A spectroscopic investigation of the complexes formed between the Pb(II) ion and D-penicillamine (H2Pen), a chelating agent used in the treatment of lead poisoning, was carried out on two sets of alkaline aqueous solutions with CPb(II) ≈ 10 and 100 mM, varying the H2Pen/Pb(II) mole ratio (2.0, 3.0, 4.0, 10.0). UV-vis. spectra of the 10 mM Pb(II) solutions consistently showed an absorption peak at 298 nm for S− → Pb(II) ligand-to-metal charge-transfer. The downfield 13C NMR chemical shift for the penicillamine COO− group confirmed Pb(II) coordination. The 207Pb NMR chemical shifts were confined to a narrow range between 1806 and 1873 ppm for all Pb(II)-penicillamine solutions, indicating only small variations in the speciation even in large penicillamine excess. Those chemical shifts are considerably deshielded relative to the solid-state 207Pb NMR isotropic chemical shift of 909 ppm obtained for crystalline penicillaminatolead(II) with Pb(S,N,O-Pen) coordination. The Pb LIII-edge extended X-ray absorption fine structure (EXAFS) spectra obtained for these solutions were well modeled with two Pb-S and two Pb-(N/O) bonds with mean distances 2.64 ± 0.04 Å and 2.45 ± 0.04 Å, respectively. The combined spectroscopic results, reporting δ(207Pb) ~1870 ppm and λmax ~ 298 nm for a PbIIS2NO site, are consistent with a dominating 1:2 lead(II):penicillamine complex with [Pb(S,N,O-Pen)(S-HnPen)]2−n (n = 0 – 1) coordination in alkaline solutions, and provide useful structural information on how penicillamine can function as an antidote against lead toxicity in-vivo.