1560 PHOTOCHEMISTRY OF CHLOROPHYLL CHAP. 35 



involved in both reactions is present in a concentration of about 1 mole- 

 cule per 200-2000 molecules of chlorophyll. If the particles in a chloroplast 

 dispersion contain 0.05 g. chlorophyll in 1 g. dry weight, have a density of 

 about 1, and contain 50% water, then one molecule chlorophyll is contained 

 in about 7 X 10"^ ju*, and one molecule of the Umiting enzyme, in about 

 1.5-15 X 10~^ M^ In particles formed by disintegration of grana, the 

 chlorophyll concentration will be higher. It was estimated as 0.2 mole/ 

 liter (or more) by Rabinowitch (1952); this corresponds to an average 

 volume of 1 X 10 ~V^ (or less) per molecule of chlorophyll, or between 2 

 and 20 X 10~ V (or less) per molecule of the postulated limiting enzyme. 

 Consequently, when particles in a grana dispersion become smaller than 

 10-^ M^ hi volume, or <0.01ju (10 mju) hi linear dimensions, an increasing 

 number of them will contain not even a single molecule of the limiting en- 

 zyme, and will thus be unable to contribute to the reaction at all. We 

 have seen above that according to French, many of the particles in his dis- 

 persion appeared to be <2 m^ under the electron microscope! 



The fact that chloroplast dispersions usually show a Tyndali cone in 

 visible light merely indicates that the linear dimensions of some of the 

 particles are > 100 ni/n; there may be enough smaller particles present to 

 account for the drop in activity. 



Thomas, Blaauw and Duysens (1953) made a direct comparison of par- 

 ticle size and photochemical activity of spinach chloroplasts, disintegrated 

 by a magnetostriction oscillator. In air the disintegration led to the loss 

 of about 50% of photochemical activity in 30 seconds, but in nitrogen the 

 decrease in activity was only 5% after a whole minute of supersonic treat- 

 ment. The active suspension, obtained in this way, was subjected to frac- 

 tional centrifugation to obtain fractions of approximately uniform size. 

 The distribution of particle sizes in each sample was observed under the 

 electron microscope, and found to possess a sharp peak. Size measure- 

 ments became unreliable below 3 m^ (limit of the resolving power of the 

 electron microscope used), and above 15 m^ (where fractional centrifugation 

 lost its effectiveness). Figure 35.15A shows photochemical activity {i. e., 

 rate of Hill reaction in the light-limited state, with quinone as oxidant) as 

 function of (mean) particle volume. Characteristic is the sudden decline 

 in activity of particles when their average volume declines below 1.5 X 

 10-^ li\ Thomas and co-workers estimated that the particles became in- 

 active when they contained less than between 40 and 120 chlorophyll mole- 

 cules (rather than between 200 and 2000 molecules, as predicted above). 



The decline in activity mth diminishing size is less sharp in saturating 

 hght [210 kerg/(cm.2 sec.)] than in limiting light. This is to be expected 

 if particles below the "critical size" are not uniformly inactive but merely 

 possessed of a statistical probability of being inactive, because some of them 



