J. W. HASTINGS AND W. D. McELROY 



259 



a specific requirement (McElroy and Green, unpublished). The puri- 

 fied enzyme acts as a typical diaphorase (McElroy and Green, 1954). 

 The overall kiminescent reaction may be written: 



FMNH2 + RCHO + O2 + enzyme -^ light + products 



None of the products has been identified as yet, although we have 

 evidence that the aldehyde is destroyed during the reaction. The pH 

 optimum for this reaction is 6.9 (Fig. 1). The luminescent reaction 



Fig. 1. Effect of pH upon bacterial luminescence. Buffer as described in text. 



does not involve free hydrogen peroxide. The level of luminescence is 

 unaflFected by added hydrogen peroxide or crystalline beef catalase. 

 This of course does not rule out the possibility that organic peroxides 

 may be functional in the system. 



In the course of our investigation of the FMN requirement we ob- 

 served that the purified enzyme without added FMN still emitted 

 appreciable (ca. 5% max.) hght when DPNH was added. Microbio- 

 logical assays using Lactobacillus casei confirmed the presence of 

 flavin in the enzyme, and numerous unsuccessful attempts were made 

 to separate it from the enzyme. Irradiation at 5° C with a Keese lamp 

 produced a marked effect in this respect. Figure 2 illustrates the type 

 of result obtained. During an initial period of irradiation added FMN 

 does not give additional stimulation. With continued irradiation, how- 

 ever, the stimulation by FMN rapidly increases until the requirement 

 may be considered essentially absolute. 



The more striking effect of irradiation is illustrated in Fig. 3. Sam- 

 ples of the enzyme were removed at intervals and assayed. Both with 



