Background: Antiplatelet, anticoagulant and fibrinolytic activities of stem bromelain (EC 3.4.22.4) are well described, but more studies are still required to clearly define its usefulness as an antithrombotic agent. Besides, although some effects of bromelain are linked to its proteolytic activity, few studies were performed taking into account this relationship.

Objective: We aimed at comparing the effects of stem bromelain total extract (ET) and of its major proteolytic compounds on fibrinogen, fibrin, and blood coagulation considering the proteolytic activity.

Methods: Proteolytic fractions chromatographically separated from ET (acidic bromelains, basic bromelains, and ananains) and their irreversibly inhibited counterparts were assayed. Effects on fibrinogen were electrophoretically and spectrophotometrically evaluated. Fibrinolytic activity was measured by the fibrin plate assay. The effect on blood coagulation was evaluated by the prothrombin time (PT) and activated partial thromboplastin time (APTT) tests. Effects were compared with those of thrombin and plasmin.

Results: Acidic bromelains and ananains showed thrombin-type activity and low fibrinolytic activity, with acidic bromelains being the least effective as anticoagulants and fibrinolytics; while basic bromelains, without thrombin-like activity, were the best anticoagulant and fibrinolytic proteases present in ET. Procoagulant action was detected for ET and its proteolytic compounds by the APTT test at low concentrations. The measured effects were dependent on proteolytic activity.

Conclusion: Two sub-populations of cysteine proteases exhibiting different effects on fibrin (ogen) and blood coagulation are present in ET. Using well characterized stem bromelain regarding its proteolytic system is a prerequisite for a better understanding of the mechanisms underlying the bromelain action.

背景：菠萝蛋白酶的抗血小板，抗凝血和纤溶活性（EC 3.4.22.4）已得到很好的描述，但仍需要进行更多研究以明确定义其作为抗血栓形成剂的作用。此外，尽管菠萝蛋白酶的某些作用与其蛋白水解活性有关，但很少考虑到这种关系进行研究。

目的：考虑到蛋白水解活性，我们旨在比较茎菠萝蛋白酶总提取物（ET）及其主要蛋白水解化合物对纤维蛋白原，纤维蛋白和血液凝固的影响。

方法：对从ET色谱分离的蛋白水解级分（酸性菠萝蛋白酶，碱性菠萝蛋白酶和凤梨苷）及其不可逆抑制的对应物进行了分析。电泳和分光光度法评估对纤维蛋白原的作用。通过纤维蛋白平板测定法测定纤维蛋白溶解活性。通过凝血酶原时间（PT）和活化的部分凝血活酶时间（APTT）测试评估对凝血的影响。将其效果与凝血酶和纤溶酶的效果进行了比较。

结果：酸性菠萝蛋白酶和凤梨苷显示出凝血酶类型的活性，且纤溶活性低，酸性菠萝蛋白酶作为抗凝血剂和纤溶蛋白的效果最低；而没有凝血酶样活性的碱性菠萝蛋白酶是ET中最好的抗凝血和纤溶蛋白酶。通过APTT试验在低浓度下检测到ET及其蛋白水解化合物的促凝作用。测得的效果取决于蛋白水解活性。

结论：ET中存在两个亚群的半胱氨酸蛋白酶，它们对纤维蛋白（基因）和凝血有不同的作用。对于蛋白质水解系统，使用特征明确的茎菠萝蛋白酶是更好地了解菠萝蛋白酶作用机理的先决条件。