Sodium heparin

Briefly, 15 mL Histopaque 1119 (Merck, Darmstadt, Germany) was overlaid with 15 mL Histopaque 1077 solution. Then, 20 mL of heparinized blood (100 I.U. sodium heparin/mL, Roth, Karlsruhe, Germany) was carefully overlaid, followed by centrifugation at 630× g without braking at 18 °C for 25 min. The mononuclear cell fraction located between the plasma and Histopaque 1077, and the second layer containing granulocytes were collected. After washing, and resuspension of cell fractions in phosphate-buffered saline (PBS without Ca++/Mg++; glucose (1 g/L; 5.5 mM), Merck, Darmstadt, Germany), cell number was determined using an ADVIA-120 blood cell counter (Siemens, München, Germany). One part of the granulocyte suspension was used for degranulation experiments (see Section 2.1.3), another part was used for direct measurement of enzyme activities.

Study approval from the local Ethical Committee of the Kanton Zurich (KEK Zurich) was received under No. 2012-0111. The methods were carried out in accordance with the approved guidelines. Whole blood from healthy volunteers was obtained at the University Hospital Zurich after receiving informed consent according to standardized guidelines. For experimental reasons, blood samples were anti-coagulated with heparin at a low concentration. Up to 60 ml of venous blood per donor was collected in vacutainer tubes (BD Vacutainer™ No Additive (Z) Plus Tubes) and heparinized upon withdrawal to a final concentration of 3IU sodium heparin (Carl Roth, Germany, Eur. Ph., 120 IU/mg) per ml blood. Blood of every donor was screened for hematogram with automated leukocyte differentiation, coagulation status and fibrinogen concentration by the Institute for Haematology and for Haemoglobin A1c (HbA1c) by the Institute for Clinical Chemistry of the University Hospital Zurich, as a measure for non-diagnosed diabetic conditions. The volunteers were classified as healthy as analysed parameters were within the normal clinical range.

Starting materials and preparation of the Rumex acetosa L. extract RA have been described recently [21] (link). Isolation and analytical characterization of proanthocyanidins from RA have been reported by Bicker et al. (2009) [24] . Structural features, sources and purity of flavan-3-ols, oligomeric proanthocyanidins, hydrolyzable tannins, depsides and building blocks of tannins used for antiviral bioassays used in this study are given in Figure 1 and Table 1. Sodium heparin (100,000 IU/g) was purchased from Roth (Karlsruhe, Germany).