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FORMULATION AND CHARACTERIZATION OF RALOXIFENE BETA-CYCLODEXTRINE INCLUSION COMPLEX FOR SOLUBILITY ENHANCEMENT

 

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ABOUT AUTHORS:
Gazi Juned M.*1, Javia A.R1, Sheth A.K1, Sachinkumar P. Chauhan, Nirmal Shah
1Department of Pharmacy, Sumandeep Vidyapeeth University,
At & Po Pipariya, Ta.- Waghodia, Dist. Vadodara-391760.
(Gujarat) India
gaziriju89@gmail.com

ABSTRACT
Raloxifene is a second genaration selective estrogen receptor modulator used to prevent osteoporosis in postmenopausal women. which is insoluble in water and half life is 27.7 hr. it has estrogen effect on bone and cholesterol metabolism but behaves as a complete estrogen antagonist on mammary gland and uterine tissue. raloxifene needs enhancement of solubility and dissolution rate to improve its oral bipavailability and therapeutic efficasy. among  the various approaches to enhance solubility and dissolution rate of poorly soluble drugs complexation with cyclodextrine is an effective and indutrially accepted technique. In the present  investigation complexation of raloxifene with β-CD was carried out by using various technique like physical blending method, kneading method, solvent evaporation method and co-precipitation method. From the various characterization studies like solubility determination, drug content determination, production yield & in vitro dissolution study, it was observed that there was a significant rise in the aqueous solubility and dissolution rate of the raloxifene from the β-cd incusion complex. from the result of statistical analysis, batch RLX-6 by kneading method was found to be optimised batch, because of the maximum enhancement in solubility and rate of dissolution.  Optimised batch was also studied for compatibility by FTIR studies, and also it was found to be stable for the period of 90 days as per the ICH guidelines.

REFERENCE ID: PHARMATUTOR-ART-2014

INTRODUCTION
Raloxifene is a second generation selective estrogen receptor modulator used to prevent osteoporosis in postmenopausal women. In the present  investigation   complexation of raloxifene with β-CD was carried out by using various technique like physical blending method, kneading method, solvent evaporation method and co-precipitation. It has estrogen agonist effects on bone and cholesterol metabolism but behaves as a complete estrogen antagonist on mammary gland and uterine tissue.[1]


Poorly soluble drugs are associated with slow drug absorption leading to inadequate and variable bioavailability. Most formulation strategies for such drugs are targeted at enhancing their dissolution rate and/or solubility in vivo by achieving their fine dispersion at absorption level.Raloxifene hydrochloride (RLX HCL) is a selective estrogen receptor modulator (SERM) shown to be effective in the prevention of osteoporosis with potential utility as a substitute for long-term female hormone replacement therapy. RLX HCl was chosen in this study, is a  poorly water soluble drug known to demonstrate dissolution and solubility limited absorption. RLX HCl being a low solubility drug with high permeability, is classified a Class II drug in BCS adopted by USFDA.

In the development of formulations, solubility of the active component in the drug product has great importance. Earlier research supports that the  inclusion of drug  β-cd complex  results in improvement in dissolution characteristic of the formulation. The present work is aimed to explore the effect of β-cd drug complex on solubility and dissolution rate of RLX HCL in different simulated gastrointestinal fluids.[2]


In 2006, the National Cancer Institute announced that raloxifene was as effective as tamoxifen in reducing the incidence of breast cancer in postmenopausal women at increased risk. A major adverse effect of tamoxifen is uterine cancer; raloxifene had fewer uterine cancers. Tamoxifen increased the risk of cataracts, but raloxifene did not. Both groups had more blood clots in veins and the lungs, but that side effect was more common with tamoxifen than raloxifene.[2][3][4] On September 14, 2007, the U.S. Food and Drug Administration announced approval of raloxifene for reducing the risk of invasive breast cancer in postmenopausal women with osteoporosis and in postmenopausal women at high risk for invasive breast cancer.[1]

MATERIALS AND METHODOLOGY
Materials :  
Raloxifene was gifted by Arti Drugs Pvt.Limited (Mumbai), Triveni Interchem Pvt. Limited (Vapi), Methanol Svu (Baroda). All other chemicals used were of analytical grade. taken in to consideration.

PREFORMULATION STUDY:
It is the first step in rational development of dosage forms of drug substance.

Following  preformulation  study check on Raloxifene solid dispersion complex.

Organoleptic property:[1,3]
The drug sample was evaluated for its colour, odour and physical appearance.

Melting point determination:[1,3]
Melting point of the drug sample was determined by capillary method.

Determination of λmax:[4]
Λmax of the drug inPhosphate buffer 5.5 was determined by UV-Visible  spectrophotometer.

Standard calibration curve of  Raloxifene in Phosphate Buffer pH 5.5: [5]
Calibration curve of  Raloxifene in Phosphate buffer 5.5  was determined by UV-Visible  spectrophotometer.

Standard  calibration curve of  Raloxifene in methanol:[5]
Calibration curve of raloxifene in methanol was determined by UV-Visible spectrophoto-meter.

Compatibility Study:[4]
Infrared spectra of pure drug raloxifene, β-cd, as well as drug β-cd mixture was taken by using FT-IR Spectrophotometer  for the evaluation of compatubility.

Solubility determination of raloxifene in water:[4]
Excess amount of drug was added in to the 100 ml water in volumetric flask and  then the solution was filtered. After filtration remaining  filtrate was evaporated to precipitate the dissolved drug. Weight of the raloxifene was ditermined after evaporation for solubility determination.

FORMULATION OF INCLUSION COMPLEX OF RALOXIFENE  WITH     β-CYCLODEXTRIN:[5]

Method of preparation:
The inclusive complex of Raloxifene with β-Cyclodextrin was prepared by following methods:

Physical mixture:
Raloxifenewith β-Cyclodextrin in different ratios (i.e. 1:1, 1:2& 1:3) were mixed in a mortar pestle for about one hour with constant trituration, passed through sieve No. 80 and stored in desiccators over fused calcium chloride.

Co-precipitation method:
The inclusion complex precipitated as a crystalline powder which was pulverized and passed through sieve No. 80 and stored in a desiccator.

Kneading method:
In this method, drug and cyclodextrine were grinded with intense trituration for 30 min. the mixture is then kneaded with hydroalcoholic solution to get paste consistency.

Solvent Evaporation Technique:
Also known as solid dispersion method, involves mixing of the alcoholic solution of drug and aqeuos solution of cyds with stirring to get a molecular dispersion.

OPTIMIZATION BY FACTORIAL DESIGN:
All 9 batches of solid dispersion of Raloxifene with β-cyclodextrin were prepared according to 32factorial designs.

EVALUATION OF INCLUSION COMPLEX OF RALOXIFENE WITH β – CD:[4,5,6]

Drug – Polymer Interaction:
It was carried out by FT-IR Spectroscopy
Infrared spectra of all batches were preparedusing FT-IR Spectrophoto-meter for the assessement of compatibility of drug and β-cd inclusion complex.

Percentage yield:
The yield was calculated by dividing the weight of the collected solid dispersion by the weight of all the non-volatile components used for preparation of the solid dispersion and expressed in terms of percentage.

Drug content estimation:
Drug content estimation was done by  the absorbance of the drug solution  measured at 289 nm using appropriate blank. The amount of  drug was calculated using calibration curve equation.

Solubility estimation:
Excess amount of inclusion complex was added in to the 100 ml water in volumetric flask and mixture was filtered and then the filtrate was evaporated to precipitate the dissolved drug. Weight of the incusion complex was ditermined after evaporation for solubility determination.

In vitro dissolution study
In-vitro dissolution of Raloxifene inclusion complex was studied in USP XXIV dissolution apparatus (Electro lab) employing a paddle stirrer. the amount of drug  released at different time interval was calculated and plotted against time and compared with pure drug.

Kinetics of Drug Release:
In order to investigate the mechanism of drug release from inclusion complex of different ratios, the release data obtained from dissolution studies were fitted to various kinetic equations.

Stability study:[7]
The stability study was carried out for optimized formulation as per ICH guidelines (Feb. 2003).

RESULT AND DISCUSSION

Result:

[Table 1: Organoleptic property of Raloxifene]

Sr. No.

Parameter

Raloxifene

1

Colour

Dark yellowish

2

Odour

Odourless

3

Appearance

powder

[Table 2: Melting point of Raloxifene]

Reported Melting Point

Observed Melting Point

144°c-146°c

143°c-147°c

 

[Table 3 :λmax (nm) of Raloxifene]

Drug

λmax (nm)

Raloxifene

289

 

[Table 4 : Calibration curve of Raloxifene in methanol.]

 

Sl.

No.

 

Concentration

(µg/ml)

 

Absorbance

 

Mean(±SD)

 

I

II

III

1

0

0

0

0

0

2

3

0.298

0.301

0.296

0.298±0.003

3

5

0.449

0.446

0.440

0.445±0.002

4

7

0.576

0.579

0.573

0.576±0.003

5

10

0.756

0.761

0.765

0.760±0.005

6

15

1.076

1.061

1.047

1.061±0.015

 

[Table 5: solubility estimation of  raloxifene]

Sl.

No

ratio

Solvent evaporation method

Kneading method

Co-ppt method

1

1:1

87.3mg/100ml

108.87mg/100ml

96.10mg/100ml

2

1:2

149.64mg/100ml

212.47mg/100ml

163.96mg/100ml

3

1:3

213.3mg/100ml

243.90mg/100ml

262.12mg/100ml

Pure drug (rlx)  

12 mg/100ml water

FT-IR Studies:

FT-IR Spectrum of Raloxifene  in range 4000 to 400 cm-1

[Figure 2: FT-IR Spectrum of pure drug Raloxifene  in range 4000 to 400 cm-1.]

FT-IR Spectrum of β-cyclodextrin in range 4000 to 400 cm-1:

[Figure 3  : FT-IR Spectrum of β-cyclodextrin in range 4000 to 400 cm-1.]

FT-IR Spectrum of  drug + cyclodextrin   in range 4000 to 400 cm-1:

[Figure 4: FT-IR Spectrum of drug + cyclodextrin in range 4000 to 400 cm-1.]

Characterization of inclusion complex :

Production yield:

The production Yield of all formulated batched is given in table.

[Table 6 : Production Yield of Raloxifene – β-cyclodextrin solid dispersion.]

Batch

Ratios

Mean Production

Yield (%)

 RLX1

 1:1

 91.83±1.89

 RLX2

 1:2

 91.22±1.01

 RLX3

 1:3

 92.08±0.38

 RLX4

 1:1

 78.33±0.76

 RLX5

 1:2

 83.44±1.38

 RLX6

 1:3

 90.08±1.18

 RLX7

 1:1

 72.33±1.04

 RLX8

 1:2

 76.21±0.50

 RLX9

 1:3

 83.33±0.76

Drug content:

The drug content of all prepared batches is given in table.

[Table 7 : Drug Content Study of Raloxifene – β-cyclodextrin solid dispersion.]

Sl

 No

Batch 

       Drug Content

(%)

1

RLX1

45.3

2

RLX2

51.96

3

RLX3

54.92

4

RLX4

51.6

5

RLX5

56.66

6

RLX6

62.7

7

RLX7

48.54

8

RLX8

55.96

9

RLX9

67.04

In vitro release studies:

The in vitrorelease studies are carried out for a period of 5 to 60 min.

[Table 8 : % Cumulative Release Data of Formulation RLX1, RLX2 and RLX3.]

Time (in min)

Formulations(SEM)

Drug

RLX1

RLX2

RLX3

0

0

0

0

0

5

9.21

18.57±0.20

25.27±0.19

31.80±0.47

10

11.96

21.12±0.13

28.44±0.33

37.12±0.12

15

13.47

24.96±0.07

31.39±0.20

41.27±0.13

30

17.36

28.65±0.27

34.55±0.08

43.38±0.07

45

19.45

31.39±0.13

36.84±0.41

45.36±0.27

60

22.38

35.12±0.26

41.20±0.13

47.90±0.34

[Figure 5 : Dissolution profile of RLX1, RLX2, and RLX3]

[Table 9 : % Cumulative Release Data of FormulationRLX4, RLX5 and RLX6.]

Time (in min)

Formulations(KM)

Drug

RLX4

RLX5

RLX6

0

0

0

0

0

5

9.21

26.37±0.19

30.35±0.13

34.42±0.14

10

11.96

39.44±0.34

41.66±0.27

44.34±0.13

15

13.47

47.02±0.13

51.02±0.13

53.87±0.42

30

17.36

59.02±0.07

62.15±0.49

67.49±0.13

45

19.45

65.88±0.26

68.62±0.19

75.14±0.40

60

22.38

69.75±0.27

78.11±0.42

87.29±0.54

[Figure 6: Dissolution profile of RLX4, RLX5and RLX6.]

[Table 10 : % Cumulative Release Data of Formulation RLX7, RLX8 and RLX9.]

Time (in min)

Formulations(co-ppt method)

Drug

RLX7

RLX8

RLX9

0

0

0

0

0

5

9.21

19.79±0.20

26.94±0.13

32.10±0.18

10

11.96

28.62±0.13

34.98±0.13

40.22±0.19

15

13.47

31.10±0.27

38.84±0.12

46.49±0.19

30

17.36

35.36±0.56

47.55±0.20

52.88±0.13

45

19.45

43.96±0.13