Quality Standards for Medicines, Supplements, and Food Ingredients throughout the World
USP Heparin Monograph Revisions — Stage 2 and Beyond Anita Szajek, Ph.D. Fabian Jameison, Ph.D.
3rd Workshop on the Characterization of Heparin Products July 27-28, 2009 USP Headquarters
Outline
• Background – USP Heparin monograph family – USP Heparin monographs: Revision timeline – USP monograph revision process: Heparin case study
• Brief recap of Stage 1 revisions • Stage 2 revisions – – – –
Summary of revisions Path to official: Implementation of stage 2 revisions Implementation of new potency RS Impact of new potency RS on other heparin monographs
• Future heparin revisions 2
USP Heparin Monographs Heparin Sodium Heparin Calcium
Heparin Lock Flush Solution Heparin Calcium Injection Heparin Sodium Injection Anticoagulant Heparin Solution Antithrombin III Human Protamine Sulfate Protamine Sulfate Injection Protamine Sulfate for Injection
LMWHs:* Enoxaparin Sodium Enoxaparin Sodium Injection Test for 1,6-Anhydro Derivative for Enoxaparin Sodium *Official on December 1, 2009
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USP Heparin Monographs: Revision Timeline
2006 Heparin Panel formed
June 18, 2008 Heparin Revision Bulletins (Stage 1) April 2008 Heparin AP F2F mtg
Aug 2008 Heparin AP F2F mtg
Dec 2008 Heparin AP F2F mtg
RS
March 2007 aIIa Assay proposed
Apr 11, Jun 12, 2008 2008 Webcast Webcast
2007-2008 Heparin crisis
Oct 1, 2009 Future Heparin IRAs + other Heparin RB revisions Official (Stage 2) RS
Public Comment
March 2009 Heparin IRAs proposed (Stage 2);
May 1, 2009 Webcast
Protamine monograph revisions
Mar 3, 2009 Webcast
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USP Monograph Revision Process: Heparin Case Study Stakeholders
USP USP Heparin Panel
FDA
USP BBP EC ballots
Web cast Web cast Public comment period 5
Heparin Monograph Updates—Process •
Stage 1 – Became official via Revision Bulletin, June 18, 2008Ref 1 No official public comment period, but USP engaged industry via web meetings – Length of time from start of revisions to official status: Three months
•
Stage 2 – Proposed IRA in March-April 2009 PF Ref 2 – Comment deadline May 15, 2009 – Initially targeted to become official in July-August PF (official August 1, 2009) – Scheduled to become official in the September-October PF (official date October 1, 2009) due to the large number of comments and to assure availability of Reference Standards
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Stage 2 Revision Timeline • • • • • • • • • • •
May 27th 2008: USP Heparin Ad hoc Advisory Panel establishes sub-teams to deliberate purity, potency, and identity tests June 19th & 20th 2008 : EDQM-NIBSC-USP Heparin Workshop August 7th & 8th 2008 : USP heparin Ad hoc Advisory Panel meets October: Heparin Ad hoc Advisory Panel updates USP Blood & Blood Products Expert Committee December 16th & 17th : Heparin Ad hoc Advisory Panel to finalize monograph February 2009: Publication of USP Immediate Revision Announcement (IRA) March 3rd 2009: USP Web Meeting #1 May 1st 2009: USP Web Meeting #2 March 15th - May 15th 2009: Public comment period July 7th – July 16th 2009: BBP EC balloted to adopt the Heparin AP recommendation. October 2009 (originally August 2009): Revised monograph to become official through IRA in PF 35(5), per panel recommendation and EC approval, new reference standards to be available
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Revisions to Heparin Sodium Monograph Overview Procedure
Stage 1 Revisions
Stage 2 Revisions
Identification
1H
NMR CE
-Expanded 1H NMR procedure -Replaced CE method with anion-exchange HPLC procedure -Added ratio of anti-factor Xa activity to anti-factor IIa potency
Potency
No revision
-Replaced sheep plasma clotting assay with chromogenic anti-factor IIa assay
Organic Impurities
No revision
-Added limit of total galactosamine in total hexosamine (a measure of dermatan sulfate and other galactosamines) -Revised protein impurities -Added nucleotidic impurities -Added residual solvents
Absence of OSCS
See Identification
-References Identification A and B
New USP Reference Standards (RS)*
-USP Heparin Sodium Identification RS -USP Heparin Sodium System Suitability RS
-USP Oversulfated Chondroitin Sulfate RS -USP Dermatan Sulfate RS -USP Galactosamine Hydrochloride RS -USP Glucosamine Hydrochloride RS -USP Heparin Sodium for Assays RS
*See USP Heparin RS flyer
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IDENTIFICATION A. 1H NMR Spectrum Rationale for retaining and expanding the 1H NMR method •
The method is sensitive in terms of LOD. – Expanded spectral window: at least 10 to -2 ppm – Increased S/N ratio requirement to at least 1000/1
•
The complexity of spectra is a drawback in the context of purity testing – A simplified method of data analysis is proposed – The method is supported by USP RS and reference spectra
•
Using the full spectrum heparin is easily distinguished from dermatan, chondroitin, and OSCS.
•
Any spectral variation alerts users in the event of contamination of unknown origin
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1H
8.00
3
ppm
Methyl of GlcNAc
ÍTSP
ÍHOD
1 2
H2 of GlcNS
4 H1 of IdoA2S
H1 of GlcNAc/GlcNS, 6S
NMR Specification for Identity and Purity of Heparin: Example
0 10
IDENTIFICATION B: Strong Anion Exchange Chromatography Method •
This method is able to detect glycosaminoglycans (GAGs) contaminants – Among the LC methods evaluated, this method provided the best separation between dermatan and heparin. – Heparin, dermatan, and OSCS are completely resolved in the proposed System suitability solution. – Dermatan sulfate and chondroitin sulfate co-eluted.
0.090 0.080 0.070
0.050 0.040 0.030 0.020
OSCS - 49.886
Dermatan - 20.603
0.060 AU
Resolution solution: Heparin [20 mg/mL], dermatan and OS-CS each at 1% (w/w)
Heparin - 32.726
0.100
0.010 0.000 0.00
5.00
10.00
15.00
20.00
25.00
30.00 35.00 Minutes
40.00
45.00
50.00
55.00
60.00
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ORGANIC IMPURITIES: Limit of Galactosamine in Total Hexosamine A measure of dermatan sulfate and galactosamine containing impurities • Oversulfated chondroitins, Dermatan, and Chondroitin Sulfate A all have the same retention time, through digestion converted to galactosamine. 30.0 nC 23D
ED_1
Glucosamine
25.0
20.0
Suitability requirements: Theoretical plates >2000 Tailing factor 0.8
