344 PHOTOMECHANICAL CONSIDERATIONS 



Chromatophores were prepared by a method similar to that de- 

 scribed by Vernon and Ash (7) with the following modifications: Tris 

 buffer concentration was increased to 0.1 M, sonication time was de- 

 creased to 2 minutes, and the last two centrifugations each were in- 

 creased to 100,000 x^ for 1 hr, Chromatophores were stored unfrozen 

 at 4°C in the dark under argon. The capacity of the preparation to 

 produce a light- induced ESR signal remained at a high level for as long 

 as two weeks. The procedure for reducing the O2 tension in the ESR 

 sample cell consisted of 5 evacuations alternated with N2 flushings. 



The chromatophore suspensions were routinely adjusted to have 

 absorbancy values of 1,5 to 1.9 at 880 m/y in the ESR cell with a light 

 path of ca. 0,03 cm. The volume within the microwave cavity was ca, 

 0,06 ml. This represents a bacteriochlorophyll concentration of from 

 100 to 120 mg BChl./ml, The ESR data were obtained with a Varian 

 V4502 spectrometer equipped with 100 kc/sec field modulation. The 

 preparations were illuminated by a focused 1000 watt projection lamp 

 after filtering with 4.5 cm of H2O and an interference filter which 

 resulted in ca. 45% transmission at 880 m/i (Xmax.) and a band width 

 at one-half height of ca. 10 mfj. 



RESULTS 



Light induces an increase in the ESR signal of chromatophores at 

 a rate that is not possible to follow with our present equipment. On 

 the other hand, the decay of the ESR signal, when the light is turned 

 off, is slower and differences with time can be resolved. The addition 

 of DPIPH2 (reduced with AA) to chromatophores caused an increase 

 in the rate of decay (measured at one magnetic field value) as shown 

 in Fig. 1. When the oxidized form of DPIP was added, it also increased 

 the decay rate of the signal from chromatophores. However, larger 

 concentrations of the oxidized form were required, and it was con- 

 cluded that some of the dye became reduced in the presence of the 

 chromatophores. Anaerobic conditions were employed, and separate 

 experiments showed a slow endogenous reduction of DPIP under these 

 conditions. The decay was accelerated to a greater extent with in- 

 creasing concentrations of DPIPH2. 



A better resolution of the rates of formation and decay of the 

 chromatophore ESR signal in the presence of DPIPH2 was obtained 

 with a fast oscillograph recorder as shown in Fig, 2. This shows that 

 the decay and formation of the ESR signal in the presence of DPIPH2 

 both have half-times on the order of 100 msec. 



The symmetrical molecule TMPD, which readily forms relatively 

 stable-free radicals upon partial oxidation, has been shown by Jacobs 

 (8) to initiate coupled electron transport in rat liver mitochondria. 

 When this molecule was added in the completely reduced form to il- 



