Meta‐analysis techniques were used to examine the effect of elevated atmospheric carbon dioxide [CO 2 ] on the protein concentrations of major food crops, incorporating 228 experimental observations on barley, rice, wheat, soybean and potato. Each crop had lower protein concentrations when grown at elevated (540–958 μmol mol −1 ) compared with ambient (315–400 μmol mol −1 ) CO 2 . For wheat, barley and rice, the reduction in grain protein concentration was ∼10–15% of the value at ambient CO 2 . For potato, the reduction in tuber protein concentration was 14%. For soybean, there was a much smaller, although statistically significant reduction of protein concentration of 1.4%. The magnitude of the CO 2 effect on wheat grains was smaller under high soil N conditions than under low soil N. Protein concentrations in potato tubers were reduced more for plants grown at high than at low concentrations of ozone. For soybean, the ozone effect was the reverse, as elevated CO 2 increased the protein concentration of soybean grown at high ozone concentrations. The magnitude of the CO 2 effect also varied depending on experimental methodology. For both wheat and soybean, studies performed in open‐top chambers produced a larger CO 2 effect than those performed using other types of experimental facilities. There was also indication of a possible pot artifact as, for both wheat and soybean, studies performed in open‐top chambers showed a significantly greater CO 2 effect when plants were rooted in pots rather than in the ground. Studies on wheat also showed a greater CO 2 effect when protein concentration was measured in whole grains rather than flour. While the magnitude of the effect of elevated CO 2 varied depending on the experimental procedures, a reduction in protein concentration was consistently found for most crops. These findings suggest that the increasing CO 2 concentrations of the 21st century are likely to decrease the protein concentration of many human plant foods.

Introduction Atmospheric concentrations of carbon dioxide (CO 2 ) have been steadily rising from preindustrial values of approximately 280 μmol mol−1 to a current global mean of approximately 380 μmol mol−1 (Keeling & Whorf, 2005; IPCC, 2007). Concentrations are projected to increase to approximately 540–958 μmol mol−1 by the year 2100 (IPCC, 2001). Numerous effects of elevated atmospheric CO 2 concentrations on plants have been documented, including changes in plant elemental composition. As growth CO 2 concentrations increase, plants typically show increased concentrations of carbon in their tissues, with correspondingly reduced concentrations of other elements, including nitrogen (Cotrufo et al., 1998; Gifford et al., 2000), phosphorus (Gifford et al., 2000) and several trace elements (Loladze, 2002). Along with changes in elemental composition, changes have frequently been noted in macromolecular composition, with proteins (which contain substantial amounts of nitrogen and sulfur) decreasing and relatively carbon‐rich molecules such as carbohydrates increasing in concentration at higher concentrations of atmospheric CO 2 (e.g. Poorter et al., 1997). Such changes in plant composition might be expected to have important implications for the growth and nutrition of animals that consume plant material. In a recent meta‐analysis, Zvereva & Kozlov (2006) found that insect herbivore performance was diminished when feeding on plants grown at elevated vs. ambient concentrations of CO 2 . Several authors have also considered the possible implications of altered chemical composition of plants in elevated CO 2 for human nutrition. Loladze (2002) argued that elevated CO 2 may lead to ‘globally imbalanced plant stoichiometry’ and negatively impact human nutrition, particularly with regard to micronutrients such as zinc and iodine. Idso & Idso (2001) in a narrative review, examined a number of studies on the effects of elevated CO 2 on food composition. They found that, for a given nutrient, the results of CO 2 enrichment varied: for example various studies have shown that CO 2 may increase, decrease or have no effect on the protein concentration of crops. In spite of the potential for elevated CO 2 to affect the nutritional composition of foods, there have been few attempts at meta‐analysis or quantitative synthesis of the available data. Loladze (2002) synthesized data from five published studies on wheat grains and found reductions in the concentration of eight elements (including nitrogen) when plants were grown at elevated CO 2 . Jablonski et al. (2002) performed a meta‐analysis of studies on the effects of elevated CO 2 on plant reproductive characteristics, including seed N concentration, for several seed/grain crops. They found that growth at elevated CO 2 resulted in significant decreases in seed N for wheat and barley, but not for soybean or rice. Neither of these studies focused exclusively on the effect of elevated CO 2 on crop nutrient composition and both surveyed only a limited number of crops and a limited selection of the available literature on those crops. In order to rigorously address the question of how elevated CO 2 affects the protein composition of food crops, we performed a comprehensive meta‐analysis, attempting to include all available data for all food crop species.

Results All of the crops included in the analysis had significantly lower protein concentrations when grown at elevated vs. ambient CO 2 (Fig. 1). For potato, the mean reduction in protein was 13.9% and for the grain crops (barley, rice and wheat) the reduction in protein was 15.3%, 9.9% and 9.8%, respectively. For soybean the reduction was a much smaller 1.4%. Figure 1 Open in figure viewer PowerPoint Response of crop protein concentrations to growth at elevated CO 2 for five major crops. Means and 95% confidence limits are depicted. Numbers of experimental observations for each species are in parentheses. For wheat and soybean, the two species with the largest sample size of studies, there were significant differences among CO 2 enrichment technologies in the effect of CO 2 on protein concentration (Fig. 2; P<0.001 for each species). For both species, the largest effects of elevated CO 2 were seen in open‐top chamber studies. For barley and for rice there were no significant differences among CO 2 enrichment technologies. Figure 2 Open in figure viewer PowerPoint Response of crop protein concentrations to growth at elevated CO 2 in studies using various CO 2 enrichment technologies. Means and 95% confidence limits are depicted. Numbers of experimental observations are in parentheses. FACE, free‐air CO 2 enrichment; OTC, open‐top chamber; CTC, closed‐top field chamber; GH, glasshouse; GC, growth chamber. There was some suggestion that CO 2 had a greater effect in studies performed in pots than in studies in which plants were rooted in the ground (Fig. 3). Comparing all studies performed in pots with those involving plants rooted in the ground, no species showed a significant rooting environment effect, although for rice there was a near‐significant trend toward a greater CO 2 effect in pot studies vs. ground studies (P=0.051; Fig. 3a). However, for both soybean and wheat it was additionally possible to make the pot vs. ground comparison focusing solely on studies performed in open‐top‐chambers (OTC; Fig. 3b). For both species, in OTC studies there was a significantly greater effect of CO 2 when plants were grown in pots (Fig. 3b; for soybean P=0.005, for wheat P=0.003). Figure 3 Open in figure viewer PowerPoint Response of crop protein concentrations to growth at elevated CO 2 in studies with plants rooted in pots vs. rooted in the ground. (a) All studies, (b) studies in open‐top chambers. Means and 95% confidence limits are depicted. Numbers of experimental observations are in parentheses. The effect of elevated CO 2 on protein concentrations was affected by environmental variables in several instances. Protein concentrations in wheat grains were reduced more when elevated CO 2 was applied to plants at low N supply than at high N supply (Fig. 4a). Across this group of studies, grain protein concentrations were decreased by 16.4% in low nitrogen treatments compared with 9.8% in high nitrogen treatments, with this difference statistically significant (P=0.038). Figure 4 Open in figure viewer PowerPoint Response of crop protein concentrations to growth at elevated CO 2 in studies that varied (a) nitrogen, (b) temperature or (c) ozone. Each point represents one study. Percent change is the percent change in protein concentration under elevated [CO 2 ]. The diagonal lines represent the 1 : 1 relationship. There was no significant difference in the effect of elevated CO 2 on wheat grain protein concentration between plants grown at high vs. low temperatures (Fig. 4b), although the trend was for elevated CO 2 to have a greater effect on protein concentrations at high than at low temperatures. The effect of ozone differed greatly between species (Fig. 4c). In potato, tuber protein concentrations were decreased by 19.3% under high ozone compared with 7.7% under low ozone, with this difference statistically significant (P=0.013). For soybean, the effect of ozone was the reverse of that seen for potato. Elevated CO 2 increased protein concentrations by 3.0% under high ozone, and decreased protein concentrations by 1.3% under low ozone, with this difference statistically significant (P=0.005). For wheat, the effect of CO 2 on protein concentration was nearly twice as large when protein was measured in grain rather than flour. (Table 1; P=0.004). Table 1. Response of crop protein concentrations to growth at elevated CO 2 for studies on wheat in which protein concentration was measured in grains or flour Itemmeasured Number ofobservations Percent decrease inprotein concentrationunder elevated CO 2 95%confidenceinterval Grain 87 11.0 8.7–13.3 Flour 28 6.0 4.1–8.2

Conclusions Rising atmospheric [CO 2 ] is likely to reduce the protein concentration for many plant crops. The magnitude of this effect is difficult to estimate, due to the sensitivity of this effect to experimental conditions. Nonetheless, decreases in protein are seen consistently for several species across a wide range of experimental techniques and environmental conditions. This effect may be partially mitigated by increased use of nitrogen fertilizers, but this seems likely to be only a partial solution to the effect of elevated CO 2 on the protein concentration of human foods. The effect of atmospheric CO 2 on crop protein therefore seems likely to be of genuine importance for human nutrition in and beyond the 21st century.

Acknowledgements This work was supported by the Fleming Fund for Collaborative Research of Southwestern University. We thank Eli Taub for assistance in typing the manuscript, Lisa Anderson for help in obtaining literature and Xianzhong Wang for valuable comments on the research and manuscript.

Appendix Appendix A. Publications with data included in the analyses. Allen LH, Vu JCV, Valle RR, Boote KJ & Jones PH (1988) Nonstructural carbohydrates and nitrogen of soybean grown under carbon dioxide enrichment. Crop Science, 28, 84–94. Amthor JS, Mitchell RJ, Runion BR, Rogers HH, Prior SA & Wood CW (1994) Energy content, construction cost and phytomass accumulation of Glycine max (L.) Merr. and Sorghum bicolor (L.) Moench grown in elevated CO 2 in the field. New Phytologist, 128, 443–150. Bai Y, Tischler CR, Booth DT & Taylor EM (2003) Variations in germination and grain quality within a rust resistant common wheat germplasm as affected by parental CO 2 conditions. Environmental and Experimental Botany, 50, 159–168. Bencze S, Veisz O & Bedö Z (2004) Effects of high atmospheric CO 2 and heat stress on phytomass, yield and grain quality of winter wheat. Cereal Research Communications, 32, 75–82. Blumenthal C, Rawson HM, McKenzie E, Gras PW, Barlow EWR & Wrigley CW (1996) Changes in wheat grain quality due to doubling the level of atmospheric CO 2 . Cereal Chemistry, 73, 762–766. Chadhuri UN, Burnett RB, Kanemaso ET & Kinkham MB (1986) Effect of Elevated Levels of CO 2 on Winter Wheat under two Moisture Regimes (Response of Vegetation to Carbon Dioxide No. 29). United States Department of Energy, Carbon Dioxide Research Division, Washington DC. Conroy JP (1992) Influence of elevated atmospheric CO 2 concentrations on plant nutrition. Australian Journal of Botany, 40, 445–456. Conroy JP, Seneweera S, Basra AS, Rogers G & Nissen‐Wooler B (1994) Influence of rising atmospheric CO 2 concentrations and temperature on growth, yield and grain quality of cereal crops. Australian Journal of Plant Physiology, 21, 741–758. Cure JD, Israel DW & Rufty TW (1988) Nitrogen stress effects on growth and seed yield of nonnodulated soybean exposed to elevated carbon dioxide. Crop Science, 28, 671–677. Donnelly A, Lawson T, Craigon J, Black CR, Colls JJ & Landon G (2001) Effects of elevated CO 2 and O 3 on tuber quality in potato (Solanum tuberosum L.). Agriculture, Ecosystems and Environment, 87, 273–285. Fangmeier A, Grüters U, Vermehren B & Jäger H‐J (1996) Responses of some cereal cultivars to CO 2 enrichment and tropospheric ozone at different levels of nitrogen supply. Angewandte Botanik, 70, 12–18. Fangmeier A, Grüters U, Högy P, Vermehren B & Jäger H‐J (1997) Effects of elevated CO 2 , nitrogen supply and tropospheric ozone on spring wheat‐ II. Nutrients (N, P, K, S, Ca, Fe, Mg, Zn). Environmental Pollution, 96, 43–59. Fangmeier A, De Temmerman L, Mortensen L, Kemp K, Burke J, Mitchell R, van Oijen M & Weigel HJ (1999) Effects of nutrients on grain quality in spring wheat crops grown under elevated CO 2 concentrations and stress conditions in the European multiple‐site experiment ‘ESPACE‐wheat’. European Journal of Agronomy, 10, 215–229. Fangmeier A, Chrost B, Högy P & Krupinska K (2000) CO 2 enrichment enhances flag leaf senescence in barley due to greater grain nitrogen sink capacity. Environmental and Experimental Botany, 44, 151–164. Fangmeier A, De Temmerman L, Black C, Persson K & Vorne V (2002) Effects of elevated CO 2 and/or ozone on nutrient concentrations and nutrient uptake of potatoes. European Journal of Agronomy, 17, 353–368. Hakala K (1998) Growth and yield potential of spring wheat in a simulated changed climate with increased CO 2 and higher temperature. European Journal of Agronomy, 9, 41–52. Heagle AS, Miller JE & Pursley WA (1998) Influence of ozone stress on soybean response to carbon dioxide enrichment: III. Yield and seed quality. Crop Science, 38, 128–134. Heagle AS, Miller JE & Pursley WA (2003) Atmospheric pollutants and trace gases Growth and yield responses of potato to mixtures of carbon dioxide and ozone. Journal of Environmental Quality, 32, 1603–1610. Israel DW & Rogers HH (1982) The effect of N 2 ‐fixing ability of Rhizobium strain on response of nodulated soybeans to atmospheric CO 2 enrichment. In Field Studies of Plant Responses to Elevated Carbon Dioxide Levels (eds. H.H. Rogers and G.E. Bingham), pp. 122–161. United States Department of Energy, Carbon Dioxide Research Division, Washington, DC. Kimball BA, Morris CF, Pinter PJ, Wall GW, Hunsaker DJ, Adamsen FJ, LaMorte RL, Leavitt SW, Thompson TL, Matthias AD & Brooks TJ (2001) Elevated CO 2 , drought and soil nitrogen effects on wheat grain quality. New Phytologist, 150, 295–303. Kleemola J, Peltonen J & Peltonen‐Sainio P (1994) Apical development and growth of barley under different CO 2 and nitrogen regimes. Journal of Agronomy and Crop Science, 173, 79–92. Lieffering M, Kim H‐Y, Kobayashi K & Okada M (2004) The impact of elevated CO 2 on the elemental concentrations of field‐grown rice grains. Field Crops Research, 88, 279–286. Manderscheid R, Bender J, Jäger H‐J & Weigel HJ (1995) Effects of season long CO 2 enrichment on cereals. II. Nutrient concentrations and grain quality. Agriculture, Ecosystems and Environment, 54, 175–185. Pleijel H, Gelang J, Sild E, Danielsson H, Younis S, Karlsson P‐E, Wallin G, Skärby L & Selldén G (2000) Effects of elevated carbon dioxide, ozone and water availability on spring wheat growth and yield. Physiologia Plantarum, 108, 67–70. Rogers GS, Gras PW, Batey IL, Milham PJ, Payne L & Conroy JP (1998) The influence of atmospheric CO 2 concentration on the protein, starch and mixing properties of wheat flour. Australian Journal of Plant Physiology, 25, 387–393. Rogers GS, Milham PJ, Gillings M & Conroy JP (1996) Sink strength may be the key to growth and nitrogen responses in N‐deficient wheat at elevated CO 2 . Australian Journal of Plant Physiology, 23, 253–264. Rogers HH, Bingham GE, Cure JD, Smith JM & Surano KA (1983) Responses of selected plant species to elevated carbon dioxide in the field. Journal of Environmental Quality, 12, 569–574. Rogers HH, Cure JD & Smith JM (1986) Soybean growth and yield response to elevated carbon dioxide. Agriculture, Ecosystems and Environment, 16, 113–128. Rogers HH, Cure JD, Thomas JF & Smith JM (1984) Influence of elevated CO 2 on growth of soybean plants. Crop Science, 24, 361–366. Rudorff BFT, Mulchi CL, Fenny P, Lee EH & Rowland R (1996) Wheat grain quality under enhanced tropospheric CO 2 and O 3 concentrations. Journal of Environmental Quality, 25, 1384–1388. Sæbø A & Mortensen LM (1996) Growth, morphology and yield of wheat, barley, and oats grown at elevated atmospheric CO 2 concentration in a cool, maritime climate. Agriculture, Ecosystems and Environment, 57, 9–15. Sánchez de la Puente L, Pérez Pérez P, Martínez‐Carrasco R, Morcuende Morcuende R & Martín del Molino IM (2000) Action of elevated CO 2 and high temperatures on the mineral composition of two varieties of wheat. Agrochimica, XLIV, 5–6. Seneweera SP & Conroy JP (1997) Growth, grain yield and quality of rice (Oryza sativa L.) in response to elevated CO 2 and phosphorus nutrition. Soil Science and Plant Nutrition, 43, 1131–1136. Terao T, Miura S, Yanagihara T, Hirose T, Nagat K, Tabuchi H, Kim H, Lieffering M, Okada M & Kobayashi K (2005) Influence of free‐air CO 2 enrichment (FACE) on the eating quality of rice. Journal of the Science of Food and Agriculture, 85, 1861–1868. Thomas JMG, Boote KJ, Allen LH, Gallo‐Meagher M & Davis JM (2003) Elevated temperature and carbon dioxide effects on soybean seed composition and transcript abundance. Crop Science, 43, 1548–1557. Thompson GB & Woodward FI (1994) Some influences of CO 2 enrichment, nitrogen nutrition and competition on grain yield and quality in spring wheat and barley. Journal of Experimental Botany, 45, 937–942. Veisz O, Bencze S & Bedö Z (2005) Effect of elevated CO 2 on wheat at various nutrient supply levels. Cereal Research Communications, 33, 333–336. Weigel HJ & Manderscheid R (2005) CO 2 enrichment effects on forage and grain nitrogen content of pasture and cereal plants. Journal of Crop Improvement, 13, 73–89. Wolf J (1996) Effects of nutrient supply (NPK) on spring wheat response to elevated atmosperic CO 2 . Plant and Soil, 185, 113–123. Wu D‐X, Wang G‐X, Bai Y‐F & Liao J‐X (2004) Effects of elevated CO 2 concentration on growth, water use, yield and grain quality of wheat under two water soil levels. Agriculture, Ecosystems and Environment, 104, 493–507. Ziska LH, Bunce JA & Caulfield F (1998) Intraspecific variation in seed yield of soybean (Glycine max) in response to increased atmospheric carbon dioxide. Australian Journal of Plant Physiology, 25, 801–807. Ziska LH, Namuco O, Moya T & Quilang J (1997) Growth and yield response of field‐grown tropical rice to increasing carbon dioxide and air temperature. Agronomy Journal, 89, 45–53. Ziska LH, Morris CF & Goins EW (2004) Quantitative and qualitative evaluation of selected wheat varieties released since 1903 to increasing atmospheric carbon dioxide: can yield sensitivity to carbon dioxide be a factor in wheat performance? Global Change Biology, 10, 1810–1819.