Development and Validation of Stability Indicating Method for Determination of Lurasidone in Bulk...

IJPRD, 2013; Vol 5(09): November-2013 (103 014) International Standard Serial Number 0974 9446

Available online on www.ijprd.com

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DEVELOPMENT AND VALIDATION OF STABILITY INDICATING METHOD FOR DETERMINATION OF

LURASIDONE IN BULK DRUG AND PHARMACEUTICAL DOSAGE FORM BY HPLC

Pawanjeet J. Chhabda1*

,

M. Balaji2, Srinivasarao .V

2, K.M.Ch.Appa Rao

2

1Department of Biochemistry, Ahmednagar College, Ahmednagar, MH,India

2Department of Chemistry, Gitam Institute of Science, GITAM University, Visakhapatnam, India.

ABSTRACT

A simple, precise, selective linear and accurate reverse phase HPLC

method was developed and validated for the assay determination

of Lurasidone in bulk drug and dosage form. Isocratic elution at

flow rate 1.0ml/min was employed on waters XBridge c18

(150x4.6) mm 5m at 30c temperature. The mobile phase

consisted of 0.1% perchloricacid: acetonitrile (50:50) (%v/v). The uv

detection wavelength was 230nm and 10l of sample injected. The

retention time for lurasidone was 5.60min and linearity was

observed in the concentration range of 30-225 g/ml with

correlation coefficient of 0.9999. The percentage relative standard

deviation in accuracy and precision studies was found to be less

than 2%. The method was successfully validated as per

International Conference on Harmonization (ICH) guidelines.

Lurasidone undergoes degradation under acidic, basic, oxidation,

dry heat and photolytic conditions, degradation impurities did not

interfere with the retention time of Lurasidone, and assay method

is thus stability indicating. The method was successfully applied for

routine analysis of lurasidone in bulk drug and dosage form.

Keywords:- Lurasidone.validation.HPLC.Stability indicating

INTRODUCTION Diabetes mellitus is regarded as a syndrome, a

Lurasidone is an atypical antipsychotic , It was

approved by the U.S. Food and Drug

Administration (FDA) for treatment

of schizophrenia on October 28, 2010 after a

review that found that two of the four Phase

III clinical trials supported efficacy, while one

showed only marginal efficacy and one was not

interpretable because of high drop-out rates. It is

currently pending approval for the treatment

of bipolar disorder in the United States, developed

by Dainippon Sumitomo Pharma(20). Each tablet

contains 20 mg, 40 mg, 80 mg, or 120 mg of

lurasidone hydrochloride market under the brand

name of LATUDA. Lurasidone is chemically

Correspondence Author

Pawanjeet J. Chhabda

Department of Biochemistry,

Ahmednagar College, Ahmednagar,

MH,India

Email: [email protected]

International Journal of Pharmaceutical Research & Development ISSN: 0974 9446


104

(3aR,4S,7R,7aS)-2-[[(1R,2R)-2-[[4-(1,2-

Benzisothiazol-3-yl)-1-piperazinyl] methyl]

cyclohexyl] methyl]hexahydro-4,7-methano-1H-

isoindole-1,3(2H)-dione hydrochloride with

empirical formula is C28H36N4O2S.HCl and molecular

weight 529.14 (21).

Various methods in the literatures involve

determination of Lurasidone in human, rat plasma

by LCMS/MS (1, 2), pharmacokinetics,

pharmacodynamics (3-13), UV Spectroscopy (14,

15) and HPLC (16, 17). However no method is

available for stability indicating method for assay of

Lurasidone in bulk drug and pharmaceutical dosage

form. In the present work we have developed a

new, simple precise and stability indicating method

for determination of Lurasidone in bulk drug and

pharmaceutical dosage form.

Figure 1: Structure of Lurasidone

Experimental

Chemicals & Reagents

Lurasidone is available as tablets with brand name

LATUDA was purchased from local market,

containing Lurasidone 120mg HPLC grade

acetonitrile, AR grade perchloric acid were

purchased from Merck, Mumbai. High pure water

was prepared by using Millipore Milli-Q plus

purification system.

Chromatographic Conditions

A Alliance e2695 separation module (Waters

corporation, Milford, MA) equipped with 2998 PDA

detector with empower 2 software used for

analysis. Buffer consisted of 0.1% perchloric acid in

water (1ml of perchloric acid in 1000 ml of water).

waters XBridge c18 (150x4.6) mm 5m column and

isocratic mixture of solution A (Buffer) solution B

(Acetonitrile)(50:50)(%v/v) used as stationary and

mobile phase respectively. Methanol used as

diluent. The column oven maintained at 30c with

1.0ml flow rate. An injection volume 10l was

used. The elution compounds were monitored at

230 nm.

Preparation of Stock and standard solutions

Accurately 150mg of Lurasidone standard dissolved

in 100ml diluent to get a concentration of

1500g/ml. Further 10ml of stock solution was

taken in 100ml flask and diluted up to the mark

with diluent to get concentration of 150g/ml.

Preparation of Tablets for assay

The formulation tablets of Latuda were crushed to

give finely powdered material. Powder equivalent

to 150mg of drug was weighed and transferred to

the 100ml flask added 10ml diluent and placed in

an ultrasonicator for 10minites made up to the

volume with diluent, and filtered through a 0.45m

nylon syringe filter. 10ml of this solution was taken

into 100 ml flask and diluted volume with diluent

to get concentration 150g/ml.

Forced Degradation studies

Acid Degradation studies

Acid decomposition was carried out in 0.1N HCL at

concentration of 1500g/ml Lurasidone and after

refluxation for 24hours at 80c, the stressed

sample was cooled, neutralized and diluted as per

requirement with diluents filtered and injected.

The resulting chromatogram is shown in fig.3 (g).

The results are tabulated in table 4.

Alkali Degradation studies

Base decomposition was carried out in 0.1N NaOH

at concentration of 1500g/ml Lurasidone after

refluxation for 24hours at 80c, the stressed

sample was cooled, neutralized and diluted as per

requirement with diluents filtered and injected.

The resulting chromatogram is shown in fig.3(i).

The results are tabulated in table 4.



105

Oxidation

Oxidation was conducted by using 4%H2O2

solution at room temperature for 5hours, 10ml of

solution was taken in 100ml flask and diluted up to

the mark with diluent to get concentration of

150g/ml filtered and injected. The resulting

chromatogram is shown in fig.3 (k). The results are

tabulated in table 4.

Temperature Stress studies

1g of Lurasidone sample was taken into a petridish

and kept in oven at 80c for 24hours. 150mg of

sample was taken into 100 ml flask diluted volume

with diluent, further 10ml to 100ml made up with

diluent. The results are tabulated in table 4.

Photo stability

1g of Lurasidone was taken in to a petridish and

kept in photo stability chamber 200 W.hr/m2

in UV

Fluorescent light and 1.2M LUX Fluorescent light.

150mg of sample was taken in 100ml flask,

dissolved in diluent, further 10ml in 100ml flask

diluted volume with diluent. The results are

tabulated in table 4.

RESULTS AND DISCUSSION

HPLC Method Development and Optimization

To develop a rugged and suitable HPLC assay

method for the determination of Lurasidone, the

analytical condition were selected after the

consideration of different parameters such as

diluents, buffer, organic solvent for mobile phase,

column and other chromatographic conditions

(19). Initial trails were performed with different

composition of buffer (acetate and formate) and

organic phase (methanol, teterhydrofuran) with

different column like c8,phenyl,cyno,amino and

basic but Lurasidone peak shape was not good.

Finally 0.1% perchoric acid and acetonitrile with

isocratic and waters XBridge c18 (150x4.6) mm 5

m column was optimized. Different diluents were

tried to dilute sample like water,buffer,

tetrahydrofuran and mixture of water: methanol

and water: teterhydrofuran, buffer:methanol and

buffer:acetonitrile. Lurasidone was not dissolved,

finally methanol was optimized. The detection

wavelength was chosen as 230nm for Lurasidone

because they have better absorption and

sensitivity at this wavelength (fig-2). Hence

selected method was best among the all trails by

many aspects.

Fig-2 wavelength spectrum of Lurasidone

Method Validation

Specificity

A study to establish the interference, blank

detection was conducted. Diluent was injected as

per the test method. Solution of standard and

sample were prepared as per test method and

injected into the chromatographic system. The

chromatograms of blank, standard and sample

were shown in the fig a, b, c.

Precision

The precision for assay method was established by

evaluating method precision and intermediate

precision study. Method precision was determined

203.3

231.4

315. 3369.8 384.2

AU

0.00

0.05

nm

20 0.00 220.00 240.00 260.00 280.00 300.00 320.00 340.00 360.00 380.00



106

by analyzing six independent assays were

performed and calculated the % RSD for replicate

assay determinations. Intermediate precision of

the analytical method was determined by

conducting method precision on another day and

another analyst under same experiment condition.

The result obtained for method precision and

intermediate precision are shown in table 3. The

percentage of RSD was calculated. The %RSD range

was obtained as 0.18 and 0.29 for method

precision and intermediate precision respectively

(Table 3) which is less than 2% indicating that the

method is more precise.

Accuracy

The accuracy of the method was estimated by

determination of recovery for three concentrations

(corresponding to 50,100 and 150% of test solution

concentration) covering the range of the method.

For each concentration three sets were prepared

and injected. The drug concentrations of

Lurasidone were calculated, the results obtained

are shown in table 2.The percentage recovery was

found to be 99.77-99.96% with %RSD 0.03 -

0.21(

International Journal of Pharmaceutical Research & Development



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(b)

(c)

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(e)

(f)

(g)

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(h)

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Fig-3 Typical chromatograms of (a) Blank (b) Standard (c) Sample (d) precision injections (e) Linearity injections

(f) Acid blank (g) Acid sample (h) Base blank (i) Base sample (j) Peroxide blank (k) Peroxide sample

of Acid (m) Purity plot of Base (n) Purity plot of Peroxide

Lura

sido

ne - 5.

578

P u r i t yA u t o T h re s h o l d

AU

- 0 . 0 2

0 . 0 0

0 . 0 2

0 . 0 4

0 . 0 6

0 . 0 8

0 . 1 0

0 . 1 2

0 . 1 4

0 . 1 6

0 . 1 8

0 . 2 0

0 . 2 2

0 . 2 4

0 . 2 6

5 . 3 0 5 .3 5 5 . 4 0 5 .4 5 5 . 5 0 5 .5 5 5 . 6 0

P u r i t yA u t o T h r e s h o ld

AU

- 0 . 0 2

0 . 0 0

0 . 0 2

0 . 0 4

0 . 0 6

0 . 0 8

0 . 1 0

0 . 1 2

0 . 1 4

0 . 1 6

0 . 1 8

0 . 2 0

0 . 2 2

0 . 2 4

5 . 3 5 5 . 4 0 5 . 4 5 5 . 5 0

Lura

sido

ne - 5.

575

P u r i t yA u t o T h re s h o ld

AU

- 0 . 0 2

0 . 0 0

0 . 0 2

0 . 0 4

0 . 0 6

0 . 0 8

0 . 1 0

0 . 1 2

0 . 1 4

0 . 1 6

0 . 1 8

0 . 2 0

0 . 2 2

0 . 2 4

5 . 3 0 5 .4 0 5 .5 0 5 . 6 0


(k)

(l)

(m)

(n)

3 Typical chromatograms of (a) Blank (b) Standard (c) Sample (d) precision injections (e) Linearity injections

(f) Acid blank (g) Acid sample (h) Base blank (i) Base sample (j) Peroxide blank (k) Peroxide sample

Purity plot of Base (n) Purity plot of Peroxide

M in u t e s5 . 6 0 5 . 6 5 5 . 7 0 5 . 7 5 5 . 8 0 5 . 8 5 5 . 9 0 5 . 9 5 6 . 0 0 6 . 0 5 6 . 1 0 6 . 1 5

Lura

sido

ne - 5.

567

M in u t e s5 . 5 5 5 . 6 0 5 . 6 5 5 . 7 0 5 . 7 5 5 . 8 0 5 . 8 5 5 . 9 0 5 . 9 5

M in u t e s5 . 7 0 5 .8 0 5 .9 0 6 . 0 0 6 . 1 0 6 .2 0 6 .3 0

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3 Typical chromatograms of (a) Blank (b) Standard (c) Sample (d) precision injections (e) Linearity injections

(f) Acid blank (g) Acid sample (h) Base blank (i) Base sample (j) Peroxide blank (k) Peroxide sample (l)Purity plot

De

gre

es

0 . 0 0

1 0 . 0 0

2 0 . 0 0

3 0 . 0 0

4 0 . 0 0

5 0 . 0 0

6 0 . 0 0

7 0 . 0 0

8 0 . 0 0

6 . 2 0 6 . 2 5 6 . 3 0

De

gre

es

0 . 0 0

0 . 5 0

1 . 0 0

1 . 5 0

2 . 0 0

2 . 5 0

3 . 0 0

3 . 5 0

4 . 0 0

5 . 9 5 6 . 0 0 6 . 0 5

De

gree

s

0 .0 0

1 0 .0 0

2 0 .0 0

3 0 .0 0

4 0 .0 0

5 0 .0 0

6 0 .0 0

7 0 .0 0

8 0 .0 0

9 0 .0 0

6 .3 0 6 . 4 0



111

Fig-4 Linearity of Lurasidone

Table-1 Results for linearity of Lurasidone

Table-2 Recoveries study for Lurasidone

Accuracy (Recovery) study

Accuracy

Level

Set No Amount

Added

Amount

Found

Recovery

(%)

Average

recovery

Std

Dev.

%

(g/ml) (g/ml) RSD

1 75.06 74.84 99.71

50% 2 75.16 75.26 100 99.77 0.21 0.21

3 75.08 74.78 99.6

1 150.04 149.8 99.84

100% 2 150.16 150 99.89 99.87 0.03 0.03

3 150.2 150.04 99.89

1 225.06 225.08 100

150% 2 225.2 224.98 99.9 99.96 0.05 0.05

3 225.14 225.1 99.98

Linearity

level

%Level Area

1 20 517237

2 40 1064284

3 80 2171507

4 100 2766401

5 120 3313426

6 150 4188742

Correlation co-efficient 0.999933

intercept -62723.4

slope 28231.25

y = 92529x - 12579R = 0.999

02000000400000060000008000000

10000000120000001400000016000000

0 50 100 150 200

Me

an

Area

Concentration(g/mL)



112

Table-3 Precision results for Lurasidone

Study Set no Assay (%) Mean

assay(%) Stdev RSD%

1 100.02

2 100.15

Method precision 3 99.78

4 99.86 100.03 0.18 0.18

5 100.1

6 100.24

1 99.75

2 100.22

Intermediate

precision 3 100.34

4 99.88 99.94 0.29 0.29

5 99.9

6 99.55

Table-4 forced degradation results for Lurasidone

Stress condition Drug recovered (%) Drug decomposed (%)

Standard drug 100

Acid degradation 99.57 0.43

Alkali degradation 83.81 16.19

Oxidation degradation 89.81 10.19

Thermal degradation 99.89 0.11

Photolytic degradation 99.90 0.10

Table -5 Robustness results for Lurasidone

Robust conditions variation Retention

time(min) USP Tailing

USP Plate

count

0.9ml 6.22 1.37 4235

Flow 1.0ml 5.6 1.3 4450

1.1ml 5.01 1.27 4568

25c 5.75 1.34 4325

Temperature 30c 5.6 1.3 4450

35c 5.25 1.24 4579

45 5.95 1.34 4321

%Acetonitrile 50 5.6 1.3 4450

55 4.98 1.21 4589



113

CONCLUSIONS

A validated RP-HPLC method has been developed

for determination of Lurasidone in presence of

degradation impurities . The proposed method was

found to be a new, simple, precise, linear, accurate

and specific. Degradation impurities did not

interfere with the retention time of Lurasidone,

and assay method is thus stability indicating.

ACKNOWLEDGEMENTS

The authors are grateful of M/S GITAM Institute of

Science, GITAM University, Visakhapatnam, India

for providing research facilities.

.

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*****

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