53.30±0.543	8.93±0.271
5	68.63±1.057	10.44±0.270
10	75.06±0.543	12.11±0.269
15	79.01±0.543	12.34±0.156
20	84.88±0.271	14.15±0.156
25	88.03±0.543	16.67±0.271
30	91.53±0.271	25.04±0.156
40	94.01±0.843	29.02±0.271
50	95.14±0.271	34.46±0.271
60	98.18±0.156	36.45±0.271
90	-	42.29±0.188

Figure 9: In-Vitro Drug release of Bio-nanocomposite of Itraconazole and Pure Drug.

DISCUSSION:

The in-vitro release of drug from the Bio-nanocomposite of Itraconazole was found to be higher as compared to pure drug suspension that showed the effect of carrier in drug release property. Table 10 indicated that in vitro release of Bio-nanocomposite showed 98% released within an hour. The release profiles of Bio-nanocomposite of Itraconazole employed yielded an immediate Itraconazole release. From the in-vitro drug release study it was found that F4 formulation showed higher drug release as compare to pure drug.

f. XRD of Bionanocomposite

Figure 10: XRD study of formulation F4

Table 11: XRD interpretation of formulation F4

Name of component	Reported 2θ	Observed 2θ
Formulation F4	17.5, 20.38, 20.40, 20.36, 20.34 and 20.32	10.90, 14.57, 16.72, 17.58, 18.0825 19.37, 20.48, 22.37, 23.59, 25.44, 27.22

DISCUSSION:

The X-ray diffraction patterns of formulation F4 is showed in table 11. The XRD of pure itraconazole X-ray diffraction patterns exhibit crystalline nature. Figure 10, the XRD pattern of formulation F4 also shows a crystalline nature but is quite different from that of pure Itraconazole. The 2Q values observed in both the cases are different. This constitutes primary evidence that a different solid phase was formed.

CONCLUSION:

There is a need for enhancement of solubility and dissolution of poorly soluble drugs. It emerges out of the analysis of most of the approaches that these are based on generating drug dispersion at nanoscale level. To maintain faster solubility and dissolution kinetic of bio nanocomposite, it is essential to keep integrity of such active pharmaceutical ingredient. From all aspects, I concluded that in vitro data obtained for bionanocomposites of itraconazole showed increase solubility in phosphate buffer having pH= 6.8 and the method of preparation of itraconazole found to be simple, reproducible, provide good solubility and drug content. Also it is obtained that there was maximum 23 fold increase in solubility when Poly vinyl pyrolidine used as a carrier in 1:4. Thus formulation prepared showed immediate release behavior and Poly vinyl pyrolidine considered as effective carrier for crystallization of itraconazole.

ACKNOWLEDGEMENT:

I am thankful to Life care neuro products, Baddi, Himachal Pradesh, for providing me the gift sample of Itraconazole. I owe a lot to Sri Sai College of Pharmacy for providing me all the necessary facilities to carry out this study at their disposal.

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Received on 25.05.2021 Modified on 29.12.2021

Asian J. Pharm. Res. 2022; 12(3):192-198.

DOI: 10.52711/2231-5691.2022.00031

S. No	Formulation Code	Molar Ratio	Drug (mg)	Polyvinyl Pyrrolidone	β-Cyclodextrin	Chitosan
1	F1	1:1	500	500	-	-
2	F2	1:2	333.37	666.63	-	-
3	F3	1:3	250	750	-	-
4	F4	1:4	200	800	-	-
5	F5	1:1	500	-	500	-
6	F6	1:2	333.37	-	666.63	-
7	F7	1:3	250	-	750	-
8	F8	1:4	200	-	800	-
9	F9	1:1	500	-	-	500
10	F10	1:2	333.37	-	-	666.63
11	F11	1:3	250	-	-	750
12	F12	1:4	200	-	-	800