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Update to USP <621> Chromatography: What does this mean?
USP 37 NF 32 1S – Current as of August
2014
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What is the USP-NF?
The United States Pharmacopeia – National Formulary (USP-NF) is a book of pharmacopeial standards – Drugs substances & preparations monographs: USP – Dietary supplements & ingredients monographs: USP – Excipient monographs: NF – More than 4500 monographs
The USP-NF is the official authority – FDA-enforceable standards – Enforcement of USP standards is the responsibility of FDA and other
government authorities in the U.S. and elsewhere – USP has no role in enforcement
The U.S. Federal Food, Drug, and Cosmetics Act designates the
USP–NF as the official compendia for drugs marketed in the United States
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Why is Chromatography <621> Important?
Question From CURRENT USP-NF Online FAQs1 – Q. How much can I modify a chromatographic procedure and still be
in compliance? Can column length, internal diameter, mobile phase composition be modified?
– A. Chromatography <621> contains a list of allowed adjustments to chromatographic systems. However, the user should verify the suitability of the method under the new conditions by assessing the relevant analytical performance characteristics potentially affected by the change (see section System Suitability under Chromatography <621>).
1http://www.uspnf.com/uspnf/scienceFAQ.html#q4
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USP <621> Chromatography Defines “Allowable adjustments”
Adjustments to a USP method may be made to meet system suitability requirements
Verification tests must be performed after changes – Full re-validation not required
Must use the same L-designation of column Isocratic hold or dwell volume adjustments are allowed
USP 37 NF 32 S1 - to be official Aug. 1, 2014 - significant changes to Chapter <621> Chromatography
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Current Changes to <621>
Flow rate (isocratic): F2 = F1 x [(dc22 x dp1)/(dc1
2 x dp2)]
L/dp = column length (L) to particle size (dp) ratio N = theoretical plate count (for solid-core particles)
Previous (USP36-NF31)
Isocratic & Gradient Isocratic Gradient Particle Size -50%
Column Length ±70% Flow Rate ±50% *based on particle size and ±50% no changes allowed
Column ID Any allowed Any allowed no changes allowed
Injection Volume Any reduction Any allowed Any allowed
Column Temperature ±10°C ±10°C ±10°C
Mobile Phase pH ±0.2 unit ±0.2 unit ±0.2 unit
Variable Since Aug 1, 2014 (USP37-NF32 S1) Allowable Adjustments in USP <621> Chromatography
per constant L/dp or N: -25% to +50% no changes allowed
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USP and EP
Variable
USP <621> Guidelines as of Aug 1, 2014 (USP37-NF32 S1) Current EP <2.2.46> V8.2 Guidelines
Isocratic Gradient Isocratic Gradient
Particle Size Per constant L/dp or N: -25%
to +50% No changes allowed -50%, no increase No changes allowed
Column Length ±70% ±70%
Flow Rate *Based on particle size and ±50% No changes allowed ±50%, *More if changing column
dimensions *Flexible ONLY if changing column
dimensions
Column ID Flexible No changes allowed ±25% ±25%
Injection Volume Flexible Flexible Decrease only for proper detection and repeatability
Decrease only for proper detection and repeatability
Column Temperature ±10°C ±10°C ±10°C ±5°C
Mobile Phase pH ±0.2 unit ±0.2 unit ±0.2 unit No changes allowed
*F2=F1 x [(dc22 x dp1)/(dc1
2 x dp2)] *F2=F1 x [(L2 x d22)/(L1 x d1
2)]
L/dp = column length (L) to particle size (dp) ratio L= Column Length
N = theoretical plate count (for solid-core particles) d= Column ID
Refer to Chapter <621> of USP37-NF32 S1 for further information and details on adjustments not shown
Refer to Section <2.2.46> of EP version 8.2 for further information and details on adjustments not shown
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• Changes made within USP <621> allowable adjustments (USP37-NF32 S1, August 2014) • 90% savings in runtime and 94% solvent savings per run! • Re-validation not required
Benefits: Isocratic USP Method Levonorgestrel/Ethinyl Estradiol
0.00 1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 0.000
0.010
0.020
0.030 HPLC XBridge C
8
0.00 1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00
AU
0.000
0.010
0.020
0.030 HPLC USP Res: 6.9
0.00 0.20 0.40 0.60 0.80 1.00
0.000
0.010
0.030
0.050
Minutes 0.00 0.20 0.40 0.60 0.80 1.00
Minutes Minutes
AU
levo
norg
estre
l ACQUITY UPLC H-Class Column: ACQUITY BEH C8, 2.1 x 50mm, 1.7µ (L7) L/dp = 29,400 Flow Rate: 0.61 mL/min, pressure 7700 psi Mobile Phase: 35:15:45 ACN:MeOH:Water Injection Volume: 3.5 µL
Alliance 2695 HPLC Column: XBridge C8, 4.6 x 150mm, 5µ (L7) L/dp = 30,000 Flow Rate: 1.0 mL/min , pressure 1400 psi Mobile Phase: 35:15:45 ACN:MeOH:Water Injection Volume: 50 µL
UPLC USP Res: 7.0
levo
norg
estre
l
ethi
nyl e
stra
diol
et
hiny
l est
radi
ol
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0.00 5.00 10.00 15.00 20.00 25.00 30.00 35.00 40.00 45.00 50.00 55.00
HPLC Separation on Alliance HPLC System
XBridgeTM C18 4.6 x 100 mm, 3.5 µm Tailing = 1.60 Rs = 7.6
Gal
anta
min
e
1 2
4 5
Benefits: USP Gradient Method Galantamine and Related compounds
AU
0.000
0.010
0.020
0.030
0.040
0.050
0.060
0.070
0.080
0.090
0.00 2.00 4.00 6.00 8.00 10.00 12.00 14.00
UPLC Separation on ACQUITY UPLC H-Class
ACQUITY UPLC BEH C18 2.1 x 50 mm, 1.7 µm Tailing = 1.43 Rs = 7.2
Gal
anta
min
e
1 2
4 5
HPLC to UPLC Transfer: Reduce analysis time by 77% (4.3X) while preserving separation integrity Tailing and Resolution Criteria were met Assay Criteria: USP Tailing < 2.0 Rs (galantamine/impurity 4) > 4.5 Re-validation will be required under USP37-NF32 S1 (effective Aug 2014).
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Empower Method Validation Manager for Verification/Validation
Improve compliance and reduce errors:
Automate testing within Empower
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Reference Experts from USP37-NF32 1S (Published)
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“Allowable adjustments”
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Changes to Particle Size
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Changes to Flow Rate
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Changes to Other factors
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Example provided