Chromogenic Substrates S-2238

The current International Standard for thrombin is the Human a-thrombin 89/588 available from NIBSC. This is a high purity preparation of a-thrombin prepared from Cohn fraction III and assayed by a clotting time method against the first International Standard for thrombin, 75/157 (1).

The National Institute of Health standard (Lot J) is also commonly used for calibration and a study conducted by Gaffney PJ et al. (2) was focused on the relationship between the two standards, and between the International Units and the NIH Units. As a result of this study, based both on a clotting and a chromogenic assay (with the chromogenic substrate S-2238), 1 NIH-U corresponds to 1.15 IU.

In an article (3) it was shown that bovine thrombin has a higher amidolytic activity than human thrombin when the same NIH-U are compared. It was also underlined that the influence of b and g-forms, that were probably contaminating the bovine enzyme, might be the reason for this discrepancy.

In the same article it was concluded that 1 NIH-U bovine thrombin was equivalent to 3.4 nkat chromogenic substrate S-2238, and that 1 NIH-U of human thrombin was equivalent to 2.7 nkat chromogenic substrate S-2238.

From an earlier publication (4) 1 NIH-U of human thrombin corresponded to 2.5 nkat chromogenic substrate S-2238. The correspondence between NIH-U or IU of thrombin and the enzyme activity expressed in nkat, depends on the substrate, the enzyme preparation (content of a-, b- and g-thrombin) and the assay conditions.

From the article of Friberger (4), 1 µg thrombin corresponds to 2.2 NIH-U or 5.5 nkat chromogenic substrate S-2238 or to 0.02 plasma equivalent units. In another study (5), 1 µg thrombin corresponds to 3.1 NIH-U.

In the experiments done in Chromogenix (see table 3 of the catalogue) 1 µg thrombin was equivalent to 3 nkat chromogenic substrate S-2238 (human) or 4.4 nkat chromogenic substrate S-2238 (bovine). It might also be added that if all prothrombin is activated in 1 ml of human plasma, about 1.5 nanomoles or 17.5 NIH-U of thrombin are formed (5).

  1. Gaffney PJ et al.
    A collaborative study to establish an International Standard for alpha-thrombin. Thromb Haemost 67, 424-427 (1992).
  2. Gaffney PJ et al.
    The International and NIH units for thrombin How do they compare? Thromb Haemost 74, 900-903 (1995).
  3. Witt I et al.
    Test system with synthetic peptide substrates in haemostaseology.
    Eur J Clin Chem Clin Biochem 29, 355-374 (1991).
  4. Friberger P et al.
    Synthetic peptide substrate assays and fibrinolysis and their application on automates. In: Sem in Thromb and Haemost, Vol. 9, 281-300 (1983).
  5. Hemker HC.
    Handbook of Synthetic Substrates. Ed. Hemker HC, Martinus Nijhoof Publishers, 96-97 (1983).