global heparin supply

2 Pharmaceutical Technology November 2015 PharmTech .com

Special Report: Global Supply Chain

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Heparin regulates hemostasis at various points of the coagulation cascade mainly through its inter-

action with antithrombin and heparin cofactor II. Because of these properties, heparin is a life-saving anticoagulant drug used in renal dialysis, cardiac surgery, and treatment for deep vein thrombosis. The drug also binds to platelets, inhibiting platelet function and contributing to the hemorrhagic effects of heparin. Bovine heparin, first approved in 1939, was widely used in the United States for more than 50 years (see Figure 1). Like all drugs, heparin can cause adverse effects, but overall, bovine heparin products were found to be safe and effective during that period.

In the late 1980s, bovine spongi-form encephalopathy (BSE, or “mad cow disease”) was reported first in the United Kingdom and later in several other countries, raising concerns about the use of bovine-sourced heparin products in humans. Because of these concerns, manufacturers of bovine heparin products voluntarily withdrew them from the US market in the 1990s. Since then, heparin products approved for use in the US and Europe have been sourced solely from pigs, with

approximately 60% of the supply of the drug coming from China. Figure 2 illustrates steps involved in manufac-turing heparin from porcine intestinal mucosa and potential impurities that are inactivated and/or removed from each manufacturing step.

Heparin is a lifesaving drug that was safely used since the 1940s. However, in 2007, contaminated heparin caused a number of deaths in the US and hun-dreds of adverse reactions worldwide (1). The contaminated heparin was found to contain over-sulfated chon-droitin sulfate (OSCS). OSCS was an inexpensive synthetic adulterant that had some anticoagulant activity and was presumably added to heparin to increase profit when the drug was in short supply due to a pig disease outbreak. This “heparin crisis” dem-onstrated the vulnerability of drug supplies produced from increasingly global manufacturing chains and high-lighted the risks inherent in reliance on one country and one animal species as the primary source for a crucial drug.

To mitigate these concerns by diversi-fying the sources of heparin drugs, FDA is considering reintroduction of bovine heparin drug product to the US market. In August 2015, the US Pharmacopeial Convention (USP) hosted the 6th Work-shop on the Characterization of Heparin Products in São Paulo, Brazil, with co-sponsors, the National Institute for Bio-logical Standards and Control (NIBSC, UK), National Health Surveillance Agency (ANVISA, Brazil), and Sao Paulo State Pharmaceutical Manufacturers As-sociation (SINDUSFARMA, Brazil). The focus of the workshop was an examina-tion of the global heparin supply chain, specifically the risks of heparin shortages, adulteration, and contamination.

The following is an overview of sci-entific research and clinical experience presented at the workshop to generate improved understanding of the differ-ences between porcine and bovine hepa-rins, the clinical implications of reintro-ducing bovine heparin in the US, and the broader ramifications of bovine heparin in the US market and worldwide.

David Keire is acting laboratory chief, Branch I,

Division of Pharmaceutical Analysis, FDA;

Barbara Mulloy is visiting professor at Institute of

Pharmaceutical Sciences, King’s College London;

Christina Chase is senior scientific writer at US

Pharmacopeial Convention (USP);

Ali Al-Hakim is acting director of Division of New

Drug API at the FDA; Damian Cairatti is head of

Latin American Regulatory Affairs at USP;

Elaine Gray is principal scientist at National Institute

of Biological Standards and Control (NIBSC) in the UK;

John Hogwood is research scientist at NIBSC;

Tina Morris is senior VP of Science Global

Biologics at USP;

Paulo Mourão is professor at Federal University

of Rio de Janeiro;

Monica Da Luz Carvalho Soares is a member of the

Deliberative Council of the Brazilian Pharmacopeia

at ANVISA and a visiting fellow at the University of

Maryland Baltimore County (UMBC); and

Anita Szajek is principal scientific liaison at USP.

The global supply chain for bovine and porcine heparin and regulatory considerations are examined.

Diversifying the Global Heparin Supply Chain: Reintroduction of Bovine Heparin in the United States?David Keire, Barbara Mulloy, Christina Chase, Ali Al-Hakim, Damian Cairatti, Elaine Gray, John Hogwood, Tina Morris, Paulo A.S. Mourão, Monica da Luz Carvalho Soares, and Anita Szajek


Porcine vs. bovine heparinsHeparin is a natural product, extracted from animals. Just as pork and beef are different from each other, heparin products made from pigs and cattle are similar but not identical. Two ses-sions of the USP workshop focused on laboratory tests used to understand the differences in structure and biological

activity between bovine and porcine heparin.

The structure of heparin is that of a linear polysaccharide consisting of repeating disaccharide motifs in which uronic acids alternate with glu-cosamine. The polysaccharide chains can vary in length and in substitu-tion with sulfates and N-acetyl groups.

Structural analysis techniques range from relatively simple, straightforward spectroscopic and chromatographic analyses to sophisticated applications of techniques in nuclear magnetic res-onance and mass spectrometry.

The biological activity of heparin, including its abilities to inhibit the en-zymes of blood clot formation in vivo and in vitro, can be quantified by sev-eral methods. Research suggests that molecular weight and disaccharide composition both play an important role in biological activity. High mo-lecular weight fractions of heparin, for example, have a greater effect than do lower molecular weight fractions on anticoagulation potency.

Overall, the combination of struc-tural and functional information avail-able provides a clear picture of heparin products from different species. Nu-merous samples have been tested by heparin manufacturers and academic and regulatory labs revealing clear and consistent differences in the structures (see Figure 3) and biological activity profiles of porcine mucosal and bovine mucosal heparin. In addition, the few samples of bovine lung heparin tested were quite distinct from either mucosal sample type.

Importantly, the data presented show that bovine heparin was signifi-


Figure 1: Historical development timeline of therapeutic heparin in United States.

Figure 2: Heparin manufacturing process.



cantly less potent, weight for weight, than porcine heparin. The relationship between laboratory testing and clini-cal experience is not straightforward for such complex products, yet a differ-ence in potency could have important clinical relevance (2). Therefore, fur-ther investigation including clinical research may be warranted.

Bovine heparin and safety There are two main safety concerns as-sociated with bovine heparin. The first is heparin-induced thrombocytopenia (HIT), an infrequent but potentially devastating adverse event (3,4). HIT is an immune response in which the body makes antibodies to large com-plexes formed between the highly sul-fated heparin chains and platelet factor 4 (5). HIT occurs in 0.2–5% of patients regardless of the type of heparin ad-ministered; porcine and bovine hepa-rin appear similar in terms of HIT risk (6,7). Another common adverse event associated with heparin is bleeding, which can be controlled through the

neutralization of heparin by protamine sulfate (8).

The second safety concern, specific for bovine heparin, is the possible pres-ence of BSE infectious agents (9). Dur-ing the BSE epidemic in the UK, some people consumed BSE-infected beef. From 1999–2000, after a long incuba-tion period, some of these individuals developed variant Creutzfeldt-Jakob disease (vCJD); 229 cases have oc-curred worldwide as of May 28, 2015 (10). Since its peak in 2000, vCJD has declined significantly but has not been eradicated, as a few cases are still de-tected every year. No known cases of vCJD, however, have been linked to use of bovine heparin. In addition, in India, Brazil, and Argentina—where bovine heparin products have been in continuous use—no cows have tested positive for BSE and no cases of vCJD have been observed. In the US, only three atypical BSE cases (i.e., different from the distinct BSE strain from the UK that causes vCJD) in cattle have been identified (11). Since the 1990s,

much has been learned about how BSE leads to vCJD in humans. In addition, methods for minimizing BSE risk in bovine materials have advanced (12, 13). Generally, risks from tissue spon-giform encephalopathy (TSE) agents are controlled in three steps: animal origin of species and supply chain control, tissue harvest controls, and chemical treatments that remove in-fectious agents (11). If bovine heparin is reintroduced, these steps will be in-strumental for ensuring patient safety. Because of the safety concerns noted previously, bovine heparin is likely to have its own USP monograph and a separate label (Physician Labelling Rule) that differentiates bovine hepa-rin from porcine heparin.

Possible reintroduction of bovine heparin into the US marketThe only approved source of heparin in most of the world is pig intestine, but the global pig supply is limited geo-graphically. In addition, there is little growth potential for porcine heparin products to be manufactured in other parts of the world. Thus, FDA is con-cerned about potential shortages due to pig disease or possible geo-political instability.

After considering the available op-tions, FDA hosted a meeting with its Science Board in June 2014 to discuss the possible reintroduction of bovine heparin in the US. Reintroduction of bovine heparin would no longer limit the source to one animal species and would extend the geographic distri-bution of source animals. If disease occurs in one animal source and/or there is geo-political instability in a major source country, the risk of sup-ply shortages could be more readily mitigated.

The original US-approved hepa-rin drugs from the 1930s were from a bovine source (cow lung) and upon approval of porcine heparin products, both were used interchangeably until the 1990s without major safety risks or concerns. Notably, bovine mucosa heparin drug product is currently available and manufactured in South

Figure 3: Part of the proton NMR spectra of (lower, red spectrum) porcine mucosal and (upper, blue spectrum) bovine mucosal heparin. Though the two spectra are similar they are not exactly the same; some differences are indicated by arrows. The NMR spectra reflect the chemical structures of porcine and bovine heparin, showing that heparin samples from the two sources are similar but not identical.

Bovine intestinePorcine intestine

PPM 5.70 5.60 5.50 5.40 5.30 5.20 5.10 5.00 4.90 4.80 4.70

Pharmaceutical Technology November 2015 7

Table I: Pharmacopeial requirements for porcine and bovine heparin. RS is reference standard.Monograph section Test Porcine heparin Bovine heparin

Identification 1H NMR No unidentified signals greater than 4% of the mean of signal height of 1 and 2 are present in the following ranges: 0.10–2.00, 2.10–3.20, and 5.70–8.00 ppm. No signals greater than 200% signal height of the mean of the signal height of 1 and 2 are present in the 3.75–4.55 ppm for porcine heparin.

• New acceptance criteria need to be generated based on batch data.

Strong anion exchange high performance liquid chromatography (SAX-HPLC) as chromatographic identity

The retention time of the major peak from the Sample solution corresponds to that of the Standard solution.

• Chrom ID method needs to be qualified/validated for bovine heparin. Depending on resolution, a new method may be needed.

• New RS may be needed

Anti-factor Xa to Anti-factor IIa ratio

Acceptance criteria: 0.9–1.1 • New acceptance criteria may be needed.• New potency standard may be needed.• Potency method needs to be qualified/

validated for bovine heparin.

Molecular weight (MW) determinations

Acceptance criteria: m24000

is NMT 20%, mw is

between 15,000 Da and 19,000 Da, and the ratio of m

8000–16000 to m

16000–24000 is NLT 1.0.

• MW method needs to be qualified/validated for bovine heparin.

• New acceptance criteria may be needed

Sodium It meets the requirements of the flame test for sodium.

It meets the requirements of the flame test for sodium.

Species and tissue identification

Disaccharide analysis Add for species identification Add for species and tissue identification

Assay Anti-factor IIa Potency Not less than (NLT) 180 USP Heparin Units/mg • Needs to be determined using batch data• New RS may be needed• New acceptance criteria are needed.

Other components Nitrogen Determination, Method I <461>

1.3%–2.5% 1.3%–2.5%

Impurities Residue on Ignition <281> 28.0%–41.0% 28.0%–41.0%

Galactosamine in Total Hexosamine

Not more than (NMT) 1% NMT 1%

Nucleotidic Impurities and Protein Impurities

NMT 0.1% NMT 0.1%

Absence of OSCS References Identification Test A and B References Identification Test A and B

BSE/TSE N/A

Specific tests Bacterial Endotoxins Test <85>

NMT 0.03 USP Endotoxin Unit/USP Heparin Unit

NMT 0.03 USP Endotoxin Unit/USP Heparin Unit

Loss on Drying <731> Loss of NMT 5.0% of weight Loss of NMT 5.0% of weight

pH <791> 5.0–7.5 in a solution (1:100) 5.0–7.5 in a solution (1:100)

Sterility Tests <71> Where it is labeled as sterile, it meets the requirements

Where it is labeled as sterile, it meets the requirements

Additional requirements

Packaging and Storage Preserve in tight containers, and store below 40 ˚C, preferably at room temperature.

Preserve in tight containers, and store below 40 ˚C, preferably at room temperature.

Labeling Label to indicate the tissue and the animal species from which it is derived

Label to indicate the tissue and the animal species from which it is derived

USP Reference Standards <11>

USP Heparin Sodium Identification RSUSP Oversulfated Chondroitin Sulfate RSUSP Dermatan Sulfate RSUSP Galactosamine Hydrochloride RSUSP Glucosamine Hydrochloride RSUSP Heparin Sodium for AssaysUSP Heparin Sodium Molecular Weight Calibrant RSUSP Adenosine RS

New RS needed:USP Bovine Heparin Sodium Identification RS



America (particularly Brazil and Argen-tina) and India. In some countries, bo-vine heparin is preferred for religious rea-sons. Thus, there have been more than 50 years of safe and effective use of bovine lung heparin in patients in the US, and bovine mucosal heparin has been used safely in South America and India. How-ever, because the heparin characteriza-tion technology has advanced in the in-terim, the specifications for the proposed bovine heparin should be modernized in a manner similar to the recent update of the USP porcine heparin monograph.

Bovine heparin use worldwideHeparin derived from bovine lungs was in general use in Europe and the US until it gradually fell out of use for two reasons: the commercial advantages of the more potent porcine product, and concerns in the 1990s about transmission of BSE to humans through contaminated beef products.

Workshop speakers from Argentina, Brazil, and India emphasized that bovine heparin has been approved and used for decades in their countries. The Argen-tinian Pharmacopeia is discussing how to manage bovine and porcine heparins, and the Brazilian Pharmacopeia is cur-rently devising specific monographs for bovine and porcine mucosal heparin.

The availability of bovine and porcine heparins in Brazil has varied considerably in recent years. In 2008, 42% of heparin was of bovine origin, but this was fol-lowed by a decrease and then total re-moval from the market in 2013. In Argen-tina, the bovine source accounts for 70% of the total heparin; this has remained unchanged in recent years.

No serious adverse effects have been associated with the use of bovine heparin in these two countries or in India. Some excess adverse events associated with heparin in cardiovascular surgery, how-ever, were reported to ANVISA in 2008 when, on short notice, bovine heparin replaced porcine heparin for cardiovas-cular surgery in Brazil. The Brazilian So-ciety of Cardiovascular Surgery (14) and ANVISA (15) published warning notes suggesting careful monitoring of antico-agulant levels when using bovine heparin.

There are no reports of clinical trials comparing heparins from bovine versus porcine intestine. Therefore, although bo-vine heparin has been used successfully in several countries for many years, caution may be required when porcine heparin is replaced with bovine heparin without warning.

Bovine heparin use in the manufacture of LMWHsLow molecular weight heparins (LMWHs) are manufactured from heparin sodium (also known as unfractionated heparin sodium, or UFH) using chemical or en-zymatic depolymerization methods (16). Currently, in the US, all forms of LMWH are made from porcine UFH. Therefore, their composition and properties are known based on their biological starting material.

The structure and composition of bo-vine UFHs differ from that of porcine, therefore a LMWH product made from bovine UFH could have different prop-erties from the same product made from porcine UFH. For example, if enoxaparin (a LMWH) is produced from bovine hepa-rin in the future, this product could not be called enoxaparin because the structure and properties would be different and the activity would probably be different as well.

Implications for public standardsThe current USP Heparin Sodium mono-graph describes system suitability and ac-ceptance criteria for identity, purity, and strength that are designed strictly for por-cine heparin (17, see Table I).

The workshop presentations showed clear differences in the structures and biological activity profiles of porcine mu-cosal, bovine lung, and bovine mucosal heparin samples, thereby suggesting the need to have a separate monograph for bovine heparin sodium. The structural differences would necessitate separate identification reference standards (RS) for 1H NMR, chromatographic identity, and molecular weight determination tests for bovine heparin (see Table I). Depending on the tissue source(s) of bovine heparin, there may be a need to establish separate identification RS for bovine lung and bo-vine mucosal heparins.

Based on the testing of numerous sam-ples, the anticoagulant activity of bovine heparin is significantly lower than that of porcine in the laboratory. Most of the current bovine products gave potencies of about 100 IU/mg and some batches were estimated to be as low as 70 IU/mg. Con-sidering the monograph specification for porcine heparin is 180 IU/mg, it is likely that the clinicians will need to give more bovine heparin than porcine heparin by weight. Many years of safe bovine hepa-rin use suggest that the higher amounts needed, as compared with porcine hepa-rin, do not impact clinical efficacy. Clini-cal issues with bovine heparin, however, will need to be monitored carefully be-cause wider clinical use could identify unforeseen differences between porcine and bovine heparin. Furthermore, bo-vine heparin requires higher doses of protamine for neutralization.

Future bovine heparin production sites (both drug substance and drug product) should be under cGMP com-pliance. Supply chains including farms, slaughterhouses, and facilities that isolate, treat, store, and ship the bovine tissue need to follow the same steps and tests described for porcine heparin (18). These steps include, but are not limited to:• Determine the species origin to ver-

ify that the ingredient comes only from the correct species.

• Confirm the absence of OSCS and ruminant material contaminants from another species.

• Ensure that all heparin suppliers are audited and inspected regularly re-garding their documentation prac-tices and compliance with cGMP.

• Reject any lots containing mucosa from another species.

ConclusionHeparin is an essential, life-saving drug that is needed worldwide. To avoid drug shortages, FDA is considering reintro-duction of bovine heparin, which would diversify the supply chain by adding a bovine source to the currently used porcine heparin. Sourcing heparin from two species could greatly reduce vulner-ability to shortages when disease strikes one species, and could also reduce reli-

Pharmaceutical Technology November 2015 9

ance on one country as the primary source. The risks involved in reliance on one species from one country were clearly illustrated by the heparin crisis of 2007–2008, when adulterated porcine heparin from China caused numerous deaths and hundreds more adverse ef-fects (1). Currently, China is the source for roughly 60% of crude porcine hepa-rin used in the US and Europe.

Bovine heparin is currently being used in some countries and was used safely for more than 50 years in the US before manufacturers voluntarily with-drew it from the market during the BSE crisis in the UK. Despite concerns about bovine products, there are no known cases of BSE contamination of bovine heparin. If bovine heparin is reintro-duced in the US, methods for inacti-vating BSE could be applied to further reduce any risk. The other main safety issue with heparin is a severe adverse ef-fect called HIT, but bovine heparin does not appear to have higher rates of HIT than does porcine heparin.

Porcine and bovine heparins are dis-tinctly different in terms of their struc-tures and biological activities. These complex products may behave differently in clinical use than in laboratory testing, but if potency does in fact differ signifi-cantly in clinical use, this will need care-ful evaluation and perhaps dosage adjust-ment to avoid giving patients too much or too little heparin. If bovine-sourced hep-arin is reintroduced, supply-chain control will be critical, with frequent inspections of slaughterhouses and processing facili-ties for cGMP compliance. From the data presented at the meeting, bovine heparin and porcine heparin are not equivalent drugs, and therefore, they will require two different compendial monographs. The next steps are for manufacturers to bring bovine products to the regulatory agencies for evaluation and possible rein-troduction to the market after a 15-year absence.

AcknowledgementsThe participants in the USP 6th Work-shop on the Characterization of Hep-arin Products are acknowledged with gratitude.

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DisclaimerThis article ref lects the views of the authors and should not be construed to represent US FDA’s views or policies.

global heparin supply

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