Malondialdehyde Adduct to Hemoglobin: A New Marker of Oxidative Stress Suitable for Full-Term and Preterm Neonates

Table 1

Derivatization of standards and samples.

	Standards (from 76 to 608 nM)	MDA-Hb
Sample volume	100 μL TMP	100 μL
Derivatization	100 μL DAN 5.8 mM in HCl 2.4 N + internal standard
Mixing, incubation for 180 min at 37°C
Hb precipitation	100 μL MPA 10%, mixing, centrifugation 3 min, 15000 × g
	200 μL supernatant for the next step
Stabilization at pH 2.0*	100 μL (NaOH 1 N + KH2PO4 300 mM pH 2.0)

*At every new preparation of both reagents, the volumes are adjusted to obtain pH 2.0 with a fixed volume of 300 μL (DAN: diaminonaphthalene; MDA: malondialdehyde; MPA: metaphosphoric acid; TMP: tetramethoxypropane).