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A rapid method for determining the efficiency of biosynthesis of plant biomass
J. theor. Biol, (1987) 128, 109-119
A Rapid Method for Determining the Efficiency of Biosynthesis of Plant Biomass N. VERTREGT AND F. W. T. PENNING DE VRIES
Centre of Agrobiological Research, P.O. Box 14, Wageningen, The Netherlands (Received 16 March 1987) A strictly linear relation was established between the carbon and ash contents of plant biomass and the relative amount of substrate required to produce this biomass. Data concerned storage organs of crops of widely different biochemical composition. Amounts of substrate required were calculated by an analysis of the metabolic pathways for the synthesis of the principal components of biomass. Carbon contents were derived from the biomass composition. A linear relation was also established between biomass carbon and ash contents and the CO2 loss during biosynthesis. The linearity of both relations and the ease with which the carbon and ash contents of new biomass samples can be determined accurately imply that the previous procedure for determining the conversion efficiency for plant biomass and growth respiration can be simplified substantially with a gain in accuracy.
Introduction The efficiency of conversion of carbohydrates, the prime products of photosynthesis, into other biochemical components is of central importance when growth and carbon balance of plants are considered. Calculation of the efficiency of growth processes requires knowledge of the quantities of substrate used for the supply of carbon skeletons, energy and reducing power during the production of biomass. Penning de Vries et al. (1974) calculated the amount of substrate required from a detailed analysis of the biochemical reactions involved in the formation of several biomass components. In their approach, the biomass is characterized in a number of typical constituents that can be analyzed with standard methods of plant analysis, viz. carbohydrates, proteins, lipids, lignin, organic anions and minerals. For each of these groups of compounds the conversion efficiency was calculated by careful analysis of the biochemical pathways followed during biosynthesis. The conversion efficiency was quantified by the production value, PV, i.e. the weight of the end product formed from one kilogram of glucose. The quantity of carbon dioxide evolved during the reactions was calculated simultaneously. These calculations referred strictly to processes within the cell. Actual growth involves also translocation of glucose molecules and active uptake into growing cells, for which probably an additional 5% of substrate glucose is required. This procedure to determine the conversion efficiency proved to yield reliable results in several experiments and studies (Penning de "Cries et al., 1983). It was concluded that biochemical conversions in higher plants proceed with nearly maximum thermodynamic efficiency (Penning de Vries et al., 1974). 109 0022-5193/87/180109+ 11 $03.00/0
© 1987 Academic Press Ltd
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However, it is difficult to use this approach in practice with full accuracy because the analytical efforts required to obtain a complete picture of the plant composition are considerable. Some components that are comparable with respect to their molecular composition can require totally different methods for their determination: starch and cellulose are one example, fats and waxes are another. No standard methods are even available for quantitatively measuring lignin precursors. And finally, the carbohydrate fraction is usually not measured directly but set equal to the complement of the sum of the results of the analysis of the other five main groups of biomass components. The carbohydrate fraction includes soluble carbohydrates, starch and cell wall constituents. As a consequence, the theoretically accurate results for substrate requirements are not obtained in practice due to limitations in analytical efforts. In this paper we present a simple and quick method to determine the efficiency of the conversion processes in plants. Its results in practice are at least as good as those obtained with a full biochemical analysis. We expect it to be applicable to all growth processes in higher plants where glucose is the substrate provided. It is assumed that energetically inefficient biochemical pathways are not used.
Elemental Composition and Energy Content of Biomass McDermitt & Loomis (198t) introduced a computation method for the energy content ofbiomass based on its elemental composition. With it, the average oxidation level of carbon in the biomass is determined from the carbon, hydrogen, nitrogen, oxygen and sulfur content in a plant sample. Starting from the carbon content and its oxidation level, the quantity of glucose is calculated that is used for the building of the carbon skeletons and for the reduction of the biomass components to the oxidation level mentioned. This quantity was called the glucose value, GV, and is expressed in g biomass per g glucose. They showed that the glucose value is highly correlated with the production value discussed earlier: for a number of seed crops the PV/GV ratio appeared to be 0.88±0-01. The results of the two calculation methods are not identical because the glucose value measures exclusively the energy content of the biomass produced, while the production value includes the amount of glucose used for the supply of energy during the formation of biomass. This additional requirement appeared to be amazingly constant. The computation of the energy content of biomass according to McDermitt & Loomis (1981) requires an accurate analysis of the carbon, hydrogen, nitrogen, oxygen and sulfur content of the sample material. Because the mineral content of the biomass can be determined only approximately from the ash content, the oxygen content has to be determined directly and cannot be calculated as a rest fraction after determination of carbon, hydrogen and nitrogen. An accurate determination of the full atomic composition of plant samples, however, cannot be performed in most laboratories, which severely limits the applicability of this method. Their line of thought, can be taken one step further.
EFFICIENCY OF PLANT BIOMASS BIOSYNTHESIS
111
Carbon Content of Organic Matter and Efficiency of Biosynthesis An increase in the energy content of the biomass corresponds with a decrease of the oxidation level of its carbon, and is reflected in an increase in carbon content. A relation between carbon content and energy content of the biomass, and also with the production value and glucose value, can therefore be expected. The question arose whether the estimation of the production value based on the biomass carbon content is sufficiently accurate for application in crop growth models. The carbon content of biomass can be measured with high accuracy with commercial C-analysers. Such a relation is investigated here by comparing sets of values for the carbon content, the production value and the glucose value of the storage organs of 23 major crops. Data on the chemical composition and the computed production values are given by Penning de Vries et al. (1983, Table 3). The tabulated crops are very diverse and cover a wide range of possible plant biomass compositions. The carbon content and the glucose value are calculated from the molecular composition for those crops (ibid., Table 2). The molecular formulae of these components are given in Table 1.
TABLE 1
Composition of organic plant components Component Carbohydrates Proteins Lipids Lignin Organic anions
Model compound starch, cellulose zein glyceroltrioleate coniferyl alcohol malate : citrate = 1 : 1
Molecular formula C6H~005 C4.sH6,TNHO1.6 C57HloaO 6 CtoHI203
C5H706
In the following, the reciprocals of the original definitions of glucose value and production value are used, because they are easier in growth equations. These inverses represent the quantities of glucose required by biosynthesis of 1 kg product, and are indicated as GVI and PVI respectively. The reciprocal production value of the organic matter is computed by means of the factors given Penning de Vries et al. (1983, Fig. 2) with exception of those for protein. Proteins are here assumed to be formed from glucose and NH~', with amides as the translocated intermediate metabolites. For the production of 1.000 g protein are required 0.948 g glucose and 0.768 g amides (for 65% of glutamine and 35% asparagine). The amount of glucose required for the synthesis of the amides is derived from data on biochemical pathways as given by Penning de Vries et al. (1974). It follows that 1.100 g glucose is used for the biosynthesis of 1-000 g amides. As a consequence the production of 1.000 g protein requires 0-948+ 1.10 x 0-768 = 1-793 g glucose. The complete equation for the computation of the reciprocal
112
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production value is: PVI = 1.211 * carbohydrates + 1.793 * protein +3.030 * lipids+ 2-119 * iignin + 0.906 * organic anions.
(1)
The reciprocal glucose value is computed by means of factors derived according to the method of McDermitt & Loomis (1981) from the molecular formulae given in Table 1, using eqn (2) GVI = 1.111 * carbohydrates + 1.363 * protein +2.714 * lipids + 1.916 * lignin + 0.688 * organic anions.
(2)
Nitrogen is again considered to be available as NH~-. The carbon content (C, g kg -~) in the organic material is computed from the molecular formulae given in Table 1 according to the equation: C = 0.444 * carbohydrates + 0.535 * protein + 0.774 * lipids + 0.667 * lignin + 0.370 * organic anions.
(3)
The inorganic material does not require glucose for synthesis and contains no carbon. Carbonates are considered to be part of the organic anion fraction. The values for C, PVI, GVI, for the concomittant CO2 evolution and for all concentrations are expressed on a dry matter basis, and are all expressed in g kg -1. The factors used in these equations are expected to be generally applicable for computations with plant material, deviations in composition within the categories of chemical components and between species being comparatively small and of little importance. The results of the computations of the carbon content and o f the reciprocal production value and glucose value o f the different storage organs are presented in Table 2. The relation between carbon content and energy content o f the biomass holds only for organic components. As the mineral contents o f different plants and organs are unequal, the analytical data have to be corrected for the mineral content. The data in Table 2 were recalculated on a mineral-free organic matter basis of yield GVIom and PVlom prior to the regression analysis on carbon content and production value. The results of these recalculations are given in Fig. 1. PVIom and GVIom appear to be strongly correlated with the carbon content of the organic fraction of the biomass, according to the following regression equations: PVIom = 5.39 * Co~ - 1191,
r = 0-997,
SEE = 20-9
(4)
GVIom = 4"63 * Corn- 988,
r = 0"993,
SEE = 21"5
(5)
Cassava, tuber (Manihot esculenta) Chickpea, pod with seed (Cicer arietinum) Coconut palm, coconut ' (Cocos nucifera) cotton, boll (Gossypium sp.) Cowpea, pod with seed (Vigna unguiculata) Field bean, pod with seed (Vicia faba) Field bean, pod with seed (Phaseolus vulgaris) Groundnut, pod with seed (Arachis hypogaea) Maize, cob (Zea mays) Miller, ear (Pennisetum typhoides) Oil palm, palm nut (Elaeis guineensis) Pigeonpea, pod with seed (Cajanus cajan)
30 190 40 210 220 290 230 270 80 90 70 200
650 390 400 610 550 600 140 750 690 370 600
protein
870
carbohydrate
20
480
40
40
390
20
10
20
230
280
60
10
lipid
100
40
120
110
140
70
70
70
80
250
40
30
lignin
40
20
30
10
30
40
40
40
40
20
30
30
organic anion
Composition of storage organ g kg -I dry matter
40
20
30
10
30
40
40
40
40
20
30
30
470
607
477
484
613
466
469
466
536
585
474
441
minerals carbon
1394
2t31
1400
1415
2159
1384
1401
1378
1764
1940
1422
1228
PVI
1213
1900
1248
1268
1871
1196
1195
1193
1536
1740
1241
1113
GVI
321
900
304
301
919
321
335
312
622
700
348
184
CPFI
glucose required and carbon dioxide produced during biosynthesis of biomass, g kg -I dry matter
Composition (columns 1-6), carbon contents and values characterizing the cost of biosynthesis of storage organs of the major crops; storage organ consists of economic product plus hull, pod or inflorescence
TABLE 2
.<
Z
7.
t-n "n
Potato, tuber (Solanum tubesum) Rice, inflorescence with seed (Oryza sativa) Sorghum, inflorescence with seed (Sorghum bicolor) Soybean, pod with seed (Glycine max) Sugar beet, beet (Beta vulgaris) Sugarcane, whole tops (Saccharum sp.) Sunflower, inflorescence with seed (Helianthus annuus) Sweet potato, tuber (Ipomoea batatus) Tomato, fruit (Lycopersicum esculentum) Wheat, inflorescence with seed (Triticum sp.) Yam,, tuber (Dioscorea sp.)
90 80 90 370 50 70 140 50 170 120 60
760 720 290 820 570 450 840 540 760 800
protein
780
carbohydrate
10
20
40
20
220
20
0
180
30
20
0
lipid
30
60
90
30
130
220
50
60
120
120
30
lignin
50
20
80
30
30
60
40
50
20
10
50
organic anion
Composition of storage organ gkg -~ dry matter
TABLE 2--continued
50
20
80
30
30
60
40
50
20
10
50
434
465
451
446
543
475
439
525
478
479
433
minerals carbon
1216
1341
1343
1258
1765
1397
1225
1732
1397
1388
1215
PVI
1090
1191
1168
1134
1558
1246
1102
1464
1248
1244
1081
GVI
192
262
316
210
598
307
187
615
296
279
194
CPF1
glucose required and carbon dioxide produced during biosynthesis of biomass, g kg -t dry matter
m <
Z 0
I'n Z Z
-t
> Z
O .4
Z < m 7¢
4~
EFFICIENCY
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PLANT
BIOSYNTHESIS
BIOMASS
115
2400
/
2200
~ ~ 2ooo 8 e
o
1800
cZ
o ~ "o o _
1600
o
o o I=n
8o ~
°dr/
i400
,,,,
1200
I000 400
I
I
I
I
450
500
55O
600
650
C a r b o n c o n l e n t {g k g - I o r g a n m a t t e r )
Fl~. 1. The relation between the reciprocal production value (PVI) and the carbon content of the biomass grown, for the storage organs o f 23 crops indicated in Table 2. The reciprocal glucose value (GVI) represents the energy retained in the biomass.
(Corn in g per kg, PVIom and GVIom in g glucose per kg of the organic fraction of the biomass). The reciprocal production value of the organic matter can be estimated quite accurately from the carbon content of the organic fraction of the plant material, as is shown by the low value for the standard error of estimate, SEE. The calculations of the reciprocal glucose values are shown for comparison only. Its values represent the heat of combustion of organic components when the ordinate units are multiplied by the heat of combustion of glucose, i.e. 1565 J g-i. The total bond energy of organic compounds cannot be calculated by the summation of contributions of its carbon, hydrogen, nitrogen and oxygen. The bond energy depends also on the molecular structure of the compounds. This is shown for a series of natural and synthetic organic compounds with carbon contents between 375 (citric acid) and 923 (benzene) g kg-L The corresponding reciprocal glucose value is calculated using the method of McDermitt & Loomis. The regression equation of the carbon contents and the reciprocal glucose values appears to be GVI = 3.781 * C - 5 1 4 ,
r = 0.975,
SEE = 225.
(6)
The accuracy of the prediction of the reciprocal glucose value for these compounds is not very high. The good result of the method introduced here (eqn (5)) was therefore rather unexpected. The precise prediction of the reciprocal production value from the carbon content of storage organs must be a consequence of the relatively uniform chemical composition of these materials, and of the high correlation between the production value and the carbon content of the major components
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(carbohydrates, proteins, lipids and organic anions) as is shown below. The deviating values of aromatic compounds like lignin are of little importance for annual crops as these usually contain rather small fractions of these constituents. The coefficient of variance for the determination of the carbon content (in plant samples by a combustion method) is as low as 1.5%. The standard error of estimate for the prediction of the reciprocal production value of biomass from the carbon content is 1-5% or less (see eqn (4)). The precision of the estimation of the reciprocal production value is therefore about 2%. This precision cannot be obtained by the long procedure involving chemical analysis of biomass components due to lack of analytical precision and insufficiently defined molecular composition of the analyzed compounds. The short method starting from the carbon content is therefore in practice a better and simpler method to estimate the reciprocal production value than the full chemical analysis. With the good correlation between the carbon content of the organic fraction of the sample and the production value, it seems worthwhile to attempt to generalize this pattern. Figure 2 shows that the pure biochemical fractions fall clearly on the line calculated for PVIom and GVIom in Fig. 1, in spite of the fact that the range is almost twice as large. Only the component lignin deviates from the calculated line.
5000 "E ~o
g "T
2OOO
~, "6
Iooo
g
o 4(30
@
800
Carbon content (g kg-I organic matter)
FIG. 2. The relation between the reciprocal production value (PVI; open dots) a n d o f the reciprocal glucose value (GVI; closed dots) and the carbon content o f the biomass (from Fig. 1). The line labeled C corresponds with the a m o u n t o f glucose required to provide the carbon for the new biomass only. The difference between the PVI a n d the C-line equals the a m o u n t of carbon lost as CO2 in the biosynthesis a n d is represented by the line CPFI. Numerals refer to the pure components: 1 = c a r b o h y d r a t e s , 2 = proteins, 3 = lipids, 4 = lignin, 5 = organic acids, 6 = glucose and 7 = COz (open dot only).
EFFICIENCY
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BIOMASS
BIOSYNTHESIS
117
As a consequence, the PVI of plant material with a very high lignin content of 30% is about 3% overestimated. A third line is added in Fig. 2 to represent the amount of glucose needed to provide only the carbon in the end product. This is a straight line (labelled C) through the origin and the point of 400 g carbon per kg glucose. This carbon line appears to cross the PVI line. At the right hand side of this point is excess glucose used to provide energy for the growth process. The difference between both lines represents the amount of CO2 that is lost during biosynthesis. At the left hand side of this point the substrate provides less energy then is contained in the product. The actual substrate requirement follows the highest of both lines: the positions in the figure of organic acids and of CO, (as an end product) confirm this. In practice, however, this deviation from the straight regression line labelled PVI can be neglected: the carbon content of the biomass is almost always higher than 400 g kg -~ and the energy derived from glucose is redistributed over components of relative higher and lower reduction levels. A fourth line is drawn to reflect the difference between the amount of carbon in the substrate and the amount retained in the product. It indicates the CO2 production during biosynthesis, and may be called CPFI (g CO2 kg -~ substrate). Its value can be determined with eqn (7) CPFIom = ~ * (PVIom * 0 . 4 0 - Corn).
(7)
It can also be calculated directly with eqn (8), derived from Table 2, last column CPFIom = 4.24 *Com- 1744,
r = 0-991,
SEE = 30"8.
(8)
Carbon Content of Biomass and Efficiency of Biosynthesis As only organic components of the biomass are taken into account in the calculation of the reciprocal production value, and because the regression line does not cross the origin, the regression equation is valid only for computations on a mineral free basis. For recalculation on a dry matter basis attention must be given to the following. The salt fraction of plants is composed of inorganic cations, (K ÷, Mg ÷÷, Ca+÷), inorganic anions (H2PO~, NO~, C1-) and organic anions like citrate and malate. As a rule of thumb, the weight of the inorganic salt constituents equals the weight of the organic anions. It is impractical to analyse for the individual ions. The analysis is therefore usually restricted to the determination of the ash content. During ashing at 550°C, organic anions and nitrate, with equivalent weights of about 60, are converted into carbonate with an equivalent weight of 30. As a consequence the inorganic mineral content and the organic anion content can both be estimated as being 67% of the ash content. This approximation is valid for storage organs with a salt content of about 60 g kg -~ dry matter and for leaf material with a salt content of about 130 g kg -~ dry matter. In organs with a high content of silicate such as rice leaves, with up to 20% SIO2, or improperly cleaned roots, the multiplication factor is higher than 0.67, as SiO2 is not modified during ashing. A separate SiO2
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determination can be useful if such high contents are expected. For SiO2-free samples the following transformations can be used to express analytical results on a dry matter basis into values for organic matter PVIom = PVIdm * 1000/(1000-0.67 aShdm)
(9)
Cor~ = Cdm * 1000/(1000-0"67 ashdm).
(10)
Values are in g per kg of organic matter (om) and dry matter (dm) respectively. Substitution of (9) in (4) gives PVIdm = 5"39 Calm+0"80 aShdm- 1191.
(11)
The amount of carbon dioxide produced during the synthesis of 1 kg biomass can be calculated by CPFIam = ~ (PVIa~ * 0 - 4 - Cdm)
(12)
or directly from the carbon and ash content by substitution of (10) in (8) CPFIdm =4.24 Cam+ 1"17 ashom- 1744.
(13)
Cost of Other Processes
Cost of processes other than biosynthesis, such as translocation within the plant or N, reduction in legumes, are not included in this analysis and calculation of the reciprocal production value. For the first process, costs are estimated at 2 ATP per glucose molecule for active passage of two membranes (Penning de Vries et aL, 1983), so that 5.3% must be added to the values calculated by eqns (4) and (11) if these costs are included in the study, as is customary for growth processes. All of this additional glucose is lost as CO2. The expense of N2 reduction in nodules of leguminous crops is estimated at 4500 g glucose per kg N; also this glucose is combusted completely. The cost of nitrate reduction is similar to that of N2-reduction, but that energy is probably often supplied free in the photosynthesis process and not by the combustion of glucose. Conclusion
The quantity of substrate glucose required for the synthesis of plant biomass can be calculated with high .precision from the carbon and ash content of the biomass, using eqn (4) or (11). These contents can be measured easily and accurately. The heat of combustion of the biomass can be estimated with comparable high precision by multiplication of the amount of glucose, derived with eqns (5) and (10), with the heat of combustion of glucose. A relatively simple chemical measurement is therefore sufficient for an accurate determination of the substrate requirement of biosynthesis. REFERENCES MCDERMI'r'r, D. K. & LOOMIS,R. S. (1981). Elemental composition of biomass and its relation to energy content, growthefficiencyand growth yield. Ann. Bot. 48, 275
EFFICIENCY
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119
PENNING DE VRIES, F. W. T., BRUNSTING, A. H. M. & VAN LAAR, H. H. (1974). Products, requirements and efficiency of biosynthesis: a quantitative approach. J. theor. Biol. 45, 339. PENNING DE VRIES, F. W. T., VAN LAAR, H. H. & CHARDON, M. C. M. (1983). flioenergetics of growth of seeds, fruits and storage organs. In: Proceedings Symposium Potential Productivity of Field Crops under Different Environments. 22-26 September 1980. pp. 37-59. Los Banos: International Rice Research Institute.
A Rapid Method for Determining the Efficiency of Biosynthesis of Plant Biomass N. VERTREGT AND F. W. T. PENNING DE VRIES
Centre of Agrobiological Research, P.O. Box 14, Wageningen, The Netherlands (Received 16 March 1987) A strictly linear relation was established between the carbon and ash contents of plant biomass and the relative amount of substrate required to produce this biomass. Data concerned storage organs of crops of widely different biochemical composition. Amounts of substrate required were calculated by an analysis of the metabolic pathways for the synthesis of the principal components of biomass. Carbon contents were derived from the biomass composition. A linear relation was also established between biomass carbon and ash contents and the CO2 loss during biosynthesis. The linearity of both relations and the ease with which the carbon and ash contents of new biomass samples can be determined accurately imply that the previous procedure for determining the conversion efficiency for plant biomass and growth respiration can be simplified substantially with a gain in accuracy.
Introduction The efficiency of conversion of carbohydrates, the prime products of photosynthesis, into other biochemical components is of central importance when growth and carbon balance of plants are considered. Calculation of the efficiency of growth processes requires knowledge of the quantities of substrate used for the supply of carbon skeletons, energy and reducing power during the production of biomass. Penning de Vries et al. (1974) calculated the amount of substrate required from a detailed analysis of the biochemical reactions involved in the formation of several biomass components. In their approach, the biomass is characterized in a number of typical constituents that can be analyzed with standard methods of plant analysis, viz. carbohydrates, proteins, lipids, lignin, organic anions and minerals. For each of these groups of compounds the conversion efficiency was calculated by careful analysis of the biochemical pathways followed during biosynthesis. The conversion efficiency was quantified by the production value, PV, i.e. the weight of the end product formed from one kilogram of glucose. The quantity of carbon dioxide evolved during the reactions was calculated simultaneously. These calculations referred strictly to processes within the cell. Actual growth involves also translocation of glucose molecules and active uptake into growing cells, for which probably an additional 5% of substrate glucose is required. This procedure to determine the conversion efficiency proved to yield reliable results in several experiments and studies (Penning de "Cries et al., 1983). It was concluded that biochemical conversions in higher plants proceed with nearly maximum thermodynamic efficiency (Penning de Vries et al., 1974). 109 0022-5193/87/180109+ 11 $03.00/0
© 1987 Academic Press Ltd
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N. V E R T R E G T
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F. W . T . P E N N I N G
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VRIES
However, it is difficult to use this approach in practice with full accuracy because the analytical efforts required to obtain a complete picture of the plant composition are considerable. Some components that are comparable with respect to their molecular composition can require totally different methods for their determination: starch and cellulose are one example, fats and waxes are another. No standard methods are even available for quantitatively measuring lignin precursors. And finally, the carbohydrate fraction is usually not measured directly but set equal to the complement of the sum of the results of the analysis of the other five main groups of biomass components. The carbohydrate fraction includes soluble carbohydrates, starch and cell wall constituents. As a consequence, the theoretically accurate results for substrate requirements are not obtained in practice due to limitations in analytical efforts. In this paper we present a simple and quick method to determine the efficiency of the conversion processes in plants. Its results in practice are at least as good as those obtained with a full biochemical analysis. We expect it to be applicable to all growth processes in higher plants where glucose is the substrate provided. It is assumed that energetically inefficient biochemical pathways are not used.
Elemental Composition and Energy Content of Biomass McDermitt & Loomis (198t) introduced a computation method for the energy content ofbiomass based on its elemental composition. With it, the average oxidation level of carbon in the biomass is determined from the carbon, hydrogen, nitrogen, oxygen and sulfur content in a plant sample. Starting from the carbon content and its oxidation level, the quantity of glucose is calculated that is used for the building of the carbon skeletons and for the reduction of the biomass components to the oxidation level mentioned. This quantity was called the glucose value, GV, and is expressed in g biomass per g glucose. They showed that the glucose value is highly correlated with the production value discussed earlier: for a number of seed crops the PV/GV ratio appeared to be 0.88±0-01. The results of the two calculation methods are not identical because the glucose value measures exclusively the energy content of the biomass produced, while the production value includes the amount of glucose used for the supply of energy during the formation of biomass. This additional requirement appeared to be amazingly constant. The computation of the energy content of biomass according to McDermitt & Loomis (1981) requires an accurate analysis of the carbon, hydrogen, nitrogen, oxygen and sulfur content of the sample material. Because the mineral content of the biomass can be determined only approximately from the ash content, the oxygen content has to be determined directly and cannot be calculated as a rest fraction after determination of carbon, hydrogen and nitrogen. An accurate determination of the full atomic composition of plant samples, however, cannot be performed in most laboratories, which severely limits the applicability of this method. Their line of thought, can be taken one step further.
EFFICIENCY OF PLANT BIOMASS BIOSYNTHESIS
111
Carbon Content of Organic Matter and Efficiency of Biosynthesis An increase in the energy content of the biomass corresponds with a decrease of the oxidation level of its carbon, and is reflected in an increase in carbon content. A relation between carbon content and energy content of the biomass, and also with the production value and glucose value, can therefore be expected. The question arose whether the estimation of the production value based on the biomass carbon content is sufficiently accurate for application in crop growth models. The carbon content of biomass can be measured with high accuracy with commercial C-analysers. Such a relation is investigated here by comparing sets of values for the carbon content, the production value and the glucose value of the storage organs of 23 major crops. Data on the chemical composition and the computed production values are given by Penning de Vries et al. (1983, Table 3). The tabulated crops are very diverse and cover a wide range of possible plant biomass compositions. The carbon content and the glucose value are calculated from the molecular composition for those crops (ibid., Table 2). The molecular formulae of these components are given in Table 1.
TABLE 1
Composition of organic plant components Component Carbohydrates Proteins Lipids Lignin Organic anions
Model compound starch, cellulose zein glyceroltrioleate coniferyl alcohol malate : citrate = 1 : 1
Molecular formula C6H~005 C4.sH6,TNHO1.6 C57HloaO 6 CtoHI203
C5H706
In the following, the reciprocals of the original definitions of glucose value and production value are used, because they are easier in growth equations. These inverses represent the quantities of glucose required by biosynthesis of 1 kg product, and are indicated as GVI and PVI respectively. The reciprocal production value of the organic matter is computed by means of the factors given Penning de Vries et al. (1983, Fig. 2) with exception of those for protein. Proteins are here assumed to be formed from glucose and NH~', with amides as the translocated intermediate metabolites. For the production of 1.000 g protein are required 0.948 g glucose and 0.768 g amides (for 65% of glutamine and 35% asparagine). The amount of glucose required for the synthesis of the amides is derived from data on biochemical pathways as given by Penning de Vries et al. (1974). It follows that 1.100 g glucose is used for the biosynthesis of 1-000 g amides. As a consequence the production of 1.000 g protein requires 0-948+ 1.10 x 0-768 = 1-793 g glucose. The complete equation for the computation of the reciprocal
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production value is: PVI = 1.211 * carbohydrates + 1.793 * protein +3.030 * lipids+ 2-119 * iignin + 0.906 * organic anions.
(1)
The reciprocal glucose value is computed by means of factors derived according to the method of McDermitt & Loomis (1981) from the molecular formulae given in Table 1, using eqn (2) GVI = 1.111 * carbohydrates + 1.363 * protein +2.714 * lipids + 1.916 * lignin + 0.688 * organic anions.
(2)
Nitrogen is again considered to be available as NH~-. The carbon content (C, g kg -~) in the organic material is computed from the molecular formulae given in Table 1 according to the equation: C = 0.444 * carbohydrates + 0.535 * protein + 0.774 * lipids + 0.667 * lignin + 0.370 * organic anions.
(3)
The inorganic material does not require glucose for synthesis and contains no carbon. Carbonates are considered to be part of the organic anion fraction. The values for C, PVI, GVI, for the concomittant CO2 evolution and for all concentrations are expressed on a dry matter basis, and are all expressed in g kg -1. The factors used in these equations are expected to be generally applicable for computations with plant material, deviations in composition within the categories of chemical components and between species being comparatively small and of little importance. The results of the computations of the carbon content and o f the reciprocal production value and glucose value o f the different storage organs are presented in Table 2. The relation between carbon content and energy content o f the biomass holds only for organic components. As the mineral contents o f different plants and organs are unequal, the analytical data have to be corrected for the mineral content. The data in Table 2 were recalculated on a mineral-free organic matter basis of yield GVIom and PVlom prior to the regression analysis on carbon content and production value. The results of these recalculations are given in Fig. 1. PVIom and GVIom appear to be strongly correlated with the carbon content of the organic fraction of the biomass, according to the following regression equations: PVIom = 5.39 * Co~ - 1191,
r = 0-997,
SEE = 20-9
(4)
GVIom = 4"63 * Corn- 988,
r = 0"993,
SEE = 21"5
(5)
Cassava, tuber (Manihot esculenta) Chickpea, pod with seed (Cicer arietinum) Coconut palm, coconut ' (Cocos nucifera) cotton, boll (Gossypium sp.) Cowpea, pod with seed (Vigna unguiculata) Field bean, pod with seed (Vicia faba) Field bean, pod with seed (Phaseolus vulgaris) Groundnut, pod with seed (Arachis hypogaea) Maize, cob (Zea mays) Miller, ear (Pennisetum typhoides) Oil palm, palm nut (Elaeis guineensis) Pigeonpea, pod with seed (Cajanus cajan)
30 190 40 210 220 290 230 270 80 90 70 200
650 390 400 610 550 600 140 750 690 370 600
protein
870
carbohydrate
20
480
40
40
390
20
10
20
230
280
60
10
lipid
100
40
120
110
140
70
70
70
80
250
40
30
lignin
40
20
30
10
30
40
40
40
40
20
30
30
organic anion
Composition of storage organ g kg -I dry matter
40
20
30
10
30
40
40
40
40
20
30
30
470
607
477
484
613
466
469
466
536
585
474
441
minerals carbon
1394
2t31
1400
1415
2159
1384
1401
1378
1764
1940
1422
1228
PVI
1213
1900
1248
1268
1871
1196
1195
1193
1536
1740
1241
1113
GVI
321
900
304
301
919
321
335
312
622
700
348
184
CPFI
glucose required and carbon dioxide produced during biosynthesis of biomass, g kg -I dry matter
Composition (columns 1-6), carbon contents and values characterizing the cost of biosynthesis of storage organs of the major crops; storage organ consists of economic product plus hull, pod or inflorescence
TABLE 2
.<
Z
7.
t-n "n
Potato, tuber (Solanum tubesum) Rice, inflorescence with seed (Oryza sativa) Sorghum, inflorescence with seed (Sorghum bicolor) Soybean, pod with seed (Glycine max) Sugar beet, beet (Beta vulgaris) Sugarcane, whole tops (Saccharum sp.) Sunflower, inflorescence with seed (Helianthus annuus) Sweet potato, tuber (Ipomoea batatus) Tomato, fruit (Lycopersicum esculentum) Wheat, inflorescence with seed (Triticum sp.) Yam,, tuber (Dioscorea sp.)
90 80 90 370 50 70 140 50 170 120 60
760 720 290 820 570 450 840 540 760 800
protein
780
carbohydrate
10
20
40
20
220
20
0
180
30
20
0
lipid
30
60
90
30
130
220
50
60
120
120
30
lignin
50
20
80
30
30
60
40
50
20
10
50
organic anion
Composition of storage organ gkg -~ dry matter
TABLE 2--continued
50
20
80
30
30
60
40
50
20
10
50
434
465
451
446
543
475
439
525
478
479
433
minerals carbon
1216
1341
1343
1258
1765
1397
1225
1732
1397
1388
1215
PVI
1090
1191
1168
1134
1558
1246
1102
1464
1248
1244
1081
GVI
192
262
316
210
598
307
187
615
296
279
194
CPF1
glucose required and carbon dioxide produced during biosynthesis of biomass, g kg -t dry matter
m <
Z 0
I'n Z Z
-t
> Z
O .4
Z < m 7¢
4~
EFFICIENCY
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PLANT
BIOSYNTHESIS
BIOMASS
115
2400
/
2200
~ ~ 2ooo 8 e
o
1800
cZ
o ~ "o o _
1600
o
o o I=n
8o ~
°dr/
i400
,,,,
1200
I000 400
I
I
I
I
450
500
55O
600
650
C a r b o n c o n l e n t {g k g - I o r g a n m a t t e r )
Fl~. 1. The relation between the reciprocal production value (PVI) and the carbon content of the biomass grown, for the storage organs o f 23 crops indicated in Table 2. The reciprocal glucose value (GVI) represents the energy retained in the biomass.
(Corn in g per kg, PVIom and GVIom in g glucose per kg of the organic fraction of the biomass). The reciprocal production value of the organic matter can be estimated quite accurately from the carbon content of the organic fraction of the plant material, as is shown by the low value for the standard error of estimate, SEE. The calculations of the reciprocal glucose values are shown for comparison only. Its values represent the heat of combustion of organic components when the ordinate units are multiplied by the heat of combustion of glucose, i.e. 1565 J g-i. The total bond energy of organic compounds cannot be calculated by the summation of contributions of its carbon, hydrogen, nitrogen and oxygen. The bond energy depends also on the molecular structure of the compounds. This is shown for a series of natural and synthetic organic compounds with carbon contents between 375 (citric acid) and 923 (benzene) g kg-L The corresponding reciprocal glucose value is calculated using the method of McDermitt & Loomis. The regression equation of the carbon contents and the reciprocal glucose values appears to be GVI = 3.781 * C - 5 1 4 ,
r = 0.975,
SEE = 225.
(6)
The accuracy of the prediction of the reciprocal glucose value for these compounds is not very high. The good result of the method introduced here (eqn (5)) was therefore rather unexpected. The precise prediction of the reciprocal production value from the carbon content of storage organs must be a consequence of the relatively uniform chemical composition of these materials, and of the high correlation between the production value and the carbon content of the major components
116
N. V E R T R E G T
AND
F. W. T. P E N N I N G
DE VRIES
(carbohydrates, proteins, lipids and organic anions) as is shown below. The deviating values of aromatic compounds like lignin are of little importance for annual crops as these usually contain rather small fractions of these constituents. The coefficient of variance for the determination of the carbon content (in plant samples by a combustion method) is as low as 1.5%. The standard error of estimate for the prediction of the reciprocal production value of biomass from the carbon content is 1-5% or less (see eqn (4)). The precision of the estimation of the reciprocal production value is therefore about 2%. This precision cannot be obtained by the long procedure involving chemical analysis of biomass components due to lack of analytical precision and insufficiently defined molecular composition of the analyzed compounds. The short method starting from the carbon content is therefore in practice a better and simpler method to estimate the reciprocal production value than the full chemical analysis. With the good correlation between the carbon content of the organic fraction of the sample and the production value, it seems worthwhile to attempt to generalize this pattern. Figure 2 shows that the pure biochemical fractions fall clearly on the line calculated for PVIom and GVIom in Fig. 1, in spite of the fact that the range is almost twice as large. Only the component lignin deviates from the calculated line.
5000 "E ~o
g "T
2OOO
~, "6
Iooo
g
o 4(30
@
800
Carbon content (g kg-I organic matter)
FIG. 2. The relation between the reciprocal production value (PVI; open dots) a n d o f the reciprocal glucose value (GVI; closed dots) and the carbon content o f the biomass (from Fig. 1). The line labeled C corresponds with the a m o u n t o f glucose required to provide the carbon for the new biomass only. The difference between the PVI a n d the C-line equals the a m o u n t of carbon lost as CO2 in the biosynthesis a n d is represented by the line CPFI. Numerals refer to the pure components: 1 = c a r b o h y d r a t e s , 2 = proteins, 3 = lipids, 4 = lignin, 5 = organic acids, 6 = glucose and 7 = COz (open dot only).
EFFICIENCY
OF PLANT
BIOMASS
BIOSYNTHESIS
117
As a consequence, the PVI of plant material with a very high lignin content of 30% is about 3% overestimated. A third line is added in Fig. 2 to represent the amount of glucose needed to provide only the carbon in the end product. This is a straight line (labelled C) through the origin and the point of 400 g carbon per kg glucose. This carbon line appears to cross the PVI line. At the right hand side of this point is excess glucose used to provide energy for the growth process. The difference between both lines represents the amount of CO2 that is lost during biosynthesis. At the left hand side of this point the substrate provides less energy then is contained in the product. The actual substrate requirement follows the highest of both lines: the positions in the figure of organic acids and of CO, (as an end product) confirm this. In practice, however, this deviation from the straight regression line labelled PVI can be neglected: the carbon content of the biomass is almost always higher than 400 g kg -~ and the energy derived from glucose is redistributed over components of relative higher and lower reduction levels. A fourth line is drawn to reflect the difference between the amount of carbon in the substrate and the amount retained in the product. It indicates the CO2 production during biosynthesis, and may be called CPFI (g CO2 kg -~ substrate). Its value can be determined with eqn (7) CPFIom = ~ * (PVIom * 0 . 4 0 - Corn).
(7)
It can also be calculated directly with eqn (8), derived from Table 2, last column CPFIom = 4.24 *Com- 1744,
r = 0-991,
SEE = 30"8.
(8)
Carbon Content of Biomass and Efficiency of Biosynthesis As only organic components of the biomass are taken into account in the calculation of the reciprocal production value, and because the regression line does not cross the origin, the regression equation is valid only for computations on a mineral free basis. For recalculation on a dry matter basis attention must be given to the following. The salt fraction of plants is composed of inorganic cations, (K ÷, Mg ÷÷, Ca+÷), inorganic anions (H2PO~, NO~, C1-) and organic anions like citrate and malate. As a rule of thumb, the weight of the inorganic salt constituents equals the weight of the organic anions. It is impractical to analyse for the individual ions. The analysis is therefore usually restricted to the determination of the ash content. During ashing at 550°C, organic anions and nitrate, with equivalent weights of about 60, are converted into carbonate with an equivalent weight of 30. As a consequence the inorganic mineral content and the organic anion content can both be estimated as being 67% of the ash content. This approximation is valid for storage organs with a salt content of about 60 g kg -~ dry matter and for leaf material with a salt content of about 130 g kg -~ dry matter. In organs with a high content of silicate such as rice leaves, with up to 20% SIO2, or improperly cleaned roots, the multiplication factor is higher than 0.67, as SiO2 is not modified during ashing. A separate SiO2
118
N. VERTREGT
AND
F. W . T . P E N N I N G
DE
VRIES
determination can be useful if such high contents are expected. For SiO2-free samples the following transformations can be used to express analytical results on a dry matter basis into values for organic matter PVIom = PVIdm * 1000/(1000-0.67 aShdm)
(9)
Cor~ = Cdm * 1000/(1000-0"67 ashdm).
(10)
Values are in g per kg of organic matter (om) and dry matter (dm) respectively. Substitution of (9) in (4) gives PVIdm = 5"39 Calm+0"80 aShdm- 1191.
(11)
The amount of carbon dioxide produced during the synthesis of 1 kg biomass can be calculated by CPFIam = ~ (PVIa~ * 0 - 4 - Cdm)
(12)
or directly from the carbon and ash content by substitution of (10) in (8) CPFIdm =4.24 Cam+ 1"17 ashom- 1744.
(13)
Cost of Other Processes
Cost of processes other than biosynthesis, such as translocation within the plant or N, reduction in legumes, are not included in this analysis and calculation of the reciprocal production value. For the first process, costs are estimated at 2 ATP per glucose molecule for active passage of two membranes (Penning de Vries et aL, 1983), so that 5.3% must be added to the values calculated by eqns (4) and (11) if these costs are included in the study, as is customary for growth processes. All of this additional glucose is lost as CO2. The expense of N2 reduction in nodules of leguminous crops is estimated at 4500 g glucose per kg N; also this glucose is combusted completely. The cost of nitrate reduction is similar to that of N2-reduction, but that energy is probably often supplied free in the photosynthesis process and not by the combustion of glucose. Conclusion
The quantity of substrate glucose required for the synthesis of plant biomass can be calculated with high .precision from the carbon and ash content of the biomass, using eqn (4) or (11). These contents can be measured easily and accurately. The heat of combustion of the biomass can be estimated with comparable high precision by multiplication of the amount of glucose, derived with eqns (5) and (10), with the heat of combustion of glucose. A relatively simple chemical measurement is therefore sufficient for an accurate determination of the substrate requirement of biosynthesis. REFERENCES MCDERMI'r'r, D. K. & LOOMIS,R. S. (1981). Elemental composition of biomass and its relation to energy content, growthefficiencyand growth yield. Ann. Bot. 48, 275
EFFICIENCY
OF PLANT B1OMASS BIOSYNTHESIS
119
PENNING DE VRIES, F. W. T., BRUNSTING, A. H. M. & VAN LAAR, H. H. (1974). Products, requirements and efficiency of biosynthesis: a quantitative approach. J. theor. Biol. 45, 339. PENNING DE VRIES, F. W. T., VAN LAAR, H. H. & CHARDON, M. C. M. (1983). flioenergetics of growth of seeds, fruits and storage organs. In: Proceedings Symposium Potential Productivity of Field Crops under Different Environments. 22-26 September 1980. pp. 37-59. Los Banos: International Rice Research Institute.