Anticipatory Life Cycle Analysis of In Vitro Biomass Cultivation for

Article pubs.acs.org/est

Anticipatory Life Cycle Analysis of In Vitro Biomass Cultivation for Cultured Meat Production in the United States Carolyn S. Mattick,*,† Amy E. Landis,‡ Braden R. Allenby,§ and Nicholas J. Genovese∥ †

School of Public Health, University of Texas, 1200 Hermann Pressler Drive, Houston, Texas 77030, United States Glenn Department of Civil Engineering, Clemson University, Clemson, South Carolina 29634, United States § School of Sustainable Engineering and the Built Environment and ASU Lincoln Center for Applied Ethics, Arizona State University, Tempe, Arizona 85287, United States ∥ Department of Medicine, University of Minnesota, Minneapolis, Minnesota 55455, United States ‡

S Supporting Information *

ABSTRACT: Cultured, or in vitro, meat consists of edible biomass grown from animal stem cells in a factory, or carnery. In the coming decades, in vitro biomass cultivation could enable the production of meat without the need to raise livestock. Using an anticipatory life cycle analysis framework, the study described herein examines the environmental implications of this emerging technology and compares the results with published impacts of beef, pork, poultry, and another speculative analysis of cultured biomass. While uncertainty ranges are large, the findings suggest that in vitro biomass cultivation could require smaller quantities of agricultural inputs and land than livestock; however, those benefits could come at the expense of more intensive energy use as biological functions such as digestion and nutrient circulation are replaced by industrial equivalents. From this perspective, large-scale cultivation of in vitro meat and other bioengineered products could represent a new phase of industrialization with inherently complex and challenging trade-offs.

1. INTRODUCTION On August 5, 2013, a hamburger prototype made from cultured, or in vitro, meat was tasted at a well-publicized event in London.1,2 The meat used for preparation of this hamburger was not grown in an animal, but rather from bovine skeletal muscle stem cells in Dr. Mark Post’s laboratory at Maastricht University in the Netherlands. While Dr. Post has suggested that commercialization of cultured meat could be 10−20 years away,1 the event may foreshadow a day when traditional livestock production has given way to large-scale cultivation of meat in factories. The environmental implications of such a transition are potentially profound. 1.1. Goal. This investigation seeks to evaluate the life cycle energy use, global warming potential (GWP), eutrophication potential (EP), and land use associated with in vitro biomass cultivation as an emerging food technology in the United States. It also compares the results to those of another speculative analysis of cultured biomass,3 as well as beef, pork, and poultry impacts from previously published studies.4−6 At the heart of this analysis is a model of a large-scale biomass cultivation facility, or carnery. It is based on established cell culture techniques, but technological change is likely to be rapid in the coming years. Inherent in anticipatory assessments is a tension between building a model that represents a workable process given present knowledge and estimating how © XXXX American Chemical Society

the future commercial process will differ. Thus, anticipatory life cycle analyses (LCAs) can provide valuable insight into how the technology might evolve and affect other coupled systems, but they should be viewed as possible future scenarios rather than predictions. 1.2. Scope. The formation of skeletal muscle tissue for cultured meat is a multistep process (shown in Figure 1) that begins with the isolation of myosatellite cells (adult skeletal muscle stem cells) from a sample of donor animal tissue. When placed in a culture medium containing the necessary factors, the myosatellite cells become proliferative myoblasts that increase in number and aggregate biomass. The culture medium is then modified to induce myoblast differentiation to nonproliferative myocytes that fuse into multinucleated myotubes, which exhibit hypertrophy, increasing in size. This analysis models cultured meat production in two primary phases: The proliferation phase is defined by an increasing myoblast population, and the differentiation phase encompasses all myotube maturation steps. Each phase is modeled as a single batch suspension culture that takes place Received: August 7, 2014 Revised: June 26, 2015 Accepted: September 4, 2015

A

DOI: 10.1021/acs.est.5b01614 Environ. Sci. Technol. XXXX, XXX, XXX−XXX

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Environmental Science & Technology

Figure 1. Phases of muscle cell development for cultured meat production.

Figure 2. System diagram for hypothetical in vitro biomass cultivation system.

transmissible diseases,8,9 serum-free media (SFM) are preferred over animal serum. Serum-free formulations have been developed for many animal cell types,10 and the adaptation of myoblasts to a SFM will be a fundamental step toward reducing costs and reliance upon livestock-derived components.11 Hydrolysates (enzymatic or acid digests) of yeast, rice, soy, and other plant and microbial materials may be added to basal media as supplementary sources of amino acids, peptides, vitamins, and trace elements.12 For this study, we assume that cultured meat will be produced in a SFM supplemented with

within a stirred-tank bioreactor. This is a simple but widely used configuration for producing viral vaccines and recombinant proteins from animal cell cultures.7 Culture media typically consist of water, glucose, and predefined mixtures called “basal media” that contain amino acids, lipids, vitamins, and salts.7 In addition, animal serum is often added to cell cultures because it contains many of the factors required for cell attachment, growth, and proliferation.8 However, for a number of reasons including high costs, unsteady supplies, lot variation, and the possible presence of B


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All three livestock LCAs reported environmental impacts in terms of live animal weight. In order to obtain a comparable functional unit of edible biomass, an economic allocation was applied such that impacts were assigned to meat and its coproducts on the basis of relative market value. Please see section S2.2 (SI) for additional information. Note that, in the United States, beef production remains largely separate from dairies,17 and broiler chickens are slaughtered prior to sexual maturity.6,18 Hence the only coproducts of meat production are derived from the carcass: hide, fat, offal, feathers, etc.

soy hydrolysate. A diagram of the cultured meat production model used for this study is given in Figure 2. 1.3. Functional Unit. The LCA described herein is a cradle-to-gate analysis of animal cell biomass expansion, in vitro, as modeled using the SimaPro 7.3.3 LCA software package from PRé Consultants.13 Metabolic requirements specific to muscle cell cultivation could not be located; therefore, the life cycle inventory is based on Chinese Hamster Ovary (CHO) cell cultivation. Thus, the functional unit is 1 kg of CHO cell biomass. Future empirical studies will be necessary to determine the relative metabolic requirements of CHO cell and skeletal muscle bioprocess production models.14 1.4. Allocation Procedures. Environmental impacts for in vitro biomass inputs and their coproducts are allocated on a gross chemical (calorific) energy basis, consistent with the approach utilized for the comparative livestock LCAs.4−6 [Please see section S2 in the Supporting Information (SI) for a detailed explanation.] 1.5. Impact Assessment. All impact assessment methods were chosen to be consistent with the beef,4 pork,5 and poultry6 LCAs (see Table S-4, SI). Industrial energy use was computed using the cumulative energy demand method,15 which converts final energy inputs to primary energy inputs using fixed heating values for fuels. Land use was found via the ecological footprint method,16 which quantifies direct land occupation associated with human activities. Although this method also estimates values for time-integrated land required to produce fuels and assimilate wastes, these indirect uses of land are excluded from this analysis. All other impacts were found via the CML (Center of Environmental Science of Leiden University) 2001 (World 1995) characterization methods,16 which express environmental impacts in terms of resource use and emissions rather than realized outcomes.4 For example, its GWP method converts greenhouse gas (GHG) emissions to carbon dioxide (CO2) equivalent mass based on heat retained over a 100-year time horizon, and its EP method expresses emissions of nitrogen- and phosphorus-containing compounds in terms of phosphate (PO4) equivalent mass. 1.6. Comparison Studies. The results of this analysis are compared to the results of four previously published studies. The first is another speculative analysis of cultured biomass performed by Tuomisto and Teixeira de Mattos.3 A detailed comparison of the two approaches is provided in Table S-1 (SI). Briefly, the prior LCA3 relied on cyanobacteria hydrolysate as the sole feedstock and did not include basal media production, a medium change between proliferation and differentiation phases, bioreactor cleaning, or a production facility. Even though that study presented results for Thailand, California, and Spain, the results included here reflect the California findings only. The functional unit in that study was 1000 kg of cultured meat, so results were divided by 1000 prior to inclusion here. The three remaining studies are LCAs of beef,4 pork,5 and broiler poultry6 production in the United States. Multiple scenarios are analyzed in the beef and pork studies, but only the results of the “feedlot” beef4 and the “commodity (high profit)” pork5 cases are presented here. These are representative of U.S. production strategies that are widely practiced and have the lowest environmental impacts of those assessed. In the poultry study, litter was considered a coproduct and credited with avoided environmental burdens of synthetic fertilizer production.6 These credits have been removed to achieve consistency with the other LCAs.

2. LIFE CYCLE INVENTORY ANALYSIS The life cycle inventory for the proliferation phase is based on the CHO cell culture data presented by Sung et al.12 The phase begins with an initial culture density of 2 × 105 cells/mL in a stirred-tank bioreactor containing basal medium. Cell proliferation then proceeds at a specific growth rate of 0.0254 cells/(cell h),12 equivalent to a doubling time of about 27 h. The buildup of ammonia is known to be a primary inhibitor of cellular growth.19,20 Therefore, it is assumed that proliferation is terminated before the ammonia concentration reaches 2 mM, the threshold at which cell growth would be impacted.20 This allows a cell concentration of 4 × 106 cells/mL, or 210 kg of biomass, to be achieved in about 5 days (see section S4 of the SI for computation). The differentiation phase begins by draining the proliferation culture medium and refilling the bioreactor with differentiation medium. The phase then proceeds for 72 h,21 during which time the number of nuclei remains constant while cells fuse into myotubes and exhibit hypertrophy. The mass increase during this phase is assumed to be proportional to the mass difference between myotubes cultured from cells donated by healthy individuals and those donated by chronic obstructive pulmonary disease (COPD) patients, whose derivative myotubes exhibit impaired hypertrophy. Mass indicators for each group were computed as the average cross-sectional areas of myotubes divided by the average number of nuclei per cell. On the basis of data from Pomiès et al.,22 a 64% mass increase is modeled during the differentiation phase. 2.1. Feedstocks and Byproducts. The primary metabolic inputs for batch suspension cell culture are assumed to be glucose, oxygen, and the amino acid glutamine; the primary byproducts are alanine, ammonia, and lactate.12 Equations for determining nutrient uptake and byproduct formation rates are given in section S4 (SI). Specific consumption rates for basal media and hydrolysates are not generally computed; instead, they are mixed at predefined concentrations. Basal media formulations are detailed in Table S-12 (SI), and the soy hydrolysate concentration is modeled at 5 g/L since this has been shown to yield optimal biomass growth.12,23 2.2. Microcarrier Beads. A scaffold, which mimics the extracellular matrices found in vivo, may be used to provide structural support during proliferation and maturation of developing tissue.24 Microcarrier bead scaffolds (spheres having diameters between 100 and 300 μm) support autonomous cellular organization and contraction2,11,25 and allow myotubes to mature into skeletal muscle tissue.2,11 Challenges surround the choice of scaffold material for cultured meat, since it must be edible, dissolve or degrade prior to consumption, or be easily excluded from cell harvest. Recently, researchers have obtained encouraging results growing skeletal muscle cells in suspension culture on microspheres made of starch, which is both edible and abundantly available.26 For the purposes of this study, the C


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Energy used within the biomass production facility was estimated using the pharmaceutical and beer industries as guides. Both industries utilize bioprocessing infrastructure featuring suspension culture systems. The bioreactor configuration in this study was modeled on the Biogen IDEC largescale pharmaceutical manufacturing plant in Research Triangle Park, NC,7,29 which houses six 15 000-L stirred-tank reactors on 245 000 ft2 of floor space.7 Due to uncertainties associated with cultured meat production and its need for cleanroom, research, and other equipment housed within pharmaceutical plants, the building size was modeled on the smaller footprint of a brewery. Thus, the plant was assumed to have 717 m2 (7717 ft2) of floorspace, and the derivation of this value can be found in section S5 of the SI. The baseline facility energy required for lighting, HVAC, and other purposes was assumed to be equivalent to a warehouse: 513 MJ/m2/year.30 The mix of fuels for cultured meat production processes was assumed to be the same as for the brewing industry and is shown in Table S-6 (SI). 2.4. Bioreactor Processes. Each phase of production requires approximately 15 000 L (the bioreactor capacity) of deionized water31 for culture medium dissolution. To avoid culture contamination, the medium must be sterilized before any cells are introduced. Since sterilization by heat could destroy critical proteins in the culture medium,31,32 sterilization via microfiltration (at 0.15 kWh/m3)33 was modeled. The bioreactor also requires energy for aerating, mixing, and regulating the temperature of the culture medium. Aeration, also known as “sparging”, is the process of delivering gases such as oxygen (to support cellular respiration) and carbon dioxide (for pH management) to the bioreactor tank. It may be accomplished by introducing pure gases and/or filtered atmospheric air into the tank via fine-bubble diffusers. This model assumed that atmospheric air could be delivered at 2 kg O2/kWh,34 and that 4% CO2 would be required for pH regulation.7 Sparged gases must be evenly distributed throughout the culture medium to ensure a favorable environment for all cells. In the stirred-tank reactor design, this is accomplished by agitating the fluid via impeller rotation. The impeller speed must be fast enough to maintain the culture suspension but not so fast that cells are damaged.35 For this reason, impeller speeds are often limited to 1.5 m/s.36 Finally, the cells must be maintained at a temperature that supports cell proliferation. It is assumed that the culture media are initially heated to 37 °C and maintained at this temperature for the duration of the proliferation and differentiation phases. In order to offset the heat produced by cellular respiration, thermal regulation was modeled as the process of pumping water at ambient temperature (23 °C) through a heat exchanger in the bioreactor walls. Detailed calculations for all modeled energy inputs can be found in section S6 (SI). 2.5. Bioreactor Cleaning in Place. Bioreactors must be cleaned and sanitized between each batch cycle to ensure sterile production conditions. Per Chisti and Moo-Young,37 this LCA assumes that cleaning procedures follow a three-step process: The tank is first rinsed with deionized water and drained; then a 1% w/v sodium hydroxide solution is added, heated from 23 to 77.5 °C, and drained; finally, the tank is rinsed again with deionized water. The inventory for the in vitro production model is summarized in Table 1. Inventories for all subprocesses are provided in section S7 (SI).

Table 1. Inventory Required To Produce In Vitro Biomass for Cultured Meata substance

per year for facility

per batch for proliferation

per batch for differentiation

per batch for cleaning

Inputs land use (m2) biomass (kg) starch microcarrier beads (kg) deionized water (1000 L) glucose (kg) glutamine (kg) oxygen (kg)c soy hydrolysate (kg) basal medium (1000 L)d carbon dioxide, sparged (kg) sodium hydroxide (kg) energy (MJ) facility sterile filtration heating water agitation aeration heat exchanger transportation (t km)e biomass (kg) lactate (kg) alanine (kg)f ammonia (kg) spent medium (1000 L)d sodium hydroxide (kg) carbon dioxide, sparged (kg)

717 10.5b 75

210

14.9

14.7

70.4 5.9 23.1 0.8

135.6 11.4 44.5

14.9

14.7

8.8

17.0

45.0

150

368033 8.1

7.9

874

859

191 41.6 0.46

116 80.2 0.64

207

165

Outputs 210 56.5 1.3 0.5 14.9

3417

75

345 108.8 2.4 1.0 14.7 150

8.8

17

a

Note that the facility as described could complete 192 batches and produce 66 000 kg per year. bSeed culture for proliferation is not included in LCA but is addressed in the sensitivity analysis. cOxygen mass is used for aeration energy computation only. dBasal medium consists of dry matter but is expressed in terms of liquid volume. Detailed inventories for the basal media are given in Table S-12 (SI). e All dry ingredients are assumed to travel 500 km by diesel truck. f Alanine was converted to chemical oxygen demand (COD) by multiplying the quantity produced by a factor of 1.36 g COD/g alanine.38

scaffold was assumed to be microcarrier beads made from corn starch, added at a concentration of 5 mg/mL in the bioreactor.27 2.3. Production Facility. To be consistent with the referenced livestock LCAs, energy embodied in buildings and capital equipment was excluded. Even though greater capital investment may be associated with industrial production versus agriculture, LCAs typically demonstrate that the contribution of infrastructure to environmental impacts is negligible for energyintensive processes (see p 185 of ref 28). D


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Figure 3. Results: Industrial energy use and GHG emissions. This figure compares the results of this study with impacts of feedlot beef,4 commodity (high-profit) pork,5 poultry,6 and in vitro biomass produced in California3 found by prior studies. “Other factors” consist predominantly of the cost of bull production for beef,4 sow replacement for pork,5 and hatchery chicks for poultry.6

3. RESULTS The environmental impact comparisons for energy consumption, GWP, EP, and land use are shown in Figures 3 and 4. Agricultural processes are shown in green, industrial processes in purple, and waste products in orange. Black error bars indicate that the results have high uncertainty. As discussed in section 4, actual production facilities and cell growth characteristics may differ from the assumptions underlying the modeled system, resulting in large deviations from the impacts reported here. 3.1. Comparison of this Study with Prior Cultured Meat LCA. The results show that energy consumption and GWP estimated by this study are approximately 3 times the prior values, due mostly to the inclusion of basal media production and the cleaning phase. Land use requirements for this study total roughly 20 times those of the previous LCA due to the use of a different feedstock and additional culture inputs, such as basal media and soy hydrolysate. The seemingly burdensome nature of the model used in this study might lead some to conclude that it is the inferior choice compared to the Tuomisto and Teixeira de Mattos3 approach. This study adhered to established cell culture protocols, but technological advances could bring commercial processes in line with the prior study’s results. Similarly, the feedstock and basal media formulas used in this study were based on peerreviewed studies demonstrating their effectiveness for support-

ing cell growth. Empirical data would be required to assess the nutritional adequacy of cyanobacteria hydrolysate alone for large-scale cell cultures. 3.2. Comparison of this Study with Conventional Meat. Figure 3 indicates that in vitro biomass will require more industrial energy than livestock production, though uncertainties suggest that cultured meat could be on par with beef under some production conditions. It follows that, because of its substantial energy requirement, cultured meat is likely to have a larger GWP than pork or poultry. Due to nitrous oxide and methane emissions from cattle manure, however, cultured meat could have a lower GWP than beef. The color scheme followed in Figure 3 suggests that nearly all industrial energy for in vitro biomass is required for industrial processes such as basal media production. Basal media contain amino acids that can be manufactured synthetically in a series of steps that begins with corn production but is followed by corn milling, saccharification of corn starch to glucose, and fermentation of the glucose. Whereas Figure 4 indicates that in vitro biomass could require less occupied land and smaller feedstock quantities than livestock, these benefits may come at the expense of industrial energy consumption. 3.3. Eutrophication Potential. The eutrophication chart in Figure 4 should be viewed with some caution. Whereas the eutrophication assessments for livestock assume managed waste E


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Figure 4. Results: Eutrophication potential and land use. This figure compares the results of this study with impacts of feedlot beef,4 commodity (high-profit) pork,5 poultry,6 and in vitro biomass produced in California3 found by prior studies. Eutrophication impacts for livestock assume managed waste flows that are ultimately applied to agricultural fields; eutrophication potential for in vitro biomass was assessed for untreated waste streams. Eutrophication impacts due to spent media were excluded from this graph. Land use consists of land occupation values for beef and feed production values for pork; it excludes area required to sequester atmospheric CO2 emissions and the land area associated with nuclear energy. “Other factors” consist predominantly of the cost of bull production for beef,4 sow replacement for pork,5 and hatchery chicks for poultry.6

flows, 4−6 the value for in vitro biomass reflects the eutrophication potential of untreated waste streams. For this reason, the columns shown in Figure 4 are not directly comparable. However, as with livestock operations, carneries may construct on-site treatment or recycling systems to limit emitted pollution either voluntarily or in response to state and federal regulations. Hence, eutrophying emissions from carneries could be on par with poultry operations or below, but it should further be noted that waste treatment methods may increase impacts in other categories. The eutrophication potential associated with the spent media for cultured meat (about 19 g PO4-equiv) was excluded from this graph. The spent media are more analogous to animal bodily fluids and, therefore, waste streams from slaughterhouses than metabolic waste products. Their exclusion facilitates a more equivalent comparison of life cycle processes in this category. 3.4. Energy Return on Investment. Energy return on investment (EROI), or energy conversion efficiency, is an expression of how much useful food energy results from the food or industrial energy invested. The EROI focuses specifically on available human-edible biomass and the gross chemical (calorific) energy contained therein. For each of the sources considered (beef, pork, poultry, and in vitro biomass), Figure 5 depicts human-edible energy output divided by human-edible and industrial energy input. It further underscores the phenomenon discussed above that in vitro muscle

biomass cultivation will utilize agricultural feedstocks more efficiently than animals at the expense of industrial energy inputs. These energy dynamics may be better understood through the analogy of the Industrial Revolution: Just as automobiles and tractors burning fossil fuels replaced the external work done by horses eating hay, in vitro biomass cultivation may similarly substitute industrial processes for the internal, biological work done by animal physiologies. That is, meat production in animals is made possible by internal biological functions (temperature regulation, digestion, oxygenation, nutrient distribution, disease prevention, etc.) fueled by agricultural energy inputs (feed). Producing meat in a bioreactor could mean that these same functions will be performed at the expense of industrial energy, rather than biotic energy. As such, in vitro biomass cultivation could be viewed as a renewed wave of industrialization.

4. SENSITIVITY ANALYSIS A Monte Carlo Analysis was performed to assess model sensitivity in identified areas of uncertainty. A summary of all uncertainty variables, their properties, and results can be found in section S9 of the SI. Each Monte Carlo simulation continued until a standard error of mean equal to 0.005 was reached. In aggregate, the analysis shows a great deal of uncertainty, with the most significant variability coming from facility size, F


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by 30% under the optimistic scenario or increase by 55% if no growth is achieved.

5. DISCUSSION This analysis suggests that in vitro biomass cultivation may constitute a new phase of industrialization. As such, it could bring a number of benefits to the world along with shifting environmental burdens: Even though industrial energy consumption may rise, agricultural land requirements could decline. To the extent that they can be foreseen, an understanding of the potential environmental implications of a technology can facilitate anticipation and mitigation of unintended consequences prior to and during commercialization. This study suggests that variances in a few key production factors could lead to large changes in energy use, GWP, EP, and land use (Figure S-4, SI); these factors might be candidates for targeted innovation. This analysis also compares the environmental impacts associated with two speculative analyses of in vitro biomass cultivation. The disparate choices inherent in the underlying models serve to underscore an important point about anticipatory LCA: Until working manufacturing facilities have been constructed, LCAs are only hypothetical scenarios. Even though significant uncertainty surrounds anticipatory analyses, they remain valuable for highlighting the possible implications of, and trade-offs associated with, emerging technologies as they advance. Future assessments may also consider factors like water requirements, potential land use changes and their subsequent effects, as well as complex impacts beyond the environmental realm, such as shifts in economic activity40 and social norms.41

Figure 5. EROI for livestock and in vitro biomass production. Industrial EROI is the human-edible energy return on industrial energy investment, and human-edible EROI is the human-edible energy return on human-edible caloric energy investment.4 The human-edible EROI for poultry was obtained from Smil (p 140 of ref 39) and assumes all poultry feed is human-edible. The sources for all other values are beef (feedlot),4 pork (commodity high profit),5 poultry,6 and in vitro production as modeled by this study. Boneless meat yields are assumed to be 43%, 56%, and 56% of live-weight cattle, hogs, and poultry, respectively. In Vitro values assume 100% edible yield. Raw, boneless meat and in vitro biomass are assumed to have an energy density of 4.63 MJ/kg.4−6 Error bars for in vitro human-edible EROI represent estimated uncertainty values; error bars for in vitro industrial EROI depict the results of the sensitivity analysis.

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ASSOCIATED CONTENT

* Supporting Information S

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.est.5b01614. A comparison of the model used in this study with that of the prior in vitro LCA; impact allocation procedures; life cycle impact methods used; cell culture model details; calculation of facility floorspace; bioreactor energy calculations; feedstock and subprocess inventories; detailed results of the impact analysis; and detailed results of the sensitivity analysis (PDF)

maximum cell concentration, and biomass increase during the differentiation phase. For the baseline model, it was assumed that the building would be similar in size to a brewery (717 m2). The sensitivity analysis explored the effects of expanding the building size to a pharmaceutical plant equivalent (22 761 m2) and found that this scenario could roughly triple energy use and GHG emissions (see Figure S-4, SI). The analysis also considered the effects of varying the maximum attainable cell concentration at the end of the proliferation phase from a minimum of 1.67 × 106 cells/mL, simulating poor growth conditions, to a maximum of 2 × 107 cells/mL, simulating more optimal conditions. The results showed that a higher cell concentration could potentially decrease environmental impacts by 50−70% in all categories considered. This can be explained by the exponential growth characteristics of cell culture: Achieving a higher cell density at the end of the proliferation cycle would result in more usable product per unit input. However, concomitant changes such as increased basal medium requirements or changes in bioreactor design (e.g., use of hollow fiber or perfusion reactors) were not part of the analysis and could serve to offset the impact reductions. Finally, uncertainties associated with biomass increase during the differentiation phase were modeled to range from no growth to twice the baseline production (128% biomass increase). The results indicated that impacts could decrease

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AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. Author Contributions

The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript. Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS The authors would like to thank the Graduate College at Arizona State University and the Lincoln Center for Applied Ethics at Arizona State University for their generous support. The authors are also grateful to People for the Ethical Treatment of Animals for their support of N.J.G.’s work. The G


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T-PA Production and Feed Glutamine Replacement to Reduce Ammonia Production. Biotechnol. Prog. 2013, 29, 165−175. (20) Altamirano, C.; Berrios, J. Advances in Improving Mammalian Cells Metabolism for Recombinant Protein Production. Electron. J. Biotechnol. 2013, 16, DOI: 10.2225/vol16-issue3-fulltext-2. (21) Doumit, M.; Cook, D.; Merkel, R. Testosterone up-Regulates Androgen Receptors and Decreases Differentiation of Porcine Myogenic Satellite Cells in Vitro. Endocrinology 1996, 137, 1385− 1394. (22) Pomiès, P.; Rodriguez, J.; Blaquière, M.; Sedraoui, S.; Gouzi, F.; Carnac, G.; Laoudj-Chenivesse, D.; Mercier, J.; Préfaut, C.; Hayot, M. Reduced Myotube Diameter, Atrophic Signalling and Elevated Oxidative Stress in Cultured Satellite Cells from COPD Patients. J. Cell. Mol. Med. 2015, 19, 175−186. (23) Chun, B.-H.; Kim, J.-H.; Lee, H.-J.; Chung, N. Usability of SizeExcluded Fractions of Soy Protein Hydrolysates for Growth and Viability of Chinese Hamster Ovary Cells in Protein-Free Suspension Culture. Bioresour. Technol. 2007, 98, 1000−1005. (24) Kosztin, I.; Vunjak-Novakovic, G.; Forgacs, G. Colloquium: Modeling the Dynamics of Multicellular Systems: Application to Tissue Engineering. Rev. Mod. Phys. 2012, 84, 1791−1805. (25) Bardouille, C.; Lehmann, J.; Heimann, P.; Jockusch, H. Growth and Differentiation of Permanent and Secondary Mouse Myogenic Cell Lines on Microcarriers. Appl. Microbiol. Biotechnol. 2001, 55, 556−562. (26) Wallin, P.; Hoglund, K.; Wildt-Persson, K.; Gold, J. Skeletal Myoblast Differentiation on Starch Microspheres for the Development of Cultured Meat. J. Tissue Eng. Regen. Med. 2012, 6, 378. (27) Nasser, A.; El-Moghaz. Factors Effects the Growth of Chinese Hamster Ovary (CHO) Cell on Microcarriers Culture. Adv. Biores. 2010, 1, 182−188. (28) Tsiropoulos, I.; Cok, B.; Patel, M. K. Energy and Greenhouse Gas Assessment of European Glucose Production from Corn − a Multiple Allocation Approach for a Key Ingredient of the Bio-Based Economy. J. Cleaner Prod. 2013, 43, 182−190. (29) Biogen’s LSM plant; on line, on time, on budget. http://www. pharmamanufacturing.com/articles/2003/100/. (30) D&R International Ltd. 2011 Buildings Energy Data Book; D&R International Ltd: Silver Spring, MD, 2012. (31) Shuler, Michael, L.; Karki, F. Bioprocess Engineering, 2nd ed.; Prentice Hall: Upper Saddle River, NJ, 2002. (32) Heinzle, E.; Biwer, A. P.; Cooney, C. L. Development of Sustainable Bioprocesses: Modeling and Assessment; John Wiley & Sons: Chichester, England, 2006. (33) Chang, Y.; Reardon, D. J.; Kwan, P.; Boyd, G.; Brant, J.; Rakness, K. L.; Furukawa, D. Evaluation of Dynamic Energy Consumption of Advanced Water and Wastewater Treatment Technologies; American Water Works Association: Arnaudville, LA, 2008. (34) Xylem. Aeration products for energy-efficient biological treatment. http://www.wwdmag.com/sites/default/files/whitepapers/ SB004-460_Sanitaire_Aeration_Products_brochure_sm_3.pdf. (35) Yang, S.-T.; Luo, J.; Chen, C. A Fibrous-Bed Bioreactor for Continuous Production of Monoclonal Antibody by Hybridoma. In Advances in Biochemical Engineering/Biotechnology; Zhong, J.-J., Ed.; Springer-Verlag: New York, 2004; pp 61−96. (36) Nienow, A. W. Reactor Engineering in Large Scale Animal Cell Culture. Cytotechnology 2006, 50, 9−33. (37) Chisti, Y.; Moo-Young, M. Clean-in-Place Systems for Industrial Bioreactors: Design, Validation and Operation. J. Ind. Microbiol. 1994, 13, 201−207. (38) World Bank Group. Pollution Prevention and Abatement Handbook, 1998; World Bank Publications: Washington, DC, 1999. (39) Smil, V. Should We Eat Meat?; Wiley-Blackwell: West Sussex, UK, 2013. (40) Mattick, C. S.; Landis, A. E.; Allenby, B. R. A Case for Systemic Environmental Analysis of Cultured Meat. J. Integr. Agric. 2015, 14, 249−254.

authors gratefully acknowledge the anonymous reviewers whose insightful suggestions helped to improve this analysis.

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ABBREVIATIONS USED EP eutrophication potential EROI energy return on investment GHG greenhouse gas GWP global warming potential LCA life cycle analysis SFM serum-free media

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REFERENCES

(1) Cultured beef: The event. http://culturedbeef.net/event/. (2) Post, M. J. Cultured Beef: Medical Technology to Produce Food. J. Sci. Food Agric. 2014, 94, 1039−1041. (3) Tuomisto, H. L.; Teixeira de Mattos, M. J. Environmental Impacts of Cultured Meat Production. Environ. Sci. Technol. 2011, 45, 6117−6123. (4) Pelletier, N.; Pirog, R.; Rasmussen, R. Comparative Life Cycle Environmental Impacts of Three Beef Production Strategies in the Upper Midwestern United States. Agric. Syst. 2010, 103, 380−389. (5) Pelletier, N.; Lammers, P.; Stender, D.; Pirog, R. Life Cycle Assessment of High- and Low-Profitability Commodity and DeepBedded Niche Swine Production Systems in the Upper Midwestern United States. Agric. Syst. 2010, 103, 599−608. (6) Pelletier, N. Environmental Performance in the US Broiler Poultry Sector: Life Cycle Energy Use and Greenhouse Gas, Ozone Depleting, Acidifying and Eutrophying Emissions. Agric. Syst. 2008, 98, 67−73. (7) Hu, W.-S. Cell Culture Bioprocess Engineering; Wei-Shou Hu: Minneapolis, MN, 2012. (8) Brunner, D.; Frank, J.; Appl, H.; Schöffl, H.; Pfaller, W.; Gstraunthaler, G. Serum-Free Cell Culture: The Serum-Free Media Interactive Online Database. ALTEX 2010, 27, 53−62. (9) Girón-Calle, J.; Vioque, J.; Pedroche, J.; Alaiz, M.; Yust, M. M.; Megías, C.; Millán, F. Chickpea Protein Hydrolysate as a Substitute for Serum in Cell Culture. Cytotechnology 2008, 57, 263−272. (10) Whitford, W. G. Fed-Batch Mammalian Cell Culture in Bioproduction. Bioprocess Int. 2006, 4, 30−40. (11) Post, M. J. Cultured Meat from Stem Cells: Challenges and Prospects. Meat Sci. 2012, 92, 297−301. (12) Sung, Y. H.; Lim, S. W.; Chung, J. Y.; Lee, G. M. Yeast Hydrolysate as a Low-Cost Additive to Serum-Free Medium for the Production of Human Thrombopoietin in Suspension Cultures of Chinese Hamster Ovary Cells. Appl. Microbiol. Biotechnol. 2004, 63, 527−536. (13) PRé Consultants. SimaPro 7.3.3; http://www.pre-sustainability. com/software. (14) Cheng, C. S.; Davis, B. N. J.; Madden, L.; Bursac, N.; Truskey, G. A. Physiology and Metabolism of Tissue-Engineered Skeletal Muscle. Exp. Biol. Med. 2014, 239, 1203−1214. (15) Frischknecht, R.; Jungbluth, N.; Althaus, H. J.; Doka, G.; Dones, R.; Hischier, R.; Hellweg, S.; Humbert, S.; Margni, M.; Nemecek, T.; et al. Implementation of Life Cycle Impact Assessment Methods; Final report ecoinvent 2000; ecoinvent: Duebendorf, Switzerland, 2003. (16) Frischknecht, R.; Jungbluth, N.; Althaus, H. J.; Doka, G.; Dones, R.; Hischier, R.; Hellweg, S.; Humbert, S.; Margni, M.; Nemecek, T.; et al. Implementation of Life Cycle Impact Assessment Methods: Data v2.0; ecoinvent report no. 3; ecoinvent: Duebendorf, Switzerland, 2007. (17) United States Department of Agriculture. Cattle & beef: Background. http://www.ers.usda.gov/topics/animal-products/cattlebeef/background.aspx#.U2B9lfldXh4. (18) US Environmental Protection Agency. Poultry production phases. http://www.epa.gov/oecaagct/ag101/poultryphases.html (accessed Jun 18, 2015). (19) Kim, D. Y.; Chaudhry, M. A.; Kennard, M. L.; Jardon, M. a; Braasch, K.; Dionne, B.; Butler, M.; Piret, J. M. Fed-Batch CHO Cell H


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