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Poult Sci 2008. 87:104-107. doi:10.3382/ps.2007-00255
© 2008 Poultry Science Association
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METABOLISM AND NUTRITION: Research Notes

Nitrogen-Corrected Apparent Metabolizable Energy Value of Crude Glycerol for Laying Hens1

P. J. Lammers*, B. J. Kerr{dagger}, M. S. Honeyman*, K. Stalder*, W. A. Dozier, III{ddagger}, T. E. Weber{dagger}, M. T. Kidd§ and K. Bregendahl*,2

* Department of Animal Science, Iowa State University, Ames 50011; {dagger} USDA-ARS, Swine Odor and Manure Management Research Unit, Ames, IA 50011; {ddagger} USDA-ARS Poultry Research Unit, Mississippi State, MS 39762; and § Department of Poultry Science, Mississippi State University, Mississippi State 39762

2 Corresponding author: kristjan{at}iastate.edu


   ABSTRACT
TOP
 ABSTRACT
INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
An experiment was conducted with laying hens to determine the AMEn value of crude glycerol, a coproduct of biodiesel production. Crude glycerol (87% glycerol, 9% water, 0.03% methanol, 1.26% Na, and 3,625 kcal/kg of gross energy) was obtained from a commercial biodiesel production facility (Ag Processing Inc., Sergeant Bluff, IA). A total of forty-eight 40-wk-old laying hens (Hy-Line W-36) were placed in metabolic cages (2 hens/ cage) and given free access to the experimental diets. A corn and soybean meal-based basal diet (18% CP, 2,875 kcal/kg of AMEn, 4.51% Ca, 0.51% nonphytate P) was formulated with 15% glucose·H2O and 1% Celite. Four dietary treatments were created by substituting 0, 5, 10, or 15% crude glycerol for glucose·H2O (3,640 kcal/kg of AMEn). After 7 d of dietary adaptation, excreta were collected twice daily for 3 d, freeze-dried, and analyzed for contents of DM, Kjeldahl N, acid-insoluble ash, and gross energy. Egg production was recorded daily, and eggs were collected on d 7 and 8 of the experiment for calculation of egg mass (egg production x egg weight). Feed consumption was measured over the 10-d experimental period. Egg-production data were analyzed by ANOVA with 4 treatments and 6 replications in a completely randomized experimental design. The AMEn value of crude glycerol was estimated as the slope of the linear relationship between the inclusion rate of dietary crude glycerol and the glucose-corrected AMEn value of the experimental diets. No significant treatment effects (P > 0.1) were apparent for egg-production rate (93.0%), egg weight (56.1 g), egg mass (52.2 g/d), or feed consumption (104 g/d). Linear regression analysis (P < 0.001, r2 = 0.92, n = 24) revealed that the AMEn value of the crude glycerol used in this study was 3,805 ± 238 kcal/kg (mean ± SEM; as-is basis) for laying hens.

Key Words: crude glycerol • biodiesel co-product • nitrogen-corrected apparent metabolizable energy • laying hen


   INTRODUCTION
TOP
 ABSTRACT
 INTRODUCTION
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
In the United States, the production of diesel fuel from vegetable oil has increased exponentially from less than 2 million liters in 1999 to almost 1 billion liters in 2005 (National Biodiesel Board, 2007). Biodiesel is produced through a NaOH- or KOH-catalyzed transesterification of the triacylglycerols in oils or fats with an alcohol, usually methanol (Ma and Hanna, 1999; van Gerpen, 2005). Through this reaction, the fatty acids are methylated to form methyl alkyl esters (i.e., biodiesel), and the principal coproduct from the process is crude glycerol (Ma and Hanna, 1999; van Gerpen, 2005; Thompson and He, 2006). Using current refinement techniques, production of 1,000 L of biodiesel results in the generation of 79 kg of crude glycerol (Thompson and He, 2006). Although chemically pure glycerol is a valuable industrial compound for use in consumer products, such as cosmetics and pharmaceuticals (Ma and Hanna, 1999; Thompson and He, 2006), purification to meet United States Pharmacopeia specifications may become uneconomical given the existing capacity and projected growth in biodiesel production. Thus, substantial amounts of crude glycerol may become available for use as livestock feed at relatively economical costs.

Free glycerol only occurs in minute amounts, if at all, in feed ingredients, but glycerol is regularly consumed as part of triacylglycerols. Glycerol is a precursor to glyceraldehyde 3-phosphate, an intermediate in the lipogenesis and gluconeogenesis pathways, and yields energy through the glycolytic and tricarboxylic-acid pathways (Lin, 1977; Tao et al., 1983; Brisson et al., 2001). Studies examining the effects of feeding chemically pure or crude glycerol from biodiesel production to broiler chickens (Simon et al., 1996; Cerrate et al., 2006), turkey hens (Rosebrough et al., 1980), and pigs (Kijora et al., 1995) have shown that glycerol can be used a source of dietary energy for livestock. There are no reports, however, in which the AMEn of crude glycerol has been directly determined. The objective of the study was to determine the AMEn of crude glycerol when fed to laying hens.


   MATERIALS AND METHODS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
A total of forty-eight 40-wk-old Single-Comb White Leghorn laying hens (Hy-Line W-36) were obtained from a commercial source and placed in metabolic cages (30.5 x 50.8 x 40.6 cm, width x depth x height, respectively), equipped for collection of excreta, in a light-controlled, fan-ventilated room at the Iowa State University Poultry Science Research Center. Each cage contained 2 hens and was equipped with a steel self-feeder and a trough waterer. Upon arrival, hens were given free access to a laying hen diet (17% CP, 2,875 kcal/kg of AMEn, 4.40% Ca, 0.43% nonphytate P) and water for a 2-wk acclimation period. Hens were provided with 16 h of light and 8 h of darkness per day, and the ambient temperature was maintained at 26°C throughout the study. After the 2-wk acclimatization period, hens were weighed and randomly assigned to cages. Each cage was randomly assigned to 1 of the 4 experimental diets. Hens were given free access to the experimental diets for a 7-d adaptation period followed by a 3-d collection period. All procedures relating to the use of live animals were approved by the Iowa State University Institutional Animal Care and Use Committee.

A total of 4 experimental diets were used, formulated from a basal diet (Table 1Go) in which 5, 10, or 15% crude glycerol (Ag Processing Inc., Sergeant Bluff, IA; Table 2Go) was substituted for glucose·H2O on an equal-weight basis (Sell et al., 2001). All diets were formulated to meet or exceed the NRC (1994) nutrient recommendations and contained 1.0% Celite to increase the content of acid-insoluble ash (an indigestible marker). The AMEn contents of the experimental diets were not equalized. The basal diet was mixed in one large batch in a horizontal ribbon mixer without glucose·H2O or crude glycerol additions; the treatment diets were subsequently prepared by mixing a portion of the basal diet with the specified amounts of glucose · H2O and crude glycerol in a Hobart mixer (model H-600; Hobart, Troy, OH). The diets were fed in mash form.


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  		Table 1. Composition of the basal diet (as-is basis)
 

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  		Table 2. Characterization of the crude glycerol1 fed to laying hens (as-is basis)
 
Egg production was recorded daily and the feed consumption was determined for the 10-d experiment. Eggs collected over a 48-h period on d 7 and 8 of the experiment were weighed, and egg mass was calculated as egg production x egg weight. After 1 wk of adaptation to the experimental diets, excreta was collected twice daily for 3 d and stored at –20°C until analysis.

Excreta samples were pooled within cage, freeze-dried, and allowed to equilibrate with room moisture prior to analysis. The moisture contents of the experimental diets and freeze-dried excreta were determined in duplicate by drying at 135°C for 3 h. Diet and excreta N were determined in duplicate using the microKjeldahl method on a Kjeltech 1028 distilling unit (US Tecator Inc., Herndon, PA). The gross energy (GE) contents of the experimental diets and the excreta were determined in duplicate using an adiabatic bomb calorimeter (model 1281, Parr Instrument Company, Moline, IL). In addition, the GE contents of the crude glycerol and of chemically pure (≥99%) glycerol (catalog number G5516; Sigma-Aldrich, St. Louis, MO) were determined in triplicate. The contents of acid-insoluble ash in the experimental diets and the excreta were analyzed in triplicate according to procedures by Vogtmann et al. (1975).

The AMEn content of the crude glycerol was estimated by a linear regression equation relating the experimental-diet AMEn values to the proportion of crude glycerol in each diet (Leeson and Summers, 2001; Sell et al., 2001). The contribution of AMEn from glucose·H2O in all diets, calculated from the glucose percentage inclusion rate and the glucose AMEn value (3,640 kcal/kg), was subtracted from glucose-containing diets (Hill and Anderson, 1958; Sell et al., 2001). The AMEn value of the experimental diet was calculated according to the method listed by Leeson and Summers (2001) as


Formula

where AMEn (kcal/kg) = N-corrected apparent metabo-lizable energy content of the diet; GEDiet and GEExcreta (kcal/kg) = GE of the diet and excreta, respectively; AIA-Diet and AIAExcreta (%) = acid insoluble ash in the diet and excreta, respectively; 8.22 (kcal/kg) = energy value of uric acid; and NRetained (g/kg) is the N retained by the hens per kilogram of diet consumed. The retained N was calculated as


Formula

where NDiet and NExcreta (%) = N contents of the diet and excreta, respectively.

The experimental design was a completely randomized design with 4 dietary treatments and 6 replications per treatment (Morris, 1999). The cage containing 2 hens was the experimental unit. The AMEn value of the crude glycerol was estimated as the slope of the linear relationship between the inclusion rate of dietary crude glycerol (independent variable) and the glucose-corrected AMEn value of the experimental diet (dependent variable) using JMP 6.0.3 (SAS Institute Inc., Cary, NC). The effects of dietary crude glycerol on egg production, egg weight, egg mass, and feed consumption were analyzed by ANOVA using JMP. The ANOVA model included only the effects of dietary crude glycerol content, and treatment means were separated using linear, quadratic, and cubic orthogonal polynomial contrasts (Morris, 1999). The GE value of crude glycerol used in the study was compared with its AMEn value using a 2-tailed t-test with n = 3 for GE values and n = 24 for AMEn values (Snedecor, 1946). Probability values less than or equal to 0.05 were considered significant. Where appropriate, means and associated SEM are reported in the text on an as-is basis.


   RESULTS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
The mean BW of the hens was 1.37 ± 0.01 kg (n = 24) at the start of the experiment with no significant difference among treatments (P = 0.60). The increases in dietary AMEn values attributed to substitution of crude glycerol for glucose H2O increased linearly with increasing crude glycerol content (P < 0.001); there were no quadratic or cubic effects (P > 0.1). The AMEn value of the crude glycerol tested was 3,805 ± 238 kcal/kg (Figure 1Go) and was not different (P > 0.1) from its GE value (3,625 ± 26 kcal/ kg) as revealed by a 2-tailed t-test. Feed consumption (104 ± 4 g/d), egg production (93.0 ± 2.6%), egg weight (56.1 ± 0.9 g), or egg mass (52.2 ± 1.9 g/d) were not affected (P > 0.1) by the dietary treatments in the 10-d experiment. The GE value of chemically pure (≥99%) glycerol was 4,305 ± 30 kcal/kg.


Figure 1
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  		Figure 1. Relationship between the dietary inclusion rate of crude glycerol and the glucose-corrected AMEn value of the experimental diets for laying hens (each dot represents data from 1 cage containing 2 hens).

 

   DISCUSSION
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
The GE content of the crude glycerol used in the present experiment (3,625 kcal/kg) was similar to the 3,596 kcal/ kg reported by Cerrate et al. (2006), who calculated the ME content as 98% of the analyzed GE content. In the present experiment, the AMEn value of crude glycerol was found to be 3,805 kcal/kg and was not different from its GE value. Rosebrough et al. (1980) used a ME value of 4,200 kcal/kg for glycerol in a turkey hen experiment, but did not specify the chemical purity or origin of the glycerol used. In comparison, the NRC (1994) lists a MEn content of corn grain of 3,350 kcal/kg and that of crude soybean oil between 8,020 to 8,650 kcal/kg. Hence, crude glycerol is estimated to contain about 14% more MEn than that of corn grain, and less than half of that contained in crude soybean oil. Moreover, crude glycerol supplies only energy and not nutrients (e.g., amino acids, phosphorus, and essential fatty acids) as do corn grain and soybean oil, which should be considered when assigning a monetary value to crude glycerol. The chemically pure sample of glycerol contained 4,305 kcal/kg of GE, suggesting that the AMEn content of crude glycerol may be a direct function of its glycerol content.

The crude glycerol tested in the present experiment had a relatively high content of Na and—because the Na content was not equalized among the treatment diets—the excreta from hens fed the 15% crude glycerol diet was considerably wetter than that from other treatments. Moreover, crude glycerol is a viscous liquid, and flow characteristics of diets containing 10 and 15% crude glycerol were noticeably poorer than that of the control diet, an effect also observed by Kijora et al. (1995) and Cerrate et al. (2006). Based on growth performance, Simon et al. (1996), Kijora et al. (1995), and Cerrate et al. (2006) recommended feeding glycerol at 5 to 10% of the diet for broiler chickens and pigs, respectively, which is consistent with the observations of physical effects on the feed from the present experiment. The results of this study show that the energy in crude glycerol is used efficiently by laying hens and has an AMEn content of 3,805 kcal/kg (as-is basis), 14% higher than that of corn grain.


   ACKNOWLEDGMENTS
 
This project was supported by the Hatch Act, State of Iowa funds, and USDA-ARS. The authors acknowledge the in-kind donation of the crude glycerol (Ag Processing Inc., Sergeant Bluff, IA), laying hens (Southwest Iowa Eggs, Massena, IA), vegetable oil (Feed Energy Company, Des Moines, IA), calcium carbonate (ILC Resources, Des Moines, IA), vitamins (DSM Nutrition, Ames, IA), and meat and bone meal (Darling International, Des Moines, IA) used in this study. Moreover, the assistance provided by personnel in the Bregendahl and Kerr laboratories and at the Iowa State University Poultry Science Research Center is greatly appreciated.


   FOOTNOTES
 
1 Mention of a trade name, proprietary product, or specific equipment does not constitute a guarantee or warranty by the USDA or Iowa State University and does not imply approval to the exclusion of other products that may be suitable. Back

Received for publication June 18, 2007. Accepted for publication October 4, 2007.


   REFERENCES
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 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
American Oil Chemists’ Society. 2000. Official methods and recommended practices of the AOCS. 5th ed. Am. Oil Chem. Soc., Champaign, IL.

American Society for Testing and Materials. 2006. Annual book of American Society for Testing and Materials Standards International, Volume 05.04, Petroleum products and lubricants (IV): D6557—Latest. ATSM Int., West Conchohocken, PA.

Association of Official Analytical Chemists. 1990. Official Methods of Analysis. 15th ed. AOAC, Arlington, VA.

Brisson, D., M.-C. Vohl, J. St-Pierre, T. J. Hudson, and D. Gaudet. 2001. Glycerol: A neglected variable in metabolic processes? Bioessays 23:534–542.[CrossRef][Web of Science][Medline]

Cerrate, S., F. Yan, Z. Wang, C. Coto, P. Sacakli, and P. W. Waldroup. 2006. Evaluation of glycerine from biodiesel production as a feed ingredient for broilers. Int. J. Poult. Sci. 5:1001–1007.

Hill, F. W., and L. Anderson. 1958. Comparison of metabolizable energy and productive energy determinations with growing chicks. J. Nutr. 64:587–603.[Abstract/Free Full Text]

Kijora, C., H. Bergner, R.-D. Kupsch, and L. Hagemann. 1995. Glycerol as a feed component in diets of fattening pigs. Arch. Anim. Nutr. 47:345–360.[CrossRef][Web of Science]

Leeson, S., and J. Summers. 2001. Nutrition of the chicken. 4th ed. Univ. Books, Guelph, Ontario, Canada.

Lin, E. C. C. 1977. Glycerol utilization and its regulation in mammals. Annu. Rev. Biochem. 46:765–795.[CrossRef][Web of Science][Medline]

Ma, F., and M. A. Hanna. 1999. Biodiesel production: A review. Bioresour. Technol. 70:1–15.[CrossRef][Web of Science]

Morris, T. R. 1999. Experimental design and analysis in animal sciences. CABI Publ., Oxon, UK.

National Biodiesel Board. 2007. Estimated US biodiesel sales. http://www.biodiesel.org/pdf_files/fuelfactsheets/Biodiesel_Sales_Graph.pdf Accessed May 23, 2007.

NRC. 1994. Nutrient requirements of poultry. 9th ed. Natl. Acad. Press, Washington, DC.

Rosebrough, R. W., E. Geis, P. James, H. Ota, and J. Whitehead. 1980. Effects of dietary energy substitutions on reproductive performance, feed efficiency, and lipogenic enzyme activity on large white turkey hens. Poult. Sci. 59:1485–1492.[Web of Science]

Sell, J. L., S. Jin, and M. Jeffrey. 2001. Metabolizable energy value of conjugated linoleic acid for broiler chicks and laying hens. Poult. Sci. 80:209–214.[Abstract/Free Full Text]

Simon, A., H. Bergener, and M. Schwabe. 1996. Glycerol—feed ingredient for broiler chickens. Arch. Anim. Nutr. 49:103–112.[CrossRef][Web of Science]

Snedecor, G. W. 1946. Statistical Methods. 4th ed. Iowa State Coll. Press, Ames, IA.

Tao, R. C., R. E. Kelley, N. N. Yoshimura, and F. Benjamin. 1983. Glycerol: Its metabolism and use as an intravenous energy source. J. Parenter. Enteral Nutr. 7:479–488.[Free Full Text]

Thompson, J. C., and B. B. He. 2006. Characterization of crude glycerol from biodiesel production from multiple feedstocks. Appl. Eng. Agric. 22:261–265.[Web of Science]

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Vogtmann, H., H. P. Pfirter, and A. L. Prabucki. 1975. A new method of determining metabolisability of energy and digestibility of fatty acids in broiler diets. Br. J. Poult. Sci. 16:531–534.[CrossRef]




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