The placental transfer and detection of 14C-carmoisine metabolites by HPLC combined with a radioactivity monitor (RAM)

Carmoisine (Azo Rubine; Ext. D&C Red No 10; C.I. (1956) No 14720; E 122) is the disodium salt of 2-(4'-sulpho-1'-naphthylazo)-1-naphthol-4-sulphonic acid, a water soluble sulphonated azodye currently permitted for food in the EEC member countries. Because of the lack of information on the metabolism and placental transfer of the red food color and its metabolites the absorption fate and excretion of 14C-carmoisine has been determined in pregnant rats together with the presence of 14C-metabolites in vivo and in vitro preparations. Pregnant rats received 14C-carmoisine (200 mg/kg; 25 μCi) by gavage on days 16-19 of gestation. The animals were killed and the maternal tissues, amniotic fluid, placentae, foetal membranes and fetuses were analyzed for radioactivity. No evidence of the transplacental transfer of 14C-carmoisine or its metabolites was obtained. More than 90% of the radioactivity was excreted in the faeces and urine within 64 h. The results suggested that there was no significant absorption of the azodye and no preferential concentration of the red food color or its metabolites in any particular tissue. The results are comparable to those obtained in a similar study in male rats. Analyses by high performance liquid chromatography (HPLC) combined with a radioactivity monitor of the urine and faeces of the animals and incubation of 14C-carmoisine under anaerobic conditions with bacterial suspensions of human faeces show the same radioactivity profile. Five radioactive peaks were present in the radiochromatogram in addition to unmodified Carmoisine. The mean peak shows half of the specific activity of Carmoisine, at the retention time and with the UV spectrum of authentic naphthionic acid. The results demonstrate the value of transplacental studies because with chemicals that are poorly absorbed from the intestine and which do not cross the placenta, they will assist in the evaluation of adverse maternal effects which may result in foetal toxicity. Furthermore, with similar results obtained in vivo and in vitro one may use the in vitro preparations to prepare these compounds in amounts suitable for their identification by further chemical methods.