PMID- 20552175
OWN - NLM
STAT- MEDLINE
DCOM- 20110414
LR  - 20191210
IS  - 1618-2650 (Electronic)
IS  - 1618-2642 (Linking)
VI  - 399
IP  - 2
DP  - 2011 Jan
TI  - Simple fluorescence assay for quantification of OSCS in heparin.
PG  - 673-80
LID - 10.1007/s00216-010-3867-5 [doi]
AB  - In 2008, heparin contaminated with oversulfated chondroitin sulfate (OSCS) 
      penetrated the worldwide market and was associated with severe adverse effects. 
      Feasible and reliable methods to test heparin for adulteration are needed. The 
      objective was to develop a simple approach based on a microplate assay for 
      quantification of heparin and sulfated glycans using the fluorescent heparin 
      sensor polymer-H (polymer-H assay). However, both heparin and OSCS 
      concentration-dependently increase the fluorescence intensity (FI) of polymer-H, 
      so that OSCS in heparin cannot be detected. The idea was a two-step procedure 
      including, first, separation of heparin by degradation with heparinase I, and 
      then measurement of the remaining OSCS. To achieve complete heparin 
      (unfractionated heparin (UFH), enoxaparin) degradation, several conditions (e.g. 
      incubation time and heparinase I concentration) were optimized by using the aXa 
      assay for monitoring. Defined UFH/OSCS mixtures incubated in this way showed a 
      concentration-dependent FI increase in the polymer-H assay (lambda ((em)) 330 nm, lambda 
      ((ex)) 510 nm). The sensitivity was unexpectedly high with an LOD/LOQ of 
      0.5%/0.6% OSCS content in heparin. Further experiments testing UFH/OSCS mixtures 
      in the aXa assay confirmed our hypothesis: OSCS inhibits heparinase I resulting 
      in incomplete heparin degradation and thus an additional FI increase of polymer-H 
      by intact heparin. This two-step microplate fluorescence assay is a sensitive, 
      rapid, and simple method for quantification of OSCS in heparin. In contrast with 
      (1)H NMR and CE, neither expensive equipment nor much experience are required. It 
      could be applied not only in the quality control of heparin, but also in clinical 
      practice, to check the applied heparin preparation when a patient suffers any 
      adverse effect.
FAU - Luhn, Susanne
AU  - Luhn S
AD  - Pharmaceutical Institute, Christian-Albrechts-University, Gutenbergstrasse 76, 
      24118, Kiel, Germany, sluehn@pharmazie.uni-kiel.de
FAU - Schiemann, Simone
AU  - Schiemann S
FAU - Alban, Susanne
AU  - Alban S
LA  - eng
PT  - Evaluation Study
PT  - Journal Article
DEP - 20100616
PL  - Germany
TA  - Anal Bioanal Chem
JT  - Analytical and bioanalytical chemistry
JID - 101134327
RN  - 0 (Anticoagulants)
RN  - 0 (Heparin, Low-Molecular-Weight)
RN  - 9005-49-6 (Heparin)
RN  - 9007-28-7 (Chondroitin Sulfates)
RN  - EC 4.2.2.7 (Heparin Lyase)
SB  - IM
EIN - Anal Bioanal Chem. 2011 Jan;399(3):1395
MH  - Animals
MH  - Anticoagulants/*chemistry
MH  - Chondroitin Sulfates/*analysis
MH  - *Drug Contamination
MH  - Heparin/*chemistry/metabolism
MH  - Heparin Lyase/metabolism
MH  - Heparin, Low-Molecular-Weight/chemistry/metabolism
MH  - Sensitivity and Specificity
MH  - Spectrometry, Fluorescence/economics/*methods
MH  - Swine
EDAT- 2010/06/17 06:00
MHDA- 2011/04/16 06:00
CRDT- 2010/06/17 06:00
PHST- 2010/03/11 00:00 [received]
PHST- 2010/05/20 00:00 [accepted]
PHST- 2010/05/12 00:00 [revised]
PHST- 2010/06/17 06:00 [entrez]
PHST- 2010/06/17 06:00 [pubmed]
PHST- 2011/04/16 06:00 [medline]
AID - 10.1007/s00216-010-3867-5 [doi]
PST - ppublish
SO  - Anal Bioanal Chem. 2011 Jan;399(2):673-80. doi: 10.1007/s00216-010-3867-5. Epub 
      2010 Jun 16.