Download - Practical issues in UV, HPLC analysis, formulation development of Solid Lipid Nanoparticles
Practical issues in UV, HPLC analysis, and formulation development of Solid Lipid Nanoparticles
Vijay Kumar E M.Pharm. Pharmaceutics
Email ID: [email protected]
Presentation outline
β’ Issue 01: Change in Emax and Valley depth
β’ Issue 02: RP-HPLC chromatograms peak splitting
β’ Issue 03: Batch to Batch Variability during replication
0
Peak 01
Peak 02
Peak 03
Valley 01Valley 02
200nm 400nm
Abso
rban
ce
(AU
)
Wavelength
Day 03Day 02
Day 01
π΄l=ππππππ π ππ .πππ .lβππ’ππ π πππ£πππ‘ πππ .l
Issue 01
1mg/mL
1.2mg/mL
1.4mg/mL
Day 01
Why do peaks attain a new high Emax as the time progresses?
π΄=πππ
b
Analyte
Day 02
Day 03
Assume analyte has no chromophoric group that absorbs at 300nm and only responsible agent for absorbance is pure solvent.
Organic
ππππππ π ππ.πππ . l 300=1.47
π΄l 300=1 .47β1 .50
Day 02π΄l 300=1 .50β1 .50
ππππππ π ππ.πππ . l 300=1.50
Day 01
ππππππ π ππ.πππ . l 300=1.45
π΄l 300=1 .45β1 .50
Day 03
Why do valley gets even more deep as the time progresses?
Consider the following case π΄l=ππππππ π ππ .πππ .lβππ’ππ π πππ£πππ‘ πππ .l
0
-0.05
300nm
Strategy(s) sought to address the issue
β’ Store at constant temperature (at 25C) (Temperature β Solubility)
β’ Adequately tighten the lid of the solution holder
AU
-0.002
0.000
0.002
Minutes
0.00 2.00 4.00 6.00 8.00 10.00 12.00 14.00
AU
0.000
0.005
0.010
Minutes
0.00 2.00 4.00 6.00 8.00 10.00 12.00 14.00
A
B
A-Chromatogram with split peaks; B-Chromatogram with no split peaks
Strategies sought to address the issue
β’ Mobile phase ratio manipulationβ’ Solvent effectβ’ Guard column replacement
Negative
Negative
Positive
One factor variation at a time
Representation of elution in normal ODS column (guard column) and simultaneous AU-Time graph
Transverse plane view
Longitudinal plane view
P1
P2
P1=P2
Representation of elution in normal ODS column and simultaneous AU-Time graph
Time
Conc
entr
ation
(mcg
/mL)
/ Ab
sorb
ance
uni
ts P1
P2
P1=P2
Representation of elution in normal ODS column and simultaneous AU-Time graph
Time
Conc
entr
ation
(mcg
/mL)
/ Ab
sorb
ance
uni
ts P1
P2
P1=P2
Representation of elution in normal ODS column and simultaneous AU-Time graph
Time
Conc
entr
ation
(mcg
/mL)
/ Ab
sorb
ance
uni
ts P1
P2
P1=P2
Representation of elution in normal ODS column and simultaneous AU-Time graph
Time
Conc
entr
ation
(mcg
/mL)
/ Ab
sorb
ance
uni
ts P1
P2
P1=P2
Peak splitting in RP-HPLC chromatograms due to guard column malfunction
Time
Conc
entr
ation
(mcg
/mL)
/ Ab
sorb
ance
uni
ts
P1
P2
P1>P2
Peak splitting in RP-HPLC chromatograms due to guard column malfunction
Time
Conc
entr
ation
(mcg
/mL)
/ Ab
sorb
ance
uni
ts
P1
P2
P1>P2
Peak splitting in RP-HPLC chromatograms due to guard column malfunction
Time
Conc
entr
ation
(mcg
/mL)
/ Ab
sorb
ance
uni
ts P1
P2
P1>>P2
Peak splitting in RP-HPLC chromatograms due to guard column malfunction
Time
Conc
entr
ation
(mcg
/mL)
/ Ab
sorb
ance
uni
ts
P1
P2
Response Replicate I Replicate II Replicate III
Particle size 291.6 513.9 290.9
PI 0.403 0.359 0.364
Replicate II
287.9
0.412
β’ Two unequal volumes of water cannot have same temperature when heated for the same time
25mL250mL
Time of heating process = 30min
A B
After heating for 30min; Temp. of A>>Temp. of B
Strategy(s) sought to address the issue
β’ Minimize the background noise (like Constant temperature maintenance, ultra-sonication duration, depth of homogenizer probe immersion )