System suitability parameters assessment by HPLC using acetone, benzene, toluene, based on polarity.

1. SYSTEM SUITABILITY PARAMETERS
ASSESMENT BY HPLC USING ACETONE,
BENZENE & TOLUENE
Presented by:
Anirban Barik.
4th Year, 7th semester
Roll no.:15901916091
Registration no.:161590210009 of 2016-17
Under the Guidance of:
Mr. Soumya Ganguly.
Dept. Of Pharmaceutics.
Calcutta Institute of Pharmaceutical
Technology and Allied Health Sciences
Banitabla,Uluberia,Howrah-711316
West Bengal. 2019

2. CONTENTS
• Introduction
• System Suitability
• Aim & Objective
• Method
• Result
• Observed chromatogram
• Discussion
• Conclusion
• References

3. INTRODUCTION
• HPLC stands for high performance liquid chromatography or
high pressure liquid chromatography.
• It is a form of liquid chromatography used to separate
compounds that are dissolved in solution.
• HPLC instruments consists of a solvent reservoir, pump, an
injector, a column, a detector and a data collection device.
• Used in biochemistry to identify, quantify and purify the
individual components of a mixture.
• Our project is based on reversed phase chromatography:
stationary phase is non-polar (hydrophobic) and mobile
phase is polar (hydrophilic). Main principle of separation is
adsorption based on polarity.

4. SYSTEM SUITABILITY
• System suitability are used to verify that the resolution and
reproducibility of the chromatographic system are adequate for the
analysis to be done.
• Some parameters which can be checked using the System
Suitability Testing are:
1. Resolution (R) =
𝟐.𝟎(𝑹𝒕 𝟐−𝑹𝒕 𝟏)
(𝑾 𝟐+𝑾 𝟏)
where, 𝑅𝑡2=Retention time for peak 2
𝑅𝑡1 = Retention time for peak 1
(𝑤1+𝑤2)= sum of peak widths at
baseline between tangent lines.
2. Plate count (N)= 𝟏𝟔(
𝒕
𝒘
) 𝟐
where, t=Retention time
w=Width of the peak at baseline

5. .
3. Retention time: It is important that retention time be fairly constant
because the data system uses retention time to identify peaks.
4. Pressure: The suitability testing must be carried out within set pressure
limits.
5. Tailing factor (T) =
𝑾 𝟎.𝟎𝟓
𝟐𝒇
where, 𝑤0.05 = width of the peak at 0.05h
f = width between left tangent
and the line of maximum peak.
• Table : The current FDA values for the Validation of Chromatographic Methods.

6. • Aim: To perform system suitability check for HPLC column
(Zorbax XDB- 150mm x 4.6mm x 5µm).
• Objective: To assess the changes in USP Resolution, USP Tailing
& theoretical plate of HPLC column (Zorbax XDB- 150mm x
4.6mm x 5µm) by using a mixture (Acetone, Benzene, Toluene)
upon changing the composition of mobile phase.
AIM & OBJECTIVE

7. 1. Preparation of mobile phase:
a) Water and acetonitrile are taken in a ratio of – water :
acetonitrile = 50 : 50. Total 500 ml solution is prepared.
b) Water and acetonitrile are taken in a ratio of – water :
acetonitrile = 40 : 60. Total 500 ml solution is prepared.
c) Sonicated.
2. Preparation of solvent mixture to be separated:
Acetone : 10 µl
Benzene : 10 µl
Toluene : 10 µl
Methanol : q.s. to 10 ml
3. 5 µl of solution is taken in syringe and injected in the rheodyne
injector.
METHOD

8. • Table 1: Reproducibility Table
Instrument parameters :
• Mobile phase – Acetonitrile : water
:: 50 : 50 (v/v)
• Detection wavelength – 254 nm
• Injection volume – 5 µl
• Column : Zorbax XDB- 150mm x
4.6mm x 5µm
• Run time – 10 min
• Table 2: Reproducibility Table
Instrument parameters :
• Mobile phase – Acetonitrile : water
:: 60 : 40 (v/v)
• Detection wavelength – 254 nm
• Injection volume – 5 µl
• Column : Zorbax XDB- 150mm x
4.6mm x 5µm
• Run time – 6 min
RESULT
Sample Retention time (min)
Inj. 1 Inj. 2 Inj. 3
Acetone 1.621 1.621 1.619
Benzene 4.627 4.620 4.601
Toluene 6.920 6.900 6.895
Sample Retention time (min)
Inj. 1 Inj. 2 Inj. 3
Acetone 1.591 1.596 1.596
Benzene 3.238 3.237 3.239
Toluene 4.328 4.327 4.329

9. • Table 3: System Suitability
Acetonitrile : Water :: 50 : 50(V/V) Acetonitrile : Water :: 60 : 40(V/V)
RESULT
Mobile phase ratio USP Resolution USP Tailing Theoretical plate
Acetone -
Benzene
Benzene -
Toluene
Benzene Toluene Benzene Toluene
Acetonitrile : water ::
50 : 50 (v/v)
5.23 3.28 1.17 1.13 951.52 1197.16
Acetonitrile : water ::
60 : 40 (v/v)
2.05 1.28 1.25 1.10 262.28 370.18

10.  Effect of change of mobile phase ratio on USP resolution,
USP tailing and theoretical plate.
• Graphical representation of USP Resolution :
5.23
3.28
2.05
1.28
0
1
2
3
4
5
6
Acetone - Benzene Benzene - Toluene
USPResolution
USP Resolution
ACN: H2O::50:50(V/V) ACN: H2O::60:40(V/V)

11. Contd.
• Graphical representation of USP Tailing :
1.17
1.13
1.25
1.1
1
1.05
1.1
1.15
1.2
1.25
1.3
Benzene Toluene
USPTailing
USP Tailing
ACN: H2O::50:50(V/V) ACN: H2O::60:40(V/V)

12. Contd.
• Graphical representation of Theoretical plates :
951.52
1197.16
262.28
370.18
0
200
400
600
800
1000
1200
1400
Benzene Toluene
Theoreticalplate
Theoretical plate
ACN: H2O::50:50(V/V) ACN: H2O::60:40(V/V)

13. • MOBILE PHASE: Water : acetonitrile :: 50 : 50 (v/v)
Run : Blank Run : 1
OBSERVED CHROMATOGRAM
I n je c t io n S u m m a r y R e p o r t
P r o je c t N a m e : C IP TR e p o r te d b y U s e r : S y s te m
R e p o r t M e th o d : In je c tio n S u m m a r y R e p o r t D a te P r in te d :
3 4 8 9 1 0 /1 9 /2 0 1 9R e p o r t M e th o d ID : 3 4 8 9
1 2 :0 2 :5 2 P M A s ia /C a lc u ttaP a g e : 1 o f 1
S A M P L E I N F O R M A T I O N
S y s te mA c q u ir e d B y :A B _ A c e B e n z T _ 2 7 0 9 2 0 1 9S a m p le N a m e :
S a m p le S e t N a m e :U n k n o w nS a m p le T y p e :
1V ia l: A c q . M e th o d S e t: B P h a r m _ A B _ A B T _ 2 6 0 9 1 9
6In je c tio n # : P r o c e s s in g M e th o d : A c e B e n z T _ A B _ A C N 5 0
5 .0 0 u lIn je c tio n V o lu m e : C h a n n e l N a m e : W 2 4 8 9 C h A
R u n T im e : 1 0 .0 M in u te s P r o c . C h n l. D e s c r .: W 2 4 8 9 C h A 2 5 4 n m
9 /2 8 /2 0 1 9 1 :5 6 :5 9 A M IS TD a te A c q u ir e d :
D a te P r o c e s s e d : 1 0 /1 9 /2 0 1 9 1 1 :4 4 :4 9 A M IS T
C h a n n e l: W 2 4 8 9 C h A ; P r o c e s s e d C h a n n e l: W 2 4 8 9 C h A 2 5 4 n m ; R e s u lt Id : 3 4 6 8 ; P r o c e s s in g M e th o d :
A c e B e n z T _ A B _ A C N 5 0
Acetone-1.621
Benzene-4.627
Tolune-6.920
AU
0 . 0 0
0 . 1 0
0 . 2 0
0 . 3 0
M in u t e s
0 . 0 0 1 . 0 0 2 . 0 0 3 . 0 0 4 . 0 0 5 . 0 0 6 . 0 0 7 . 0 0 8 . 0 0 9 . 0 0 1 0 . 0 0
1
2
3
P r o c e s s e d
C h a n n e l D e s c r .
P e a k N a m e R T A r e a % A r e a H e ig h t
W 2 4 8 9 C h A 2 5 4 n m
W 2 4 8 9 C h A 2 5 4 n m
W 2 4 8 9 C h A 2 5 4 n m
A c e to n e
B e n z e n e
T o lu n e
1 .6 2 1
4 .6 2 7
6 .9 2 0
2 4 1 6 9 1
1 8 7 7 0 7 9
2 2 6 8 3 1 7
5 .5 1
4 2 .7 9
5 1 .7 0
3 0 5 3 2
2 3 6 6 3 2
2 1 3 4 4 1
P r o c e s s e d C h a n n e l D e s c r . : W 2 4 8 9 C h A 2 5 4 n m
I n je c t io n S u m m a r y R e p o r t
P r o je c t N a m e : C IP TR e p o r te d b y U s e r : S y s te m
R e p o r t M e th o d : In je c tio n S u m m a r y R e p o r t D a te P r in te d :
3 4 8 9 1 0 /1 9 /2 0 1 9R e p o r t M e th o d ID : 3 4 8 9
1 2 :0 2 :3 0 P M A s ia /C a lc u ttaP a g e : 1 o f 1
S A M P L E I N F O R M A T I O N
S y s te mA c q u ir e d B y :A B _ B la n k _ 2 7 0 9 2 0 1 9S a m p le N a m e :
S a m p le S e t N a m e :U n k n o w nS a m p le T y p e :
1V ia l: A c q . M e th o d S e t: B P h a r m _ A B _ A B T _ 2 6 0 9 1 9
5In je c tio n # : P r o c e s s in g M e th o d : A c e B e n z T _ A B _ A C N 5 0
5 .0 0 u lIn je c tio n V o lu m e : C h a n n e l N a m e : W 2 4 8 9 C h A
R u n T im e : 1 0 .0 M in u te s P r o c . C h n l. D e s c r .: W 2 4 8 9 C h A 2 5 4 n m
9 /2 8 /2 0 1 9 1 :4 2 :1 9 A M IS TD a te A c q u ir e d :
D a te P r o c e s s e d : 1 0 /1 9 /2 0 1 9 1 1 :4 8 :3 7 A M IS T
C h a n n e l: W 2 4 8 9 C h A ; P r o c e s s e d C h a n n e l: W 2 4 8 9 C h A 2 5 4 n m ; R e s u lt Id : 3 4 7 7 ; P r o c e s s in g M e th o d :
A c e B e n z T _ A B _ A C N 5 0
AU
0 . 0 0
0 . 1 0
0 . 2 0
0 . 3 0
M in u t e s
0 . 0 0 1 . 0 0 2 . 0 0 3 . 0 0 4 . 0 0 5 . 0 0 6 . 0 0 7 . 0 0 8 . 0 0 9 . 0 0 1 0 . 0 0
1
2
3
P r o c e s s e d
C h a n n e l D e s c r .
P e a k N a m e R T
W 2 4 8 9 C h A 2 5 4 n m
W 2 4 8 9 C h A 2 5 4 n m
W 2 4 8 9 C h A 2 5 4 n m
A c e to n e
B e n z e n e
T o lu n e
1 .6 0 0
4 .6 5 0
6 .9 0 0
P r o c e s s e d C h a n n e l D e s c r . :
W 2 4 8 9 C h A 2 5 4 n m

14. MOBILE PHASE: Water : acetonitrile :: 50 : 50 (v/v)
Run : 2 Run : 3
In je c tio n S u m m a r y R e p o r t
P r o je c t N a m e : C IP TR e p o r te d b y U s e r : S y s te m
R e p o r t M e th o d : In je c tio n S u m m a r y R e p o r t D a te Pr in te d :
3 4 8 9 1 0 /1 9 /2 0 1 9R e p o r t M e th o d ID : 3 4 8 9
1 2 :0 3 :0 8 PM A s ia /C a lc u ttaP a g e : 1 o f 1
S A M P L E I N F O R M A T I O N
S y s te mA c q u ir e d B y :A B _ A c e B e n z T _ 2 _ 2 7 0 9 2 0 1 9S a m p le N a m e :
S a m p le S e t N a m e :U n k n o w nS a m p le T y p e :
1V ia l: A c q . M e th o d S e t: B P h a r m _ A B _ A B T _ 2 6 0 9 1 9
7In je c tio n # : P r o c e s s in g M e th o d : A c e B e n z T _ A B _ A C N 5 0
5 .0 0 u lIn je c tio n V o lu m e : C h a n n e l N a m e : W 2 4 8 9 C h A
R u n T im e : 1 0 .0 M in u te s P r o c . C h n l. D e s c r .: W 2 4 8 9 C h A 2 5 4 n m
9 /2 8 /2 0 1 9 2 :1 0 :3 5 A M IS TD a te A c q u ir e d :
D a te Pr o c e s s e d : 1 0 /1 9 /2 0 1 9 1 1 :4 4 :4 9 A M IS T
C h a n n e l: W 2 4 8 9 C h A ; P r o c e s s e d C h a n n e l: W 2 4 8 9 C h A 2 5 4 n m ; R e s u lt Id : 3 4 6 9 ; P r o c e s s in g M e th o d :
A c e B e n z T _ A B _ A C N 5 0
Acetone-1.621
Benzene-4.620
Tolune-6.900
AU
0 . 0 0
0 . 1 0
0 . 2 0
0 . 3 0
M in u t e s
0 . 0 0 1 . 0 0 2 . 0 0 3 . 0 0 4 . 0 0 5 . 0 0 6 . 0 0 7 . 0 0 8 . 0 0 9 . 0 0 1 0 . 0 0
1
2
3
P r o c e s s e d
C h a n n e l D e s c r .
P e a k N a m e R T A r e a % A r e a H e ig h t
W 2 4 8 9 C h A 2 5 4 n m
W 2 4 8 9 C h A 2 5 4 n m
W 2 4 8 9 C h A 2 5 4 n m
A c e to n e
B e n z e n e
T o lu n e
1 .6 2 1
4 .6 2 0
6 .9 0 0
2 6 2 5 0 1
2 0 2 8 2 7 7
2 4 6 0 6 8 8
5 .5 2
4 2 .6 9
5 1 .7 9
3 3 0 5 4
2 5 5 8 1 2
2 3 1 6 1 0
P r o c e s s e d C h a n n e l D e s c r .: W 2 4 8 9 C h A 2 5 4 n m
In je c tio n S u m m a r y R e p o r t
P r o je c t N a m e : C IP TR e p o r te d b y U s e r : S y s te m
R e p o r t M e th o d : In je c tio n S u m m a r y R e p o r t D a te Pr in te d :
3 4 8 9 1 0 /1 9 /2 0 1 9R e p o r t M e th o d ID : 3 4 8 9
1 2 :0 3 :2 0 PM A s ia /C a lc u ttaP a g e : 1 o f 1
S A M P L E I N F O R M A T I O N
S y s te mA c q u ir e d B y :A B _ A c e B e n z T _ 3 _ 2 7 0 9 2 0 1 9S a m p le N a m e :
S a m p le S e t N a m e :U n k n o w nS a m p le T y p e :
1V ia l: A c q . M e th o d S e t: B P h a r m _ A B _ A B T _ 2 6 0 9 1 9
8In je c tio n # : P r o c e s s in g M e th o d : A c e B e n z T _ A B _ A C N 5 0
5 .0 0 u lIn je c tio n V o lu m e : C h a n n e l N a m e : W 2 4 8 9 C h A
R u n T im e : 1 0 .0 M in u te s P r o c . C h n l. D e s c r .: W 2 4 8 9 C h A 2 5 4 n m
9 /2 8 /2 0 1 9 2 :2 4 :1 1 A M IS TD a te A c q u ir e d :
D a te Pr o c e s s e d : 1 0 /1 9 /2 0 1 9 1 1 :4 4 :4 9 A M IS T
C h a n n e l: W 2 4 8 9 C h A ; P r o c e s s e d C h a n n e l: W 2 4 8 9 C h A 2 5 4 n m ; R e s u lt Id : 3 4 7 0 ; P r o c e s s in g M e th o d :
A c e B e n z T _ A B _ A C N 5 0
Acetone-1.619
Benzene-4.601
Tolune-6.895
AU
0 . 0 0
0 . 1 0
0 . 2 0
0 . 3 0
M in u t e s
0 . 0 0 1 . 0 0 2 . 0 0 3 . 0 0 4 . 0 0 5 . 0 0 6 . 0 0 7 . 0 0 8 . 0 0 9 . 0 0 1 0 . 0 0
1
2
3
P r o c e s s e d
C h a n n e l D e s c r .
P e a k N a m e R T A r e a % A r e a H e ig h t
W 2 4 8 9 C h A 2 5 4 n m
W 2 4 8 9 C h A 2 5 4 n m
W 2 4 8 9 C h A 2 5 4 n m
A c e to n e
B e n z e n e
T o lu n e
1 .6 1 9
4 .6 0 1
6 .8 9 5
2 4 9 5 6 9
1 9 3 0 6 5 4
2 3 3 6 2 8 9
5 .5 3
4 2 .7 5
5 1 .7 3
3 2 4 6 1
2 4 5 2 6 4
2 2 0 8 8 6
P r o c e s s e d C h a n n e l D e s c r .: W 2 4 8 9 C h A 2 5 4 n m

15. MOBILE PHASE: Water : acetonitrile :: 40 : 60 (v/v)
Run : Blank Run : 1
In je c tio n S u m m a r y R e p o r t
P r o je c t N a m e : C IP TR e p o r te d b y U s e r : S y s te m
R e p o r t M e th o d : In je c tio n S u m m a r y R e p o r t D a te Pr in te d :
3 4 8 9 1 0 /1 9 /2 0 1 9R e p o r t M e th o d ID : 3 4 8 9
1 1 :5 4 :5 7 A M A s ia /C a lc u ttaP a g e : 1 o f 1
S A M P L E I N F O R M A T I O N
S y s te mA c q u ir e d B y :A B _ B la n k _ 0 3 1 0 1 9 _ A C N 6 0S a m p le N a m e :
S a m p le S e t N a m e :U n k n o w nS a m p le T y p e :
1V ia l: A c q . M e th o d S e t: B P h a r m _ S R _ A c e A c e n a p _ 0 3 1 0 1 9
7In je c tio n # : P r o c e s s in g M e th o d : A c e B e n z T _ A B _ A C N 6 0
5 .0 0 u lIn je c tio n V o lu m e : C h a n n e l N a m e : W 2 4 8 9 C h A
R u n T im e : 1 0 .0 M in u te s P r o c . C h n l. D e s c r .: W 2 4 8 9 C h A 2 5 4 n m
1 0 /3 /2 0 1 9 1 :2 8 :4 6 P M IS TD a te A c q u ir e d :
D a te Pr o c e s s e d : 1 0 /1 9 /2 0 1 9 1 1 :4 9 :4 4 A M IS T
C h a n n e l: W 2 4 8 9 C h A ; P r o c e s s e d C h a n n e l: W 2 4 8 9 C h A 2 5 4 n m ; R e s u lt Id : 3 4 8 2 ; P r o c e s s in g M e th o d :
A c e B e n z T _ A B _ A C N 6 0
AU
0 . 0 0
0 . 1 0
0 . 2 0
0 . 3 0
M in u t e s
0 . 0 0 1 . 0 0 2 . 0 0 3 . 0 0 4 . 0 0 5 . 0 0 6 . 0 0 7 . 0 0 8 . 0 0 9 . 0 0 1 0 . 0 0
1
2
3
P r o c e s s e d
C h a n n e l D e s c r .
P e a k N a m e R T
W 2 4 8 9 C h A 2 5 4 n m
W 2 4 8 9 C h A 2 5 4 n m
W 2 4 8 9 C h A 2 5 4 n m
A c e to n e
B e n z e n e
T o lu n e
1 .6 0 0
3 .2 4 3
4 .3 2 7
P r o c e s s e d C h a n n e l D e s c r .:
W 2 4 8 9 C h A 2 5 4 n m
In je c tio n S u m m a r y R e p o r t
P r o je c t N a m e : C IP TR e p o r te d b y U s e r : S y s te m
R e p o r t M e th o d : In je c tio n S u m m a r y R e p o r t D a te Pr in te d :
3 4 8 9 1 0 /1 9 /2 0 1 9R e p o r t M e th o d ID : 3 4 8 9
1 1 :5 7 :1 7 A M A s ia /C a lc u ttaP a g e : 1 o f 1
S A M P L E I N F O R M A T I O N
S y s te mA c q u ir e d B y :A B _ A c e B e n z T _ S e t 1 _ 0 3 1 0 1 9 _ A C NS a m p le N a m e :
S a m p le S e t N a m e :U n k n o w nS a m p le T y p e :
1V ia l: A c q . M e th o d S e t: B P h a r m _ S R _ A c e A c e n a p _ 0 3 1 0 1 9
1 0In je c tio n # : P r o c e s s in g M e th o d : A c e B e n z T _ A B _ A C N 6 0
5 .0 0 u lIn je c tio n V o lu m e : C h a n n e l N a m e : W 2 4 8 9 C h A
R u n T im e : 6 .0 M in u te s P r o c . C h n l. D e s c r .: W 2 4 8 9 C h A 2 5 4 n m
1 0 /3 /2 0 1 9 2 :0 5 :4 0 P M IS TD a te A c q u ir e d :
D a te Pr o c e s s e d : 1 0 /1 9 /2 0 1 9 1 1 :4 9 :4 4 A M IS T
C h a n n e l: W 2 4 8 9 C h A ; P r o c e s s e d C h a n n e l: W 2 4 8 9 C h A 2 5 4 n m ; R e s u lt Id : 3 4 8 5 ; P r o c e s s in g M e th o d :
A c e B e n z T _ A B _ A C N 6 0
Acetone-1.591
Benzene-3.238
Tolune-4.328
AU
0 . 0 0
0 . 1 0
0 . 2 0
0 . 3 0
M in u t e s
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 4 . 5 0 5 . 0 0 5 . 5 0 6 . 0 0
1
2
3
P r o c e s s e d
C h a n n e l D e s c r .
P e a k N a m e R T A r e a % A r e a H e ig h t
W 2 4 8 9 C h A 2 5 4 n m
W 2 4 8 9 C h A 2 5 4 n m
W 2 4 8 9 C h A 2 5 4 n m
A c e to n e
B e n z e n e
T o lu n e
1 .5 9 1
3 .2 3 8
4 .3 2 8
2 4 7 8 9 9
1 9 4 3 9 6 3
2 3 9 3 0 2 8
5 .4 1
4 2 .4 0
5 2 .1 9
3 4 8 0 4
3 0 1 2 6 6
3 1 8 6 7 7
P r o c e s s e d C h a n n e l D e s c r .: W 2 4 8 9 C h A 2 5 4 n m

16. MOBILE PHASE: Water : acetonitrile :: 40 : 60 (v/v)
Run : 2 Run : 3
In je c tio n S u m m a r y R e p o r t
P r o je c t N a m e : C IP TR e p o r te d b y U s e r : S y s te m
R e p o r t M e th o d : In je c tio n S u m m a r y R e p o r t D a te Pr in te d :
3 4 8 9 1 0 /1 9 /2 0 1 9R e p o r t M e th o d ID : 3 4 8 9
1 1 :5 7 :4 7 A M A s ia /C a lc u ttaP a g e : 1 o f 1
S A M P L E I N F O R M A T I O N
S y s te mA c q u ir e d B y :A B _ A c e B e n z T _ S e t 2 _ 0 3 1 0 1 9 _ A C NS a m p le N a m e :
S a m p le S e t N a m e :U n k n o w nS a m p le T y p e :
1V ia l: A c q . M e th o d S e t: B P h a r m _ S R _ A c e A c e n a p _ 0 3 1 0 1 9
1 1In je c tio n # : P r o c e s s in g M e th o d : A c e B e n z T _ A B _ A C N 6 0
5 .0 0 u lIn je c tio n V o lu m e : C h a n n e l N a m e : W 2 4 8 9 C h A
R u n T im e : 6 .0 M in u te s P r o c . C h n l. D e s c r .: W 2 4 8 9 C h A 2 5 4 n m
1 0 /3 /2 0 1 9 2 :1 3 :4 0 P M IS TD a te A c q u ir e d :
D a te Pr o c e s s e d : 1 0 /1 9 /2 0 1 9 1 1 :4 9 :4 4 A M IS T
C h a n n e l: W 2 4 8 9 C h A ; P r o c e s s e d C h a n n e l: W 2 4 8 9 C h A 2 5 4 n m ; R e s u lt Id : 3 4 8 6 ; P r o c e s s in g M e th o d :
A c e B e n z T _ A B _ A C N 6 0
Acetone-1.596
Benzene-3.237
Tolune-4.327
AU
0 . 0 0
0 . 1 0
0 . 2 0
0 . 3 0
M in u t e s
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 4 . 5 0 5 . 0 0 5 . 5 0 6 . 0 0
1
2
3
P r o c e s s e d
C h a n n e l D e s c r .
P e a k N a m e R T A r e a % A r e a H e ig h t
W 2 4 8 9 C h A 2 5 4 n m
W 2 4 8 9 C h A 2 5 4 n m
W 2 4 8 9 C h A 2 5 4 n m
A c e to n e
B e n z e n e
T o lu n e
1 .5 9 6
3 .2 3 7
4 .3 2 7
2 4 7 9 8 0
1 9 5 2 3 0 9
2 3 8 9 9 5 6
5 .4 0
4 2 .5 3
5 2 .0 7
3 5 3 4 1
3 0 2 7 3 8
3 1 8 7 1 3
P r o c e s s e d C h a n n e l D e s c r .: W 2 4 8 9 C h A 2 5 4 n m
In je c tio n S u m m a r y R e p o r t
P r o je c t N a m e : C IP TR e p o r te d b y U s e r : S y s te m
R e p o r t M e th o d : In je c tio n S u m m a r y R e p o r t D a te Pr in te d :
3 4 8 9 1 0 /1 9 /2 0 1 9R e p o r t M e th o d ID : 3 4 8 9
1 1 :5 7 :5 9 A M A s ia /C a lc u ttaP a g e : 1 o f 1
S A M P L E I N F O R M A T I O N
S y s te mA c q u ir e d B y :A B _ A c e B e n z T _ S e t 3 _ 0 3 1 0 1 9 _ A C NS a m p le N a m e :
S a m p le S e t N a m e :U n k n o w nS a m p le T y p e :
1V ia l: A c q . M e th o d S e t: B P h a r m _ S R _ A c e A c e n a p _ 0 3 1 0 1 9
1 2In je c tio n # : P r o c e s s in g M e th o d : A c e B e n z T _ A B _ A C N 6 0
5 .0 0 u lIn je c tio n V o lu m e : C h a n n e l N a m e : W 2 4 8 9 C h A
R u n T im e : 6 .0 M in u te s P r o c . C h n l. D e s c r .: W 2 4 8 9 C h A 2 5 4 n m
1 0 /3 /2 0 1 9 2 :2 2 :5 5 P M IS TD a te A c q u ir e d :
D a te Pr o c e s s e d : 1 0 /1 9 /2 0 1 9 1 1 :4 9 :4 4 A M IS T
C h a n n e l: W 2 4 8 9 C h A ; P r o c e s s e d C h a n n e l: W 2 4 8 9 C h A 2 5 4 n m ; R e s u lt Id : 3 4 8 7 ; P r o c e s s in g M e th o d :
A c e B e n z T _ A B _ A C N 6 0
Acetone-1.596
Benzene-3.239
Tolune-4.329
AU
0 . 0 0
0 . 1 0
0 . 2 0
0 . 3 0
M in u t e s
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 4 . 5 0 5 . 0 0 5 . 5 0 6 . 0 0
1
2
3
P r o c e s s e d
C h a n n e l D e s c r .
P e a k N a m e R T A r e a % A r e a H e ig h t
W 2 4 8 9 C h A 2 5 4 n m
W 2 4 8 9 C h A 2 5 4 n m
W 2 4 8 9 C h A 2 5 4 n m
A c e to n e
B e n z e n e
T o lu n e
1 .5 9 6
3 .2 3 9
4 .3 2 9
2 4 4 1 3 7
1 9 0 6 2 6 3
2 3 3 8 5 6 6
5 .4 4
4 2 .4 7
5 2 .1 0
3 4 2 8 5
2 9 5 7 3 0
3 1 2 7 7 0
P r o c e s s e d C h a n n e l D e s c r .: W 2 4 8 9 C h A 2 5 4 n m

17. • As per polarity table given below the polarity of the used compounds are
as Acetone > Benzene > Toluene.
• Naturally in Reverse phase chromatography, acetone will be eluted first
followed by benzene and toluene, which in evident in the
chromatograms.
• Upon decreasing the polarity of the mobile phase (adding extra
acetonitrile to the mobile phase) the runtime of the experiment was
reduced.
• We assessed that even after reducing the polarity the separation was good
but decreased by quite an amount, but it hardly affected the USP tailing.
DISCUSSION
Solvent Relative polarity Eluent strength
Toluene 0.099 0.24
Benzene 0.111 0.32
Acetone 0.355 0.56

18. • Separation of the compounds (acetone, benzene, toluene) in a
mixture was not hampered by change in composition of mobile
phase.
• The peak shape and orientation were not hampered upon change
of mobile phase.
• The resolution between the chromatogram was also very good.
• It can be concluded that the mixture of acetone, benzene &
toluene, could be successfully employed as system suitability
check solution.
CONCLUSION

19. • IP 2007, VOLUME I, Section: 2.4.14
• USP NF 2002; 621(Chromatography): 1990-1991
REFERENCES