TIZANIDINE HYDROCHLORIDE METHOD OF ANALYSIS SOP

TIZANIDINE HYDROCHLORIDE METHOD OF ANALYSIS SOP

1.0  OBJECTIVE:

To lay down a procedure of analytical report for raw active material of the Tizanidine hydrochloride from the Pharmacopoeial specification.

2.0  SCOPE:

This SOP shall be applicable in Q.C laboratory.

3.0  RESPONSIBILITY:

3.1  Q.C Analysts.

4.0  ACCOUNTABILITY:

4.1  Q.C Manager

5.0  PROCEDURE:

5.1  Identification tests:

5.1.1        Chlorides test:

5.1.1.1  Material and equipment:

5.1.1.1.1        Centrifuge machine.

5.1.1.1.2        Diluted nitric acid.

5.1.1.1.3        Purified water.

5.1.1.1.4        Silver nitrate.

5.1.1.1.5        Ammonia.

5.1.1.1.6        Excess of 6N ammonium hydroxide.

5.1.1.2  Sample:

5.1.1.2.1        10mg/ml.

5.1.1.3  Method:

5.1.1.3.1        Take a test tube add in it 10mg/ml with the help of pipette.

5.1.1.3.2        Add silver nitrate. Observe the changes.

5.1.1.3.3        A white, curdy ppt is formed that is insoluble in nitric acid but is soluble in a slight excess of 6N ammonium hydroxide.

5.1.1.3.4        Observe the changes.

5.1.1.4  Observations:

5.1.1.4.1        A white, curdy ppt is soluble in a slight excess of 6N ammonium hydroxide.

5.2  pH determination test:

5.2.1        Material and equipment:

5.2.1.1  Glassware (according to the requirement).

5.2.1.2  pH meter.

5.2.2        Sample:

5.2.2.1  10mg/ml.

5.2.3        Method:

5.2.3.1  Firstly clean the pH meter with clean dry cloth, according to SOP

5.2.3.2  Operate the pH meter according to SOP for operation of pH meter i.e. SOP

5.2.3.3  Rejuvenate the electrode before use according to SOP instructions, if there is any need of.

5.2.3.4  Calibrate the electrode of the pH meter according to SOP

5.2.3.5  Perform the test on 10mg/ml solution.

5.2.3.6  Take a beaker of 100.0ml and add 10mg/ml solution in it. Such that it immersed electrodes in it completely.

5.2.3.7  Maintain the temperature of sample at 25^oC±2^oC.

5.2.3.8  Dip the electrode along with temperature sensor into the sample.

5.2.3.9  When dipping electrode into sample, it must be completely immersed in it.

5.2.3.10 Stir the probe gently in the sample to create a homogeneous sample.

5.2.3.11 Allow the reading to stabilize for a time.

5.2.3.12 Record the observed values of pH & temperature in the respective Annexure-1.

5.2.3.13    Wash the electrodes again after use and store the electrode in storage solution as recommended in SOP of cleaning i.e. SOP. Calibrate the instrument before use according to SOP.

5.2.4        Observation:

5.2.4.1  4.3-5.3.

5.3  Loss on drying:

5.3.1        Material and equipment:

5.3.1.1  Glassware (according to requirement).

5.3.1.2  Analytical weighing balance.

5.3.1.3  Oven.

5.3.2        Sample:

5.3.2.1  0.5g.

5.3.3        Method:

5.3.3.1  Weigh 0.5g of the test sample.

5.3.3.2  Set the oven apparatus. Operate it according to the SOP

5.3.3.3  Place the sample into the tray and dry it.

5.3.3.4  Set the temperature of oven at 105^oC for at least 3 hours.

5.3.3.5  And wait till the sample loses its moisture.

5.3.3.6  After 3 hours weigh the sample again by using analytical weighing balance i.e. the final weight.

5.3.3.7  Note down readings on given Annexure-2

5.3.4        Observation:

5.3.4.1  NMT 0.5%.

5.4  Assay:

5.4.1        Buffer:

5.4.1.1  Take beaker of 1000.0ml add 6.8g/L of monobasic potassium phosphate.

5.4.1.2  Adjust pH with5.3N potassium hydroxide to a pH of 7.5±0.05.

5.4.2        Mobile phase:

5.4.2.1  Take a beaker of 100.0ml.

5.4.2.2  Add acetonitrile and buffer (20:80), stir by using magnetic stirrer.

5.4.3        System suitability solution:

5.4.3.1  46μg/ml of USP Tizanidine hydrochloride RS and 0.12μg/ml of USP Tizanidine related compound C RS in mobile phase.

5.4.4        Standard solution:

5.4.4.1  0.046mg/ml of USP Tizanidine hydrochloride RS in mobile phase.

5.4.5        Sample solution:

5.4.5.1  0.046mg/ml of Tizanidine hydrochloride in mobile phase.

5.4.6        Chromatographic system:

5.4.6.1  Mode: Liquid chromatography.

5.4.6.2  Detector: UV 230nm.

5.4.6.3  Column: 4.6mm x 15cm; packing L7.

5.4.6.4  Column temperature: 35^o.

5.4.6.5  Flow rate: 1.0ml/min.

5.4.6.6  Injection size: 20μL.

5.4.7        System suitability:

5.4.7.1  Samples: System suitability solution and Standard solution.

[NOTE: The relative retention times for Tizanidine related compound C and Tizanidine are 0.5 and 0.1, respectively.]

5.4.7.2  Suitability requirements:

5.4.7.2.1        Resolution: NLT 6 between Tizanidine and Tizanidine related compound C, System suitability solution.

5.4.7.2.2        Tailing factor: NMT 2.0 for Tizanidine, standard solution.

5.4.7.2.3        Relative standard deviation: NMT 2.0%, standard solution.

5.4.8        Analysis:

5.4.8.1  Samples: Standard solution and sample solution.

5.4.9        Calculate the percentage of Tizanidine  hydrochloride (C₉H₈ClN₅S.HCl) in the portion of Tizanidine  hydrochloride taken:

Result: (r_U/r_S) x (C_S/C_U) x 100

r_U= peak area of Tizanidine from the sample solution.

r_S= peak area of Tizanidine from the standard solution.

C_S= concentration of USP of Tizanidine hydrochloride RS in the standard solution (mg/ml).

C_U= concentration of the sample solution (mg/ml).

5.4.10    Equilibrate the column and detector with mobile phase at specified flow rate until a constant signal is received.

5.4.11    Separately inject equal volumes about 20μL of the standard solution and sample solution.

5.4.12    Record the chromatogram.

5.4.13    Measure the responses for the major peaks.

5.4.14    Analyze as directed in the monograph.

5.5  Limit:

5.5.1        98.0 to 102.0% on the dried basis.

6.0  REVISION LOG:

Revision No.	Effective Date	Reason
00		New SOP

7.0  REFERENCES:

7.1  USP38NF33 Volume-6 Official Monograph/ Tizanidine hydrochloride: 2015, pp.: 5590-5591.

8.0  ANNEXURES:

Annexure 1: pH measurement.

Annexure 2: Observations of Percentage Loss of drying by using oven.

Annexure 3: Observations and calculations of HPLC method.

Annexure: 1

pH measurement

Sr.#	Temperature	pH

Result: _________________________________________________________________

Annexure: 2

Observations of percentage loss of drying by using Oven

Percentage loss of drying by using Oven Weight of Sample = _____________ Time period = _____________ Pressure= _________________ Sr.# Time (min) Weight of sample (g) % Loss of Moisture Initial weight Final weight Average % Loss of Moisture: _____________ % Loss of Moisture: Remarks: _______________________________________________________________

Annexure: 3

Observations and calculations of HPLC method

Analysis on HPLC Instrument: ___________________                                           Date: _________________ Model: ___________________ Column size: Length= θ= Stationary phase: Temperature: Mobile phase: Flow rate: Injection size: Detector: Wavelength: λ= Sample solution: _______________________ Reference standard solution: ______________ Impurities: ____________________________ (calculate each component calculation separately) OBSERVATIONS: Attach chromatogram. CALCULATIONS: 1.      Retention time:                                                                                n= no. of peak Retention time of unretained peak (tm)= _____________ No. of peaks Retention time of peak of interest (tr)n Height of peak of interest (h)n Width of peak of interest (w)n Area of peak of interest A=1/2(h x w) 2.      Retention volume: Flow rate= _______________ml/min. No. of peaks Retention time of peak of interest (tr)n Retention volume = retention time x flow rate 3.      Retention factor: Retention time of unretained peak (tm)= _____________ No. of peaks Retention time of peak of interest (tr)n Retention factor of a component k= (tr-tm)/tm 4.      Separation factor (α): No. of peaks Retention factor of a component (kn) Relative retention of two adjacent peaks α = k2/k1 5.      Resolution: Retention time of unretained peak (tm)= _____________ No. of peaks Retention time of peak of interest (tr)n Width of peak of interest (w)n Resolution Rs = 2 (tr2-tr1)         (w1-w2) 6.      Efficiency: No. of peaks or components Retention time of peak of interest (tr)n Width of peak of interest (w)n Efficiency (No. of theoretical plates) N= 16 (tr/w)2 7.      Height equivalent to a theoretical plate (HETP): Length of column = ________________________ No. of peaks or components No. of theoretical plates (N) Height equivalent to a theoretical plate HETP = L/N 8.      Symmetry factor (tailing factor): No. of peaks or components Distance from the peak max. to leading edge of the peak (f) Width w Symmetry factor At 5% At 10% As = w5%        2f As = w10%        2f 9.      Response factor & Relative response factor: Conc. (mg/ml)= ___________________ No. of peak Peak area Response factor = (peak area/conc.) Relative response factor = (response factor of impurity/response factor of API) 10.  Relative standard deviation (%RSD): Use formula of relative standard deviation where it is required i.e., pic 11.  Percentage of content: Percentage content = (rU/rS) x (CS/CU) x 100. rU= peak response of substance from the sample solution. rS= peak response of substance from the standard solution. CS= concentration of substance in the standard solution (mg/mL). CU= concentration of substance in the sample solution (mg/mL). RESULTS: ________________________________________________________________________________________________________________________________________________

9.0  ABBREVIATIONS:

Abbreviation	Expanded Form
SOP	Standard operating procedure
&	And
No.	Number
Ltd.	Limited
Sr.#	Serial number
Q.C	Quality control
Vol	Volume
g	Grams
ml	Milliliters
ppt	Precipitates
mg	Milligrams
g/L	Gram per liter
mg/ml	Milligram per milliliter
UV	Ultra violet
USP	United states pharmacopoeia
nm	Nanometer
mm	Millimeter
cm	Centimeter
μm	Micron
oC	Degree centigrade
ml/min.	Milliliter per minute
μL	Microliter
NMT	Not more than
%	Percentage
NF	National formulary
QCA	Quality control active ingredient
F	Format