Abstract
The distribution of two quinones with coenzyme Q activity which are broadly distributed in higher plants has been determined. Particulate fractions from spinach leaves, the white and green portions of Pandanus leaves, and the roots an(d shoots of corn seedlings have been examined with respect to the amount of coenzyme Q and Q2s4 present. Coenzyme Q is found in both white and green tissue and appears to be concentrated in mitochondria, whereas Q254 is concentrated in chloroplasts and chloroplast-containing tissue. It is proposed that coenzyme Q is involved in mitochondrial electron transport and that Q2s4 may be involved in photosynthetic electron transport.

PLANT PIIYSIOLOGY system, it cannot be treate(d as a homogeneous system since it is structurally complex with physiological specialization closely associated with this structural complexity. Generally speaking, the potentialitv for active synthesis and growth is restricted to the embryonic axis, while the cotyledlons and/or endospermi are primarily concerned with the storage and subsequent mobilization of reserve materials. Since a seed is heterogeneous and complex, investigations of the germination process mlust treat the seed in its complexity and study the physiology of the individutal organs and their interrelations. Such an approachi was suggested by Shermaln (15) but has received relatively little experimental treatment.

MIost dry seeds contain all essential elements for germination except water. However, in many seeds the simple uptake of water will not permit germination because the growvth of the embryo is blocked by the presence of a resting or dormant condition (see (2) and (13) for discussion of terminology). Crocker (3) described 6 classes of blocked growth which are obviously associated with blocks at dlifferent stages of germination and in different parts of the seed. It is clear in Crocker's classification that mechanical restriction of transport or expansion is relatively simple compared with "embryo dormancy" and "secondary dormancy." These latter, wNhile classified by Crocker as distinct types, probablv are identical (1, 6, 17, 20) and are closely similar to, if not identical with, the rest period of buds of trees (1, 6) . We will refer to the result of any block to growth which resides within the embryo as the rest period.

The rest period is frequently studied because of its importance in agriculture and forestry. However, rest is also of great theoretical importance because it results from a block to growth within the growing cells; understanding the nature of this block could contribute directly to the understanding of the factors which control cellular growth and development. However, it is much easier to define the rest period in writing than it is to measure it in the laboratory. For example, germinationi is generally measured as the emergence of the primary root from the seed coat. However the question arises as to whether growth by cell division or enlargement prior to rupture of the seed coat is part of germinationi. To avoid this difficulty Toole et al (18) defined germination as "the resumption of growth" and Evenari (4) coInsiders germination to be those processes preceding the resumption of growth. The rest period mav be defined as a blocking of germination, but we must recognize that this is not an adequate working definition for at least 2 reasons: 1) The block to growth may not be complete, thus some growth may continue during the rest period as in the embryos described in this paper; and 2) the rest period can only be defined in terms of failure to grow. In the laboratory, it is normal practice to remove the block to growth bv after-ripening at low temperatures and then to test for growth by placing the material at higher temperatures. There is, however, ample evidenice (1) that these higher temiperatures themselves can reverse the effects of low temperature and tend to re-establish the rest period; according to Crocker's terminology (3), secondlary dlormancy is establishedl. Thus it is essential to have a measure of the actual growth potential of the systemii wlhich at least minimizes this reversing effect. It is for this reason that recent workers such as Abbott (1) and Visser (20) use a more conmprehensive measure of "germination capacity" rather than germination, and(I a similar procedure is followed in the work presented in this paper.

In the experiments reported in this paper the cherry seed was chosen as experimiiental miaterial since it contains an emnbryollic axis \vith a well developed root andl slhoot, has a (lefiinite rest period (8, 15, 19) , andl is readily available. In early experimelnts, we found that low-tempel-ature after-ripening increases the respiration rate of cherrv seeds. We then investigated the relative effect of low temperature on the various seed organs and found that the respiration of the parts of the embryonic axis increases much more than the whole seed. figure 3 .

ANALYTICAL METHODS: If we examine a cherrv seed, we find that the amount of experimental material available in the embryonic axis and leaf primordia is only a very small part of that in the whole seed. For example, an average non-after-ripened seed has a dry weight of 56.5 mg and contains an embryonic axis with a dry weight of 405 ,ug and 2 leaf primordia each with a dry weight of about 12 ,ug. Thus in colmlparing changes in leaf primordia with changes in whole seeds, we are dealing with materials differing in size over a 1000-fold range. Since only a limited number of seeds can be dissected within a practical period of time, it has been necessary to use a range of methods, from the "conventional" to the "ultramicro."

Several measurements were made to compare the size of the seedl parts and to serve as a basis for expressing other data. Linear measurements on the leaf primordia and embryonic axis were made with an ocular micrometer in a 10 X dissecting microscope. Dry weights were obtained on these same organs using "fishpole" type quartz fiber balances (12) . Several balances were used; the size of the balances was chosen to give dry weights of the organs accurate to at least 1 06 Dry weights of whole seeds were obtained with a Roller-Smith balance sensitive to 0.02 mg. Cell counts were obtained using the maceration technique described by Sunderland andl Brown (16) . We have also studied changes in total nitrogen and phosphorus; these (lata will be presented in another paper. During the course of these investigations we attempted to obtain the volume of the embryonic leaf primordia and axis by the method of L(pvtrup (11) .

Briefly, this method involves measuring the reduced weight of an organism with the Cartesian diver balance (10) and the density from the level at which the organism comes to rest in a physiologically-inert density gradient. From these 2 measurements, the volume of the organism can be calculated. While the reduced weight measurement offered no problem,. density measurement proved difficult. The axis and leaf primordia are relatively dense, 1.06 to 1.08, in comparison with the amoeba, density 1.02, for which the method was originally developed. The solutions available for these densities were so viscous that it was difficult to find a definite rest point in the gradient. We used a high molecular weight dextran preparation ( Respiration, measured as oxygen uptake, was determined by 3 methods. In each case, the gas atmosphere was air and a temperature of 250 C was used. The rates were essentially constant for the period of 4 to 6 hours during which measurements were made. For whole seeds, standard Warburg techniques were used. Groups of 10 to 20 seeds were placedl on moist filter paper in the bottom of a 20 ml vessel, and measurements were made without shaking. For the embryonic axis, the differential volumeter described by Grunbaum et al (7) was employed. The particular instruments used were made with capillaries of 0.8 to Respiration of leaf primordlia wx-as measured using standlardI Cartesian diver techniques ( (9) and other papers from the Carlsberg Laboratory). Figure 2 illustrates the filling schedule for the various experiments. All solutions were prepare(d using water redistilled from an all-glass (Pvrex) still. The oxygeen consumption per diver was kept at a minimum of about 10 X 10' ul/hr by using pairs of leaf primordia from non-after-ripened seedls and from seedls afterripened at 25°C. With seeds after-ripene(d at 50 C a single primordium was placed in a diver. In most cases, this resultecl in an oxygen uptake below 80 x 10-' ,l/hr/diver although a few rani as high as 140 X 10-,ul/hr/diver. However, empirical measurements showed that even with these high rates, diffusion of oxygen to the primordium and absorption of carbon dioxide were not limiting.

In experiments with 2,4-dinitrophenol (DNP). the normal respiration rate was measured for a period of 2 to 3 hours before the seal containing the leaf primordium was mixed with a side drop containing DNP. The rate of oxygen uptake in the presence of DNP was allowed to stabilize for about 1 hour and the linear rate was then measured during the periodl 1 to 3 hours after mixing. Early experiments with DNP indicated that respiratory rates during these rather long experiments were stabilized somewhat if the primordia were suspended in 0.5 M sucrose in A1/30 phosphate buffer pH 5.3. Other than this slight stabilizing effect, the sucrose used in this series of experiments seemed to have no influence on respiratory rates.

For cyanide inhibition measurements. the Ca (CN) 2-Ca(OH) 2 method of Robbie ( 14) wvas used, with the HCN concentrations calculated as for the XVarburg apparatus. Two measurements were made for each primordium. First a normal rate was determined during a 2 to 3 hour period by absorbing the C02 in a seal of 0.1 N KOH. This seal was then replaced with the appropriate Ca(CN) 2-Ca(OH)2 mixture anl the linear rate of oxvgen uptake was determined for a period of 2 to 3 hours after the initial exposure to HCN. Although in the Cartesian diver the Ca(CN)2-Ca(OH)2 mixture is not as effective in C02 absorption as KOH, probably because of the formation of a surface film of Ca(CO3)2, empirical measurements showed that the rate of C02 absorption did not become limiting.

The sampling procedure followed in these experiments involved analyses of non-after-ripened seeds and of seeds after-ripened for 4, 8, 12, and 16 weeks. The non-after-ripened seeds were sterilized and allowed to imbibe wvater for 2 to 5 (lays before analysis. At each of the subsequent time periods the measurements with living material were completed within a 3 to 5 day period. At each sampling time 1 group of 15 seeds from each treatment was removed from after-ripening conditions and was immediatelv excised and the respiration rate of 1 or both leaf primordia measured. One primordium was then used for cell counting and the other was (lried and used for dry weight measurement and total nitrogen analvsis. The embryonic axis was maceratedl for cell counting. Another group of 15 seeds was excised and the leaf primordia driedl for total phosphate analysis while the embryonic axes, in groups of 5, were used for respiration measuremenetts. These were then dried and used for drv weights and total nitrogen and phosphate analysis. Three groups of 10 seeds each were used for respiration measuremeints on whole seeds and were then excisedl for growtth measuremllents. Finally, 20 to 30 seeds were used to measure the germination of intact seeds. It shouldl be noted in the sampling procedure for leaf primor(lia that the primordia from any one seed are so closely matched that a "matched leaf" technique could have been emi)loyed.
RESULTS
The data obtained show that after-ripening at 50 C leads to 5 % germination after 12 to 16 weeks: a high germination percentage is not achieved under these conditions until after 18 weeks. After-ripening at 250 C does not lead to normal germination. As shown in figure 3 , excision of the embryo leads to much earlier growth, according to the classification used almost full capacity after 16 weeks at 5°C. Excision of embryos after-ripened at 250 C leads to a slight increase in growrth capacity by 16 weeks.
RESULTS
One of the qualitative characters taken as a measure of growth is a change in the growing portions of the embryo from a white opaque to a yellow translucent appearance. This change is apparently due to 2 factors: 1) the synthesis of pigments and 2) the emptying of the reserve materials from the cells. The latter can be seen clearly in microscope sections of the material. This means that growth in these cells is accompanied by utilization of storage materials or their conversion into protoplasm and cell wall material in situ. For this reason, dry weight or total nitrogen may be poor bases for expressing other data. We have therefore investigated such other measures of growth as organ size and number of cells. 
POLLOCK AND OLNEY-CHANGES IN THE AFTER-RIPENING SEEI)
In figures 4 and 5 are presented data on changes in the embryonic axis and leaf primordia during afterripening at 50 C and at 25°C. A number of changes are obvious and fairly constant, although the exact times of these changes are not necessarily synchronized in the axis and leaf primordia. Specifically, at 50 C both the axis and leaf primordia show an increase in length roughly correlated with the increase in the growth capacity. This increase in length is due at least in part to the production of new cells. The increase in length and number of cells is also accompanied by an increase in the total dry weights of axis and leaf primordia (figures 4 C and 5 C). In the axis' the dry weight per cell tends to drop and then increase again (figure 6), while in the leaf primordia it tends to drop, although somewhat erratically because the rate of cell division is not constant. The difference in amount of variability between leaf primordia and axis in this respect is magnified by the fact that (luring the 1st 4 weeks of after-ripening the increase in cell number in the axis could be accounted for by a single division in 15 % of the cells, while in the leaf primordia it was the equivalent of a single division in 51 % of the cells. Thus cell division in the leaf primordia is proportionately much more active. It should be noted that the dry weight per cell in the leaf primordium is less than half that in the axis. This is at least partially the result of a difference in cell size as can be seen in microscope sections.
POLLOCK AND OLNEY-CHANGES IN THE AFTER-RIPENING SEEI)
Whether this also represents a difference in amount of stored material cannot now be determined.
POLLOCK AND OLNEY-CHANGES IN THE AFTER-RIPENING SEEI)
In comparing embryos after-ripened at 50 C, and thus being prepared for active growth, with embryos after-ripened at 250 C and increasing little in growth capacity, a number of differences are obvious. According to most of the measurements made, increases with after-ripening time at 50 C are not paralleled by increases in embryos after-ripened at 250 C. However, if we examine the changes during the 1 st 4 weeks of after-ripening, we find that these are actually quite similar at both 50 C and 250 C, the divergence appears in most cases only after 4 weeks.
POLLOCK AND OLNEY-CHANGES IN THE AFTER-RIPENING SEEI)
It should be recognized that the growth shown in figures 4 and 5 can be seen only by means of the precise measurements used to obtain these data. On the other hand, the data use(l to estimate the growth PLANT PHYSIOLOGY capacity as presented in figure 3 represent changes of a much higher order of magnitude, actually growth visible to the naked eye.
POLLOCK AND OLNEY-CHANGES IN THE AFTER-RIPENING SEEI)
In figure 8 the changes in respiration rate in the various parts of the seed are presented as a function of after-ripening time and temperature. Because of the great difference in absolute amounts, the data are expressed on a percentage basis, using the rates for non-after-ripened seeds as 100 %. For the whole seed, embryonic axis, and indiviclual leaf primordium this basic value is 5.29, 0.246 and 11.1 X 10-' btl/hr/ individual or organ, respectively. Figure 8 A shows that the respiration of whole seeds after-ripened at 50 C increases during after-ripening. However, this increase for whole seeds, a maximum of about 70 %, is small compared to the increase for the embrvonic axis and leaf primordia of almost 600 % in 16 weeks.
POLLOCK AND OLNEY-CHANGES IN THE AFTER-RIPENING SEEI)
It should be noted that for both the leaf primordia and axis this increase is almost linear with time and does not show a 4 week lag as (loes the growth capacitv and increase in dry weight of the embryonic axis.
POLLOCK AND OLNEY-CHANGES IN THE AFTER-RIPENING SEEI)
In seeds after-ripened at 250 C the respiration rate of whole seeds actually declines to less than half that originally present. The respiration rate of the embryonic axis and leaf primordiumii does not change significantly with time. At 250 C after-ripening has little effect on the total oxygen uptake of the axis and leaf primordium cells, with a decrease apparent in the seed as a whole.
POLLOCK AND OLNEY-CHANGES IN THE AFTER-RIPENING SEEI)
Recognizing the great increase in respiration rate which precedes and accompanies an increase in growth capacity, the question arises as to whether this increase is due to the activation of enzymes previously present, to the synthesis of new enzymes, to the synthesis of other possible rate-limiting factors, or to the transport of such rate-limiting factors from the cotyledons to the growing cells of the embryo. In an attempt to answer these questions, 2,4-dinitrophenol (DNP) was used as an uncoupling agent to obtain an approximate measure of the rate-limiting step in the respiratory chain.
POLLOCK AND OLNEY-CHANGES IN THE AFTER-RIPENING SEEI)
Preliminary experiments with leaf primordia showed that DNP can increase respiration at a concentration of 3 X 10-' M (fig 7) . These experiments also showed that the degree of increase is greater in non-after-ripened than in after-ripened primordia. Therefore, as part of the experiment previously described, DNP at 3 X 10-' M was added to leaf primordia after the normal respiratory rate had been measured. The results are shown in figure  9 . Figure 9 A figure 10 with the results calculated on a per cell basis. If we then calculate the actual respiration rate of the leaf primordia as a percentage of their respiratory capacity, it is clear (fig 9 C) that the percent of utilization climbs more or less continuouisly during after-ripening at 50 C, but at 25°C it dleclines very sharply during the 1st 4 weeks, thereafter rising somewhat with time but never getting to much over half the degree of utilization shown by the leaf primordia from seeds after-ripening at 50 C. Wlhile this experiment was terminated prior to much actual growth, other experiments with more advanced material, such as that shown in figure 7, in(licate that this trend for more complete utilization of the respiratory potentialities of the system continues at least into the early stages of active growth.
POLLOCK AND OLNEY-CHANGES IN THE AFTER-RIPENING SEEI)
The fact that the stimulation by DNP declines as the system approaches a growing condition suggests that the rate-limiting step in respiration shifts with the degree of after-ripening and changing growth capacity. This shift seems to be from the phosphorylation system toward a rate-limiting step in the respiratory chain itself, although these data alone might equally indicate a rate limited by respiratory substrate. However, evidence for the respiratory chain as the rate-limiting step is shown in figure 11 , where cvanide sensitivity curves for non-after-ripened and afterripened (15 to 18 weeks at 50 C) leaf primordia are shown. In this case, the after-ripening was carried somewhat closer to actual growth than in the previous case, and the DNP increase was comparable to that shown for after-ripenedl primordia in figure 7 . These (lata show that after-ripened primordia are more sensi- inhibition, than are non-after-ripened primordia. This is what would be expected if the capacity of the respiratory chain itself became more the limiting factor in respiration.
DIscussIoN
In their recent review article, Toole et al (18) noted that one of the first evidences of the onset of germination is the increase in respiration rate, but that the sequence of enzymatic events has not been established. Thus the data presented in this paper need to be interpreted relative to the normal germination process as well as to the rest period. Indeed, it is possible to consider that the changes which occur slowly during a period of low temperature after-ripening are similar to those which must occur very quickly during the 1st hours of germination in a seed germinating without a rest period.
DIscussIoN
One of the interesting conclusions from these data is that, for the cherry seed at least, early growth of the embryo is not restricted to cell elongation, but includes cell division. It is particularly interesting to note that in the leaf primordia a significant number of cell divisions occur during the 1st 4 weeks, and occur equally at 50 C and 250 C. At this time the embryo is not capable of any further growtlh. This might be interpreted to mean that these early cell divisions are independent of the rest period block. possibly a continuation of development which was arrested by seed maturity.
DIscussIoN
One (5) suggests that excision of parts of the cotyledons, while making the plants smaller, has no specific effect on breaking the rest period. However, this does not answer the question since it is possible that the stimulus directing translocation arises within the growing cells and that the rest period involves failure of this stimulus. It is equally possible that the failure of the translocation stimulus is just one of many secondary effects of a block elsewhere in metabolism. In other words, we cannot yet distinguish cause from result in this situation.
DIscussIoN
The most striking changes we have noted during after-ripening have been associated with the respiratory mechanism of the cells. The data show that the respiration rate of both the embryonic axis and the leaf primordia from seeds after-ripened at 50 C rises linearly with time of after-ripening; there is no obvious difference in response between the axis and leaf primordia. It is particularly interesting to note that the respiratory rate increases from the beginning of after-ripening, while the growth capacity rises only after a 4-week lag. Thus the increase in respiration appears to precede the renmoval of the rest period block. It is particularly interesting to examine the leaf primordia and the changes in respiratory capacity which occur within these 1st critical 4 weeks of afterripening. At 50 C the respiratory rate and respiratory capacity rise together; this increase is accompanied by division of the equivalent of 51 % of the original cells. At 250 C the cells dividle at the sanme rate, and the respiratory capacity rises almost as rapidly as at 50 C but the respiratory rate fails to jenkinschristopher@example.net. With further after-ripening the rate and capacity both climb at 50 C but the respiratory capacity declines while the respiratory rate remains constant at 250 C. These changes strongly suggest that the initial effect of after-ripening at 50 C is to increase the efficiency with which the respiratory enzymiie system is utilized, hence increasing the supply of energy available for synthesis and growth. In contrast, at 2°5 C synthesis appears to start normally aind then come to a halt for lack of available energy. A sonmewlhat similar situation has been described by Evenari (4) for the lettuce seed.
DIscussIoN
WVhat does the termi "respiratory capacity" mleani in the biochemical meclhanismiis of the cell ? Dinitrophenol is usually considered to act by uncoupling mitochondIrial oxidations from phosphorlation. If DNP acts to increase respiration, it may therefore be as- figure 10 , during the 1st 4 wveeks after-ripening at 5°C the enzyme must be synthesized (or activated) in a quantity sufficient not only to supply the newly-formed cells but also to increase the quantity within these cells. This increase must continue through the period 4 to 8 weeks when no cell divisions occur. Then from 8 to 12 weeks, when more frequent cell divisions occur, the synthetic rate is not adequate to maintain the rate per cell constant and the respiratory capacity per cell drops. This does not mean that synthesis stops, because the increase in total respiratory capacity per primordium continues to increase at a more or less constant rate; it mere4y means that cell division and synthesis are not precisely linked.
DIscussIoN
The changes in respiratory capacity with afterripening at 25°C are also interesting. Accompanying the cell divisions during the 1st 4 weeks, there would appear to be a considerable amount of synthesis. It is not clear however whether the subsequent decline indicates a destruction of enzyme, its conversion to an inactive form, or a shift to a new rate-limiting step in respiration. It is perhaps significant that the reduction is to, but not below, the original level in non-after-ripened seeds.
DIscussIoN
In considering the present status of our knowledge of the mechanism of the rest period, it is clear that rest may be considered to involve at least 1 primary block to growth which will have many secondary effects on the blocked cells. Regardless of whether this primary block is in the nature of a chemical inhibitor, a deficiency of some essential hormone or cofactor, or some other type of mechanism, this primary block must operate through the metabolism of the affected cells and, in turn, must be generated and removed by changes in cell metabolism. The problem then is to construct a tinmetable of metabolic changes and a flouw sheet of such compounds as nutrients and hormones sufficiently precise to permit identification of the metabolic block. together with its mode of generation and removal.
SUMMARY
The changes associated witlh after-ripening in resting cherry seeds have been investigated by utilizing a combination of "conventional" and "ultramicro" analytical methods. \With these nmethods, it has been possible to deternmine changes in the wlhole seeds and, more important, in the potentially growing cells of the embryo. In seeds after-ripened at 50 C, the capacity for growth develops linearly with time after a lag of 4 weeks. This capacity for growth is parallele(d in the embryonic axis by an increase in dry weight, indicating translocation of material from the cotyledons, but is preceded by cell divisions and a sharp increase in respiration rate. Using 2,4-dinitrophenol to obtain a measure of respiratory capacity, we find that the increase in respiration is paralleled by an increase in respiratory capacity. In contrast, in seeds after-ripened at 250 C growth capacity does not develop and little translocation to the potentially growing cells of the embryo is evident. The respiratory rate remains approximately constant but the respiratory capacity rises, accompanied by a significant nunmber of cell divisions, within the 1st 4 weeks of afterripening, but thereafter declines to its original level.
SUMMARY
We interpret these results as suggesting that one cause of the breaking of the rest period may be the increased availability of energy to the embryo, possibly by an increase in the supply of phosphate acceptors. Whether this is the primary reaction responsible for breaking the rest period or just one of many secondary reactions which follow activation of the cells is not clear.
SUMMARY
tainedl from the National Bureau of Standards, and solutions of these sugars were prepared by dissolving 5 mg of the sugar in 5 ml of water.
SUMMARY
Synthetic phenylglucoside was obtained by the method described by Helferich and Schmitz-Hillebrecht (2) . The position of phenylglucoside on the chromatograms was determined by placing the chromatogram over a calcium tungstate screen and irradiating it with ultraviolet light (2537 A maximum).
SUMMARY
The phenylglucoside appeared as a blue-violet spot onl the chromatogram and as a dark shadow on the screen.
SUMMARY
Barlev or wheat leaves were fed the solutions of labeled sugar by placing the leaf base in a tube with 100 1Al of the sugar solution with or without phenol. After the leaf had taken up this solution, a continual supply of water was provided. At the end of the experiment, the leaves were ground in liquid nitrogen, and the powder was taken up in water and immediately heated in a boiling water bath. After separation of the supernatant liquid from the solid debris, the extract was concentrated, and its components separated by the chromatographic procedures previously used (4). The C'4-labeled compound suspected of being phenylglucoside was separated by these procedures and located on the chromatograms by radioautographic techniques. The area was cut out and eluted with distilled water. The eluate was then again chromatographed in an ethyl alcohol: 15 % acetic acid (8: 1 by volume) solvent system in order to separate any contaminating phenol. The C'4-labelerl compound (phenylglucoside) was then eluted from the chromatograms. and its ab-

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