428 M. D. Kamen and R. G. BartscH 



the Chromatiwn preparation, using the customary methods for iron analysis, 

 based on colour development with 1,10-phenanthroline, after acid or alkaline 

 ashing (Sandell, 1944; Drabkin, 1941). The R. rubrum preparation has been 

 somewhat more amenable (Bartsch and Kamen, 1958). It yielded acceptably- 

 reproducible iron assays after wet-ashing with nitric acid (Sandell, 1944), but 

 not with alkaline peroxide (Drabkin, 1941). The iron content of R. rubrum 

 RHP indicated a minimum molecular weight of 3 1 ,000 which was reckoned 

 to be high rather than low owing to incomplete recovery of iron, often met 

 with in assay of standard cytochrome c preparations using this method. 

 Inasmuch as the molecular weight determined, either by physical methods or 

 by spectroscopic means using the derivative pyridine haemochrome, was close 

 to 28,000, it was concluded that no iron was present in excess of that accounted 

 for by the haem. 



Early studies on the mixed RHP-cytochrome c preparation originally 

 obtained in impure and partially denatured form from Chromatium (Newton 

 and Kamen, 1956) indicated considerable iron in excess of that which could 

 be attributed to the haem moieties. 



Recently, after failure of our preliminary attempts to determine the iron 

 content of the Chromatium RHP using wet-ashing and colorimetric pro- 

 cedures, we have begun a collaborative research with Dr. B. L. Vallee, who, 

 with his colleagues, has developed accurate methods for analysis of a wide 

 variety of trace metals in biological systems using dry-ashing and arc or 

 spark emission spectroscopy (Vallee, 1955). We cannot provide definitive 

 data as yet, but tentative results obtained in the first experiments on Chro- 

 matium RHP show iron contents which are in accord with the haem content. 



Since the spectroscopic emission methods can be readily extended to 

 determine all trace metals of possible interest, we plan to investigate the 

 possibihty that not only iron but other metals such as copper, manganese, 

 cobalt, nickel, and zinc may be present. It is by no means certain that iron 

 is the only functional metal in these preparations. The close association 

 of RHP and some other haematin compounds with the photosynthetic 

 system raises many questions about a possible photochemistry of these 

 compounds. Such a possibility is diflScult to conceive on the basis that iron 

 is the only metal atom present, but would become real should it be demon- 

 strated that metals are present which are photochemically active as bound 

 ions or porphyrin chelates. 



Enzyme Properties 



Little is known about enzymic activities or cellular functions of RHP. 

 In R. rubrum extracts, as well as in Chromatium extracts, there are diphospho- 

 pyridine-nucleotide (DPN)-l inked cytochrome c reductases, very active 

 diaphorases, and catalases. The cytochrome c reductase of these bacteria 

 can reduce RHP using reduced diphosphopyridine nucleotide (DPNH) 



