Your SlideShare is downloading. ×
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Efresh
Upcoming SlideShare
Loading in...5
×

Thanks for flagging this SlideShare!

Oops! An error has occurred.

×
Saving this for later? Get the SlideShare app to save on your phone or tablet. Read anywhere, anytime – even offline.
Text the download link to your phone
Standard text messaging rates apply

Efresh

448

Published on

Published in: Education, Technology
0 Comments
0 Likes
Statistics
Notes
  • Be the first to comment

  • Be the first to like this

No Downloads
Views
Total Views
448
On Slideshare
0
From Embeds
0
Number of Embeds
0
Actions
Shares
0
Downloads
6
Comments
0
Likes
0
Embeds 0
No embeds

Report content
Flagged as inappropriate Flag as inappropriate
Flag as inappropriate

Select your reason for flagging this presentation as inappropriate.

Cancel
No notes for slide

Transcript

  • 1. THE USE OF ELECTRICAL IMPEDANCE TO RAPIDLY PREDICT BEEF TENDERNESS. BY TIMOTHY MARK NATH A thesis submitted in partial fulfillment of the requirements for the Master of Science Major in Animal Science South Dakota State University 2008
  • 2. ii THE USE OF ELECTRICAL IMPEDANCE TO RAPIDLY PREDICT BEEF TENDERNESS. This thesis is approved as a creditable and independent investigation by a candidate for the Master of Science degree and is acceptable for meeting the thesis requirements for this degree. Acceptance of this thesis does not imply that the conclusions reached by the candidate are necessarily the conclusions of the major department. Dr. Duane Wulf Thesis Advisor Date Dr. Robert Thaler Head, Animal & Range Sciences Date
  • 3. iii Acknowledgements I would like to thank several people for their help throughout graduate school and the completion of this thesis. First of all, I am sincerely grateful for all the help and advice I received from my advisor Dr. Wulf. I appreciate all the time and effort Dr. Wulf put into teaching and guiding me through my research. I would also like to thank him for allowing me to work in the meat lab. The skills I have learned working in the meat lab will help guide me through my career. I would also like to thank Dr. Weaver for all of her advice and support. She has been great help with my lab work and the writing of this thesis. I would like to thank Dr. Thaler and Dr. Cartrette for serving on my committee. I would also like to thank Michaeal Singer for providing the electrical impedance instrument and his support through my research. I would like to especially thank Deon Simon for all her technical help with lab work. A huge thank you goes to Adam Rhody and his capable meat lab crew for all their help with shear force for my project. I can’t thank the meat science graduate students enough for their help, but a very sincere thank you goes to Tanner Machado, Dustin Mohrhauser, Sarah Wells, Andrew Everts, and Amanda Everts. I would also like to thank my parents Mike and Janet Nath and the rest of my family for being supportive and encouraging me throughout my education experience. Most of all I would like to thank my wife Carissa for her help and support through my graduate career. To our beautiful daughter Madison for giving me even more reasons to enjoy life. Lastly, I would like to thank God for allowing me to live my life each and every day.
  • 4. iv ABSTRACT THE USE OF ELECTRICAL IMPEDANCE TO RAPIDLY PREDICT BEEF TENDERNESS. Timothy M. Nath December 2008 A three-phase study was conducted to determine the usefulness of electrical impedance to rapidly predict beef tenderness. Objectives of phase I were to determine optimal measurement location and impedance’s potential to predict tenderness. Measurements of the ribeye were the most accurate at predicting tenderness and impedance had potential to predict beef tenderness. Objective of phase II was to compare impedance to existing technology (BeefCam and NIR) to predict beef tenderness. Impedance was the most effective technology at sorting out very tender carcasses. Instruments were additive when combined to predict beef tenderness. Objectives of phase III were to determine the effect of breed type, suspension method, and aging on impedance measurements. Impedance decreased slightly from d 1 to d 7 postmortem. Impedance was related to sarcomere length, proteolysis and Warner-Bratzler shear force. In conclusion, electrical impedance was weakly related to beef tenderness, but may be useful to sort out very tender beef carcasses and predict postmortem aging.
  • 5. v Table of Contents Page Abstract………………………………………………………………………..................iv List of Tables……………...………………………………………………………….….vii List of Figures……………...………………………………………………………….…ix Appendixes……………………………………………………………………………….x Chapter 1. Review of Literature Introduction…………………………………………………………………….....1 Beef Tenderness…………………………………………………………………...3 Proteolysis …………………………………….…………………………..6 Sarcomere Length …………………………….……………………….…..7 Connective Tissue………………………….………………………….…...8 Tenderness Prediction……………………………………………………………..9 Tendertec…………………………………………………………………10 BeefCam...…………………………………….………….………………11 NIR………………………………………………….………………........12 Slice Shear Force………………………………………………………...12 Electrical Impedance… …………………………….…………………..13 Literature Cited…………………………………………………………………..15
  • 6. vi Chapter 2. The use of electrical impedance to rapidly predict beef tenderness Introduction…………………………………………………….............................20 Materials and Methods……………………………………………………………22 Results and Discussion……………………………………………………....…....34 Implications………………………………………………………………...……..43 Literature Cited…………………………………………………………...………64
  • 7. vii List of Tables Table Page 2.1 Simple statistics for carcass and muscle traits for Phase I………………… ……44 2.2 The ability of electrical impedance measurements from various electrode depth and placement combinations to predict Warner-Bratzler shear force (average of d4 and d14)…...……………………….……….…......……………...45 2.3 Evaluating the ability of electrical impedance from tagged (carcass side sample was removed) and untagged (carcass side sample was not removed) sides to predict Warner-Bratzler shear force (average of d5 and d14)……....…..46 2.4 Predicting d5 and d14 Warner-Bratzler shear force using electrical impedance measurements from d4, d5, d14……………………...……………...47 2.5 R-squares for various prediction models to predict Warner-Bratzler shear force (d14)…………………...………..…………………………….…......48 2.6 Simple statistics for carcass and muscle traits for Phase II……………………...49 2.7 R-squares for various prediction models to predict Warner-Bratzler shear force (d14)....................................................................................................50 2.8 Percentage of tough (>49.0 N) carcasses by certification rate and instrument………………………………………………………………………..51 2.9 Percentage of very tender (<34.3 N) carcasses by certification rate and instrument………………………………………………………………………..52 2.10 Average Warner-Bratzler shear force (N) for sort groups (fifths) by instrument………………………………………………………………………..53
  • 8. viii 2.11 Effect of day postmortem on impedance measurements (n = 100)….………......54 2.12 Simple statistics for carcass and muscle traits for Phase III…………...………..55 2.13 Least square means of Warner-Bratzler shear force, sarcomere length, troponin T degradation, total collagen, resistance, reactance, and phase angle (averaged across all aging periods)..............................................................56 2.14 Effect of day on Warner-Bratzler shear force, sarcomere length, and troponin T degradation………….………………………………..…………...…57 2.15 Correlations of Warner-Bratzler shear force, sarcomere length, proteolysis, total collagen, resistance, reactance, and phase angle (*P < 0.05)………….......58
  • 9. ix List of Figures Figure Page 2.1 Electrode arrangement and penetrating depths…………………………………..59 2.2 The effect of breed type on resistance, reactance, and phase angle from 1 to 7 days postmortem (*P < 0.05)……......................................…….….....…...60 2.3 The effect of suspension on resistance, reactance, and phase angle from 1 to 7 days postmortem (*P < 0.05) ………………………..….…...……………61 2.4 Representative micrographs of myofibrils with normal (1.82 µm, left) and hip suspension (2.43 µm, right) sarcomeres of longissimus muscle (Magnification 400X)…………………………………………………………...62 2.5 Representative western blot of whole muscle protein extracts from bovine longissimus muscle with normal (NS) and hip (HS) suspension and postmortem aging periods were 1, 4, 7, and 10 days (d0 was used for quantification)……………………………………………………………...….....63
  • 10. x Appendix Appendix Page A. The effect of breed type on resistance, reactance, and phase angle impedance measurements of longissimus lumborum steaks from d 1 to d 21 postmortem.....67 B. The effect of suspension on resistance, reactance, and phase angle impedance measurements of longissimus lumborum steaks from d 1 to d 21 postmortem….68 C. The effect of breed type on resistance, reactance, and phase angle impedance measurements of semitendinosus steaks from d 1 to d 10 postmortem………….69 D. The effect of suspension on resistance, reactance, and phase angle impedance measurements of semitendinosus steaks from d 1 to d 10 postmortem……...…..70 E. The effect of breed type on resistance, reactance, and phase angle impedance measurements of psoas major steaks from d 1 to d 10 postmortem……………..71 F. The effect of suspension on resistance, reactance, and phase angle impedance measurements of psoas major steaks from d 1 to d 10 postmortem……………..72
  • 11. 1 CHAPTER I Review of Literature Introduction Palatability of meat consists of three main components: flavor, juiciness, and tenderness. The National Beef Tenderness Survey reported tenderness is the single most important factor affecting consumers’ perception of taste (Morgan et al., 1991). However, tenderness is a leading cause of consumer dissatisfaction, due to the variation of tenderness and inability to predict this variation among steaks. Many factors affect meat tenderness including: genetics of the animal, gender, and physiological maturity (Wulf, Tatum, Green, Morgan, Golden, & Smith, 1996; Huff-Lonergan, Parrish, & Robson, 1995; Purslow, 1985). Additionally, postmortem muscle characteristics known to influence meat tenderness include: proteolysis of myofibril proteins, sarcomere length, and connective tissue (Koohmaraie, Kent, Shackelford, Veiseth, & Wheeler, 2002). These numerous factors affecting tenderness make this a difficult palatability trait to predict on-line at packing plant speeds. Currently, the beef industry segregates carcasses into palatability groups according to the amount of marbling in the ribeye and the percentage of ossification in the thoracic buttons. However, marbling has been reported to be a poor predictor of tenderness and can explain no more than 5% of the variation in palatability traits (Morgan, et al., 1991; Wheeler, Cundiff, & Koch, 1994). Consumers are dissatisfied when purchasing steaks from the same quality grade that vary greatly in tenderness.
  • 12. 2 Studies have shown that consumers can detect the difference in beef tenderness and are willing to pay a premium for guaranteed tender beef (Boleman et al., 1997; Shackelford, Wheeler, Meade, Reagan, Byrnes, & Koohmaraie, 2001; Miller, Carr, Ramsey, Crockett, & Hoover, 2001). The most accurate technique to predict tenderness and satisfy consumers is the Warner-Bratzler shear force method (Bratzler, 1932; Warner, 1952). However, this machine is not practical for use in the industry due to time constraints and cost. Many researchers have tried to develop a machine or method that will predict beef tenderness. However, most of these machines or methods are unable to provide sufficient information at commercial packing-plant chain speeds to predict beef tenderness accurately. Therefore the meat industry is looking for a simple, fast, noninvasive way to predict beef tenderness and sort carcasses into palatability groups (NCA, 1994). Providing a value associated with tenderness will provide an economic incentive for the beef industry to market beef more efficiently to consumers. Electrical impedance is a simple, fast, noninvasive technology currently used in the medical field that utilizes phase angle to predict the life expectancy of a person dying from a terminal illness such as cancer (Gupta et al., 2004). Phase angle is a calculated value from the two components of electrical impedance: resistance (related to water loss) and reactance (related to cell membrane breakdown) (Lukaski, 1996). As the value of phase angle decreases over time the life expectancy shortens for that person (Gupta et al., 2004). In terms of predicting beef tenderness, it is known that as a beef animal ages and proceeds through proteolysis there is a loss of water and a breakdown of cell membranes. Lepetit, Salè, Favier, and Dalle (2002) have shown that electrical impedance could be
  • 13. 3 related to meat aging. Therefore, the objective of this research was to determine the usefulness of electrical impedance to rapidly predict beef tenderness. Beef Tenderness Tenderness as defined by Takahashi (1996) is the sum of the total of the mechanical strength of skeletal muscle tissue and its weakening during post-mortem ageing of meat. The National Beef Tenderness Survey revealed tenderness is the single most important factor affecting consumers’ perception of taste (Morgan et al., 1991). However, tenderness is the leading cause of consumer dissatisfaction, due to the variation of tenderness and inability to measure this variation among steaks. Currently the beef industry segregates carcasses into palatability groups based on the amount of marbling in the ribeye and the ossification of the thoracic buttons. Morgan and associates revealed that the current USDA grading system does a poor job at segregating carcasses into tenderness groups (1991). In addition, Wheeler and associates have stated “marbling explained at most 5% of the variation in palatability traits,” (1994). These authors concluded that there is not a lack of tender steaks, but rather an inconsistency of tenderness among steaks with similar quality grades. Thus, the National Cattlemen’s Association (NCA, 1994) listed “development of an instrument or procedure that can adequately measure quality, cutability and tenderness in beef carcasses in modern packing plants” as a top priority of the beef industry. There is an incentive for the beef industry to market tender beef based upon the increase in branded beef programs and the
  • 14. 4 data that demonstrates a significant segment of consumers that are willing to pay a premium for guaranteed tender beef. Boleman and associates (1997) were the first to show that consumers can detect differences in beef tenderness and are willing to pay a premium for guaranteed tender beef. In this study, beef strip steaks of known shear force values were placed into three color-coded categories [2.27 to 3.58 kg (Red), 4.08 to 5.40 kg (White), and 5.90 to 7.21 kg (Blue)] with a premium of $1.10 per kg placed between each category. The color- coded streaks were offered to randomly-selected consumers and after sampling consumers purchased 94.6, 3.6, and 1.8 percent of the Red, White, and Blue steaks, respectively. In addition, Shackelford and associates (2001) reported that 50% of consumers would be willing to pay a $1.10 per kg premium for guaranteed tender USDA Select loin steaks. In a study conducted by Miller and associates (2001), 78% of consumers were willing to pay more for steaks that the retailer guaranteed as tender. These results indicate the importance to the beef industry to be able to segregate between tender and tough steaks and to market accordingly. Genetic differences in tenderness have been found both among (Crouse, Cundiff, Koch, Koohmaraie, & Seideman, 1989) and within (Shackelford, Koohmaraie, Cundiff, Gregory, Rohrer, & Savell, 1994) breeds of cattle. Wulf and associates (1996) reported genetic differences existing in tenderness within and between the Charolais and Limousin breeds. In addition, breed type greatly affects beef tenderness; where Brahman influenced cattle tend to create tougher meat due to a higher level of calpastatin than non-
  • 15. 5 Brahman influenced cattle (Whipple, Koohmaraie, Dikeman, Crouse, Hunt, & Klemm, 1990; O’Connor, Tatum, Wulf, Green, & Smith, 1997). A method for improving meat from Brahman influenced cattle would be aging the meat for longer periods of time postmortem (O’Connor et al., 1997). Physiological maturity of the animal affects tenderness in that as cattle mature, intramuscular collagen solubility decreases, resulting in tougher beef (Purslow, 1985). Results from Huff-Lonergan and associates (1995), reported older cattle to be tougher than younger cattle due to an increased level of connective tissue, and bulls to be tougher than steers due to an increased level of calpastatin. Animal’s genetics, physiological maturity, and gender are just a few of the factors that can affect meat tenderness ante- mortem. There are also many factors that affect meat tenderness postmortem. Research has shown the three main factors that affect meat tenderness postmortem are: proteolysis of myofibril proteins, sarcomere length, and connective tissue (Koohmaraie et al., 2002). Connective tissue content increases with animal age and is related to the workload for each muscle (Purslow, 1985). Shorter sarcomere length leads to an increase in toughness (Herring, Cassens, & Briskey, 1965; Hostetler, Landman, Link, & Fitzhugh, 1970). Proteolysis is the breakdown of proteins by calpains (Huff-Lonergan, Mitsuhashi, Beckman, Parrish, Olson, & Robson, 1996). Research has shown meat tenderizes when held at refrigerated temperatures during postmortem aging (Taylor, Geesink, Thompson, Koohmaraie, & Goll, 1995). Since sarcomere length and connective tissue do not change during normal refrigerated temperature, proteolysis of
  • 16. 6 myofibril proteins is the major mechanism behind the tenderization of meat during postmortem aging. Proteolysis Proteolysis is the degradation of structural proteins by cellular enzymes called proteases (Tornberg, 1996). These proteolytic enzymes do not degrade the major filaments of myosin and actin, but rather the structural proteins (titin, nebulin, desmin, tropomyosin, and troponin) that regulate the integrity of the sarcomere (Goll, 1991, Huff- Lonergan et al., 1996). Koohmaraie has shown that the degradation of these proteins improves tenderness by reducing the organization of the filamentous structure within the sarcomere (1996). The calpain protease system has been shown by many researchers to be responsible for the improved tenderness during postmortem storage (Goll 1991; Huff- Lonergan et al., 1996). The calpain system includes µ-calpain and m-calpain (Koohmaraie and Geesink, 2006). The calcium concentration required is different for each enzyme as µ-calpain requires micromolar calcium concentrations and m-calpain requires millimolar calcium concentrations (Goll, 1991). Calpains are responsible for the degradation of titin, nebulin, tropomyosin, and troponin (Goll, 1991). The protease µ-calpain is thought to be the primary protease responsible for the breakdown of these proteins postmortem (Koohmaraie, 1996; Geesink, Kuchay, Chrishti, & Koohmaraie, 2006). Calpastatin is the specific inhibitor of µ-calpain and m-calpain, and thus inhibits the rate and extent of postmortem proteolysis (Geesink and Koohmaraie, 1999). Bos
  • 17. 7 indicus cattle have an increased calpastatin level and therefore do not go through this process of proteolysis as rapidly as Bos taurus cattle, and is one reason for the increased toughness in Bos indicus (Whipple et al., 1990). Shackelford and associates (1994) reported that the heritability of calpastatin activity was high for Bos indicus and accounted for much of the genetic variation in Warner-Bratzler shear force values. The effect of calpains has been well documented by many researchers to affect meat tenderness, but there may be other enzymes that could also affect proteolysis. The rate of proteolysis is highly variable and could be a factor in the inconsistency of beef tenderness, meaning that some beef may take a few days and others take weeks to complete proteolysis. Sarcomere Length A sarcomere is defined as a segment between two neighboring z-lines that is also the basic unit where muscle contraction and relaxation occur (Aberle, Forrest, Gerrard, & Mills, 2001). Many studies have shown that sarcomere length is correlated with tenderness, with a longer sarcomere having less resistance to shearing than a shorter sarcomere (Hostetler et al., 1970; Rhee, Wheeler, Shackelford, & Koohmaraie, 2004). During normal postmortem aging conditions, sarcomere length does not change due to the permanent formation of actomyosin cross-bridges which locks the thick and thin filaments in place (Savell, Mueller, & Baird, 2005). Sarcomere length can be affected by variations in hanging of the carcass pre-rigor to alter the sarcomere lengths of different muscles.
  • 18. 8 Hip suspension also known as “Tenderstretch” is a method of hanging the carcass from the aitch bone pre-rigor, allowing the weight of the hind limb to stretch muscles in the round and loin. Hip suspension is utilized to increase the sarcomere length of several muscles such as the longissimus and semitendinosus; however this method shortens sarcomeres in other muscles such as the psoas major. This method would improve the tenderness of the longissimus and semitendinosus muscles, but would cause detrimental effects on the already tender psoas major muscle (Hostetler et al., 1970). Tenderstretch is not practiced in commercial packing plants due the detrimental effects of the psoas major as well as space limitations within the plant. Tendercut™ is another prerigor treatment which subjects muscles to more tension by severing the bones, minor muscles, and connective tissues in the loin and(or) round area 45 to 90 min postmortem (Wang, Claus, & Marriot, 1996). This method increases the sarcomere length of the following muscles: longissimus, biceps femoris, semitendinosus, and semimembranosus, (Wang et al, 1996; Ludwig, Claus, Marriott, Johnson, & Wang, 1997; Beaty, Apple, Rakes, & Kreider, 1999). Inceasing the sarcomere length of these muscles would result in an improved tenderness. Tendercut™ could be used in the industry as this method requires no new equipment, and it is adaptable to the current design of existing plants. Connective Tissue Connective tissue is a measure of collagen cross-linking. The number of cross- linked chains explains a large amount of the tenderness variation, and varies greatly
  • 19. 9 between muscles, species, breeds, and with animal age (Judge & Aberle, 1982; Purslow, 1985; Lepetit, 2007). As collagen cross-linking increases with animal age, the tougher and less palatable the muscle becomes (Judge and Aberle, 1982). Unlike some myofibrillar proteins, collagen is not degraded postmortem which contributes to a fixed amount of background toughness. Perimysium is a sheath of connective tissue that groups individual muscle fibers into bundles. Perimysium remains intact during cooking and is the primary source of cooked meat toughness. Connective tissue content must be addressed either ante-mortem via genetics and physiological age at slaughter, or postmortem via cooking (Purslow, 1985). Tenderness Prediction Past surveys have revealed the inability of the current USDA beef quality grading system to accurately segregate carcasses into palatability groups (Morgan et al., 1991). Studies have shown that consumers can detect the difference in beef tenderness and are willing to pay a premium for guaranteed tender beef (Boleman et al., 1997; Miller et al., 2001; Shackelford et al., 2001). Thus, the National Cattlemen’s Association (NCA, 1994) listed “development of an instrument or procedure that can adequately measure quality, cutability and tenderness in beef carcasses in modern packing plants” as a top priority of the beef industry. There have been many attempts to identify instrumental methods for predicting meat tenderness. Most of these were intended for laboratory research tools and varied widely in their efficacies.
  • 20. 10 In more recent investigations of objective predictions of meat tenderness, the goal has been to develop on-line systems for grading carcasses based on tenderness. The ideal system would involve an objective, noninvasive, tamper-proof, accurate technology. Past technologies evaluated for their potential as on-line tenderness grading tools include Tendertec (Ferguson, 1993), BeefCam (Wyle, Cannell, Belk, Goldberg, Riffle, & Smith, 1998), near-infrared spectroscopy (Park, Chen, Hruschka, Shackelford, & Koohmaraie, 1998), and slice shear force (Shackelford, Wheeler, & Koohmaraie, 1999). Many instruments have been developed to predict tenderness, but few are as accurate as Warner-Bratzler shear force (Bratzler, 1932; Warner, 1952). Warner-Bratzler shear force is the most commonly used objective method to measure tenderness, but is costly, time consuming, and difficult to fit into industry operations because it must be done on cooked steaks. Therefore, the industry is looking for a machine that would provide sufficient information at commercial packing-plant chain speeds to accurately predict beef tenderness. Tendertec Initial studies performed by Ferguson (1993) showed promise for the Tendertec to predict beef tenderness. The Tendertec is an instrument equipped with a 14-cm piston that encountered two decleration stops occurring at 2 and 6 cm and an overall predetermined depth of 8 cm. The probe tip is inserted perpendicularly between the dorsal spinous processes of thoracic and lumbar vertebra through the multifidus dorsi and into the longissimus lumborum. George and associates (1997) reported that the Tendertec
  • 21. 11 probe detected some differences in connective tissue, however it was not better than USDA quality grade at segmenting A-maturity carcasses into tenderness categories. Belk and associates (2001) concluded that Tendertec was able to sort carcasses of older, mature cattle based on tenderness, but failed to consistently detect the differences in steaks derived from youthful carcasses. In addition, the Tendertec did not meet the demands of a noninvasive system as this machine would puncture holes into the meat. BeefCam Researchers have reported that objective color measurements of beef longissimus to be related to tenderness and can sort carcasses into palatability groups using lean color (Wulf, O’Connor, Tatum, & Smith, 1997). These findings led researchers at Colorado State University and Hunter Associates Laboratory (Reston, VA) to develop a prototype video imaging system (prototype BeefCam) that would be able predict beef tenderness. The prototype BeefCam instrument would quickly provide a noninvasive visual image of the ribeye. Pilot studies performed by Wyle and associates (1998) indicated that the prototype BeefCam instrument could quickly identify carcasses likely to produce steaks that were tender, based on Warner-Bratzler shear force values, after a 14 day aging period. However, the prototype BeefCam instrument did have limitations that prevented its use in a commercial setting. According to the National Beef Instrument Assessment Planning Symposium (NLSMB, 1994), for an instrument to be successful, it must be tested under real-world conditions.
  • 22. 12 Vote, Belk, Tatum, Scanga, & Smith (2003) reported that the Computer Vision System equipped with a BeefCam module was able to capture and segment video images at commercial packing-plant chain speeds and produce information useful in explaining observed variation in Warner-Bratzler shear force values of steaks. This information could be used to sort carcasses according to expected palatability differences, even in carcasses with similar marbling scores. BeefCam measurements could aid in the selection of tender or tough steaks and therefore improve consumer satisfaction. Near-Infrared Near-infrared reflectance (NIR) spectroscopy is a rapid, nondestructive system that gathers information from samples through measurements of reflected light. The light reflected back through the NIR system contains information about properties associated with meat tenderness (Park et al., 1998). Shackelford, Wheeler and Koohmaraie (2004) developed a commercially available tenderness prediction system based on visible and NIR spectroscopy that could be used on-line. Rust and associates (2008) reported that the NIR system was able to successfully sort tough from tender longissimus lumborum samples to 70% certification levels. These authors concluded that NIR scanning offers an in-plant opportunity to sort carcasses into tenderness outcome groups and more importantly guaranteed-tendered branded beef programs (Rust et al., 2008). Slice Shear Force Shackelford and associates (1999) developed a system for classifying beef tenderness based on a rapid, simple method of measuring cooked longissimus shear force.
  • 23. 13 Longissimus steaks (2.54 cm thick) were trimmed free of subcutaneous fat and bone then rapidly cooked using a belt grill. A 1-cm-thick, 5-cm-long slice was removed from the cooked longissimus parallel with the muscle fibers to measure shear force. Slices were sheared with a flat, blunt-end blade using an electronic testing machine. The entire process was completed in less than 10 min. Therefore, in commercial application, this process could be completed during the 10- to 15-min period that carcasses are normally held to allow the ribeye to bloom for quality grading. Wheeler and associates (2002) concluded that slice shear force, not a prototype BeefCam or colorimeter systems, accurately identified “tender” beef. However, the industry is reluctant to implement this system because of cost and a loss of product. Electrical Impedance Electrical impedance is a simple, fast, noninvasive technology currently used in the medical field that utilizes phase angle to predict the life expectancy of a person dying from a terminal illness such as cancer (Gupta et al., 2004). According to Foster and Lukaski (1996), electrical impedance is measured by introducing a small alternating current into the body and measuring the potential difference that results. Electrical impedance utilizes two components, resistance and reactance. Resistance is the pure opposition of a biological conductor to the flow of an alternating current. Reactance is the voltage stored by a condenser for a brief period of time. Phase angle is a calculated value from resistance and reactance (Lukaski, 1996). As the value of phase angle decreases, the life expectancy shortens for that person (Gupta et al., 2004).
  • 24. 14 In terms of predicting beef tenderness, it is known that as a beef animal ages and proceeds through proteolysis, there is a change from bound water to free water and a breakdown of membranes. Past researchers have shown that electrical impedance could be related to tenderness (Byrne, Troy, & Buckley, 2000). Lepetit and associates (2002) showed a decrease in electrical impedance values during postmortem aging and concluded that electrical impedance could be related to meat aging. Based on these results, we hypothesized that electrical impedance will predict beef tenderness through the resistance, reactance, and phase angle. This research was conducted in three phases and the objectives for each phase were to: (1) determine optimal electrode arrangement and anatomical measurement location and determine if electrical impedance had potential to predict beef tenderness, (2) compare electrical impedance to existing technology (BeefCam and NIR) to predict beef tenderness, and (3) determine the effect of breed type (Bos taurus vs. Bos indicus), suspension method (traditional vs. hip), and postmortem aging on electrical impedance measurements.
  • 25. 15 Literature Cited Aberle, E. D., J. C. Forrest, D. E. Gerrard, and E. W. Mills. (2001). Principles of Meat Science. 4th ed. Kendall/Hunt Dubuque, IA. pp. 13-14. Beaty, S.L., J.K. Apple, L. K. Rakes, and D. L. Kreider. (1999). Early postmortem skeletal alterations effect on sarcomere length, myofibrillar fragmentation, and muscle tenderness of beef from light-weight, Brangus heifers. Journal of Muscle Foods, 10, 67-78. Belk, K. E., M. H. George, J. D. Tatum, G. G. Hilton, R. K. Miller, M. Koohmaraie, J. O. Reagan, and G. C. Smith. (2001). Evaluation of the Tendertec beef grading instrument to predict the tenderness of steaks from beef carcasses. Journal of Animal Science, 79, 688-697. Boleman, S. J., S. L. Boleman, R. K. Miller, J. F. Taylor, H. R. Cross, T. L. Wheeler, M. Koohmaraie, S. D. Shackelford, M. F. Miller, R. L. West, D. D. Johnson, and J. W. Savell. (1997). Consumer evaluation of beef of known categories of tenderness. Journal of Animal Science, 75, 1521–1524. Bratzler, L. J. (1932). Measuring the tenderness of meat by mechanical shear. M.S. Thesis, Kansas State College, Manhattan. Byrne, C. E., D. J. Troy, and D. J. Buckley. (2000). Postmortem changes in muscle electrical properties of bovine M. longissimus dorsi and their relationship to meat quality attributes and pH fall. Meat Science, 54, 23-34. Crouse, J. D., L. V. Cundiff, R. M. Koch, M. Koohmaraie and S. C. Seideman. (1989). Comparisons of Bos indicus and Bos taurus inheritance for carcass beef characteristics and meat palatability. Journal of Animal Science, 67, 2661-2668. Ferguson, D. M. (1993). Objective evaluation of meat-quality characteristics. In: Proceedings Australian Meat Industry Reciprocal Conference Gold Coast, QLD. pp 1-8. Foster, K. R. and H. C. Lukaski. (1996). Whole-body impedance – what does it measure? The American Journal of Clinical Nutrition, 64, 388S-396S. Geesink, G. H. and M. Koohmaraie. (1999). Technical note: a rapid method for quantification of calpain and calpastatin activities in muscle. Journal of Animal Science, 77, 3225-3229.
  • 26. 16 Geesink, G. H., S. Kuchay, A. H. Chrishti, and M. Koohmaraie. (2006). µ-Calpain is essential for postmortem proteolysis of muscle proteins. Journal of Animal Science, 84, 2834-2840. George, M. H., J. D. Tatum, H. G. Dolezal, J. B. Morgan, J. W. Wise, C. R. Calkins, T. Gordon, J. O. Reagan, and G. C. Smith. (1997). Comparison of USDA quality grade with Tendertec for the assessment of beef palatability. Journal of Animal Science 75, 1538–1546. Goll, D. E. (1991). Role of proteinases and protein turnover in muscle growth and meat quality. Proceedings Reciprocal Meat Conference, 44, 25-36. Gupta, D., C. A. Lammersfeld, J. L. Burrows, S. L. Dahlk, P. G. Vashi, J. F. Grutsch, S. Hoffman, and C. G. Lis. (2004). Bioelectrical impedance phase angle in clinical practice: implications for prognosis in advanced colorectal cancer. The American Journal of Clinical Nutrition, 80, 1634-1638. Herring, H. K., R. G. Cassens, and E. J. Briskey. (1965). Further studies on bovine muscle tenderness as influenced by carcass position, sarcomere length, and fiber diameter. Journal of Food Science, 30, 1049-1054. Hostetler, R. L., W. A. Landmann, B. A., Link, and H. A. Fitzhugh, Jr. (1970). Influence of carcass position during rigor mortis on tenderness of beef muscles: Comparison of two treatments. Journal of Animal Science, 31, 47-50. Huff-Lonergan, E., F. C. Parrish Jr., and R. M. Robson. (1995). Effects of postmortem aging time, animal age, and sex on degradation of titin and nebulin in bovine longissimus muscle. Journal of Animal Science, 73, 1064-1073. Huff-Lonergan. E., T. Mituhashi, D. D. Beckman, F. C. Parrish Jr., D. G. Olson, and R. M. Robson. (1996). Proteolysis of specific muscle structural proteins by µ-calpain at low pH and temperature is similar to degradation in postmortem bovine muscle. Journal of Animal Science, 74, 993-1008. Judge, M.D. and E. D. Aberle. (1982). Effects of chronological age and postmortem aging on thermal shrinkage temperature of bovine intramuscular collagen. Journal of Animal Science, 54, 68-71. Koohmaraie, M. (1996). Biochemical factors regulating the toughening and tenderization processes of meat. Meat Science, 43, S193-S201. Koohmaraie, M., M. P. Kent, S. D. Shackelford, E. Veiseth, and T. L. Wheeler. (2002). Meat tenderness and muscle growth: Is there any relationship? Meat Science, 62, 345-352.
  • 27. 17 Koohmaraie, M. and G. H. Geesink. (2006). Contribution of postmortem muscle biochemistry to the delivery of consistent meat quality with particular focus on the calpain system. Meat Science, 74, 34-43. Lepetit, J., P. Salè, R. Favier, and R. Dalle. (2002). Electrical impedance and tenderization in bovine meat. Meat Science, 60, 51-62. Lepetit, J. (2007). A theoretical approach of the relationship between collagen content, collage cross-links and meat tenderness. Meat Science, 76, 147-159. Ludwig, C. J., J. R. Claus, N. G. Marriott, J. Johnson, and H. Wang. (1997). Skeletal alterations to improve beef longissimus muscle tenderness. Journal of Animal Science, 75, 2404-2410. Lukaski, H. C. (1996). Biological indexes considered in the derivation of the bioelectrical impedance analysis. American Journal of Clinical Nutrition, 64, 397S-404S. Miller, M. F., M. A. Carr, C. B. Ramsey, K. L. Crockett, and L. C. Hoover. (2001). Consumer thresholds for establishing the value of beef tenderness. Journal of Animal Science, 79, 3062–3068. Morgan, J. B., J. W. Savell, D. S. Hale, R. K. Miller, D. B. Griffin, H. R. Cross, and S. D. Shackelford. (1991). National Beef Tenderness Survey. Journal of Animal Science, 69, 3274-3283. NCA. (1994). National Beef Tenderness Conference: Executive Summary. National Cattlemen’s Association, Englewood, CO. NLSMB. (1994). National beef instrument assessment plan. National Live Stock and Meat Board, Chicago, IL. O’Connor, S. F., J. D. Tatum, D. M. Wulf, R. D. Green, and G. C. Smith. (1997). Genetic effects on beef tenderness in Bos indicus composite and Bos taurus cattle. Journal of Animal Science, 75, 1822-1830. Park, B., Y. R. Chen, W. R. Hruschka, S. D. Shackelford, and M. Koohmaraie. (1998). Near-infrared reflectance analysis for predicting beef longissimus tenderness. Journal of Animal Science, 76, 2115-2120. Purslow, P. P. (1985). The physical basis of meat texture: Observations on the fracture behaviour of cooked bovine M. Semitendinosus. Meat Science, 12, 39-60.
  • 28. 18 Rhee, M. S., T. L. Wheeler, S. D. Shackelford, and M. Koohmaraie. (2004). Variation in palatability and biochemical traits within and among eleven beef muscles. Journal of Animal Science, 82, 534-550. Rust, S. R., D. M. Price, J. Subbiah, G. Kranzler, G. G. Hiton, D. L. Vanoverbeke, and J. B. Morgan. (2008). Predicting beef tenderness using near-infrared spectroscopy. Journal of Animal Science, 86, 211-219. Savell. J. W., S. L. Mueller, and B. E. Baird. (2005). The chilling of carcasses. Meat Science, 70, 449-459. Shackelford, S. D., M. Koohmaraie, L. V. Cundiff, K. E. Gregory, G. A. Rohrer, and J. W. Savell. (1994). Heritabilities and phenotypic and genetic correlations for bovine postrigor calpastatin activity, intramuscular fat content, Warner-Bratzler shear force, retail product yield, and growth rate. Journal of Animal Science, 72, 857-863. Shackelford, S. D., T. L.Wheeler, and M. Koohmaraie. (1999). Evaluation of slice shear force as an objective method of assessing beef longissimus tenderness. Journal of Animal Science, 77, 2693–2699. Shackelford, S. D., T. L. Wheeler, M. K. Meade, J. O. Reagan, B. L. Byrnes, and M. Koohmaraie. (2001). Consumer impressions of Tender Select beef. Journal of Animal Science, 79, 2605-2614. Shackelford, S. D., T. L. Wheeler, and M. Koohmaraie. (2004). Development of optimal protocol for visible and near-infrared reflectance spectroscopic evaluation of meat quality. Meat Science, 68, 371-381. Takahashi K. (1996). Structural weakening of skeletal muscle tissue during post- mortem ageing of meat: The non-enzymatic mechanism of meat tenderization. Meat Science, 43, S67-S80. Taylor, R. G., G. H. Geesink, V. F. Thompson, M. Koohmaraie, and D. E. Goll. (1995). Is z-disk degradation responsible for postmortem tenderization? Journal of Animal Science, 73, 1351-1367. Tornberg, E. (1996). Biophysical aspects of meat tenderness. Meat Science, 43, S175- S191. Vote, D. J., K. E. Belk, J. D. Tatum, J. A. Scanga, and G. C. Smith. (2003). Online prediction of beef tenderness using a computer vision system equipped with a BeefCam module. Journal of Animal Science, 81, 457-465.
  • 29. 19 Wang, H., J. R. Claus, and N. G. Marriott. (1996). Prerigor treatment and endpoint temperature effects on U.S. Choice beef tenderness. Journal of Muscle Foods, 7, 45-54. Warner, K. F. (1952). Adventures in testing meat for tenderness. Proceedings Reciprocal Meat Conference, 5, 156–160. Wheeler, T. L., L. V. Cundiff, and R. M. Koch. (1994). Effect of marbling degree on beef palatability in Bos taurus and Bos indicus cattle. Journal of Animal Science, 72, 3145-3151. Wheeler, T. L., D. Vote, J. M. Leheska, S. D. Shackelford, K. E. Belk, D. M. Wulf, B. L. Gwartney, and M. Koohmaraie. (2002). The efficacy of three objective systems for identifying beef cuts that can be guaranteed tender. Journal of Animal Science, 80, 3315-3327. Whipple, G., M. Koohmaraie, M. E. Dikeman, J. D. Crouse, M. C. Hunt and R. D. Klemm. (1990). Evaluation of attributes that affect longissimus muscle tenderness in Bos taurus and Bos indicus cattle. Journal of Animal Science, 68, 2716-2728. Wulf, D. M., J. D. Tatum, R. D. Green, J. B. Morgan, B. L. Golden, and G. C. Smith. (1996). Genetic influences on beef longissimus palatability in Charolais- and Limousin-Sired steers and heifers. Journal of Animal Science, 74, 2394-2405. Wulf, D. M., S. F. O’Connor, J. D. Tatum, and G. C. Smith. (1997). Using objective measures of color to predict beef longissimus tenderness. Journal of Animal Science, 75, 684–692. Wulf, D. M. and J. K. Page. (2000). Using measurements of muscle color, pH, and electrical impedance to augment the current USDA beef quality grading standards and improve the accuracy and precision of sorting carcasses into palatability groups. Journal of Animal Science, 78, 2595–2607. Wyle, A. M., R. C. Cannell, K. E. Belk, M. Goldberg, R. Riffle, and G. C. Smith. (1998). An evaluation of the portable HunterLab Video Imaging System (BeefCam) as a tool to predict tenderness of beef carcasses using objective measures of lean and fat color. Final Report to the National. Cattlemen’s Beef Association, Englewood, CO. Department of Animal Science, Colorado State University, Fort Collins.
  • 30. 20 CHAPTER II THE USE OF ELECTRICAL IMPEDANCE TO RAPIDLY PREDICT BEEF TENDERNESS. Timothy M. Nath Department of Animal & Range Sciences South Dakota State University, Brookings 57007 Introduction Palatability of meat consists of three main components: flavor, juiciness, and tenderness. The National Beef Tenderness Survey revealed tenderness as the single most important factor affecting consumers’ perception of taste (Morgan et al., 1991). However, tenderness is a leading cause of consumer dissatisfaction, due to the variation of tenderness and inability to predict this variation among steaks. Currently, the beef industry segregates carcasses into palatability groups according to the amount of marbling in the ribeye and the physiology maturity of the carcass. However, marbling has been reported to be a poor predictor of tenderness and can explain approximately 5% of the variation in palatability traits (Morgan, et al., 1991; Wheeler, et al., 1994). Consumers are dissatisfied when purchasing steaks from the same quality grade that vary greatly in tenderness. Studies have shown that consumers can detect differences in beef tenderness and are willing to pay a premium for guaranteed tender beef (Boleman et al., 1997; Shackelford et al., 2001; Miller et al., 2001). Therefore, the meat industry is looking for a simple, fast, noninvasive way to predict beef tenderness and sort carcasses into palatability groups (NCA, 1994). Electrical impedance is a simple, fast, noninvasive
  • 31. 21 technology currently used in the medical field that utilizes phase angle to predict the life expectancy of a person dying from a terminal illness such as cancer (Gupta et al., 2004). Phase angle is a calculated value from the two components of electrical impedance: resistance (related to water loss) and reactance (related to cell membrane breakdown) (Lukaski, 1996). In terms of predicting beef tenderness, it is known that as postmortem beef muscle ages and proceeds through proteolysis there is a loss of water and a breakdown of cell membranes. Therefore, we hypothesized that electrical impedance will predict beef tenderness through the resistance, reactance, and phase angle.
  • 32. 22 Materials and Methods Description of the Instrument Electrical impedance was measured with a prototype instrument utilizing four electrodes in a linear arrangement (outside two electrodes = electrical source; middle two electrodes = detection) that measured reactance and resistance (Figure 2.1). Phase angle is calculated from reactance and resistance with the following equation: [Phase Angle = degrees(arctangent(reactance/resistance))]. Phase I Carcass Selection & Electrical Impedance Phase I was conducted in a commercial beef packing plant (Swift & Company, Greeley, Colorado). Electrical impedance was measured on the exposed longissimus muscle of 300 randomly-chosen beef carcasses to define population variation. The four electrical impedance electrodes were attached to four separate probes in a linear arrangement penetrated the exposed longissimus muscle 5 cm (Figure 2.1a). The same arrangement of electrodes and probe depth was used to select carcasses (n=92) to equally represent all quantiles of the population. These carcasses were then moved to a separate rail for further testing. Carcasses were measured with five different electrode depth and placement combinations. The first three combinations were all in a linear arrangement with penetrating depths of 5 cm, 1.3 cm, or 0 cm (Figure 2.1a,b,c). Another linear arrangement expanded 50 cm over the strip loin with the detection and electrical source probes 2.5 cm apart on each end, with each probe penetrating the strip loin 5 cm. The
  • 33. 23 last combination measured from strip to round with the detection and electrical source probes 2.5 cm apart with two probes in the strip loin and two probes in the round at 5 cm penetration. South Dakota State University (SDSU) personnel recorded hot carcass weight (HCW), longissimus muscle area (REA), adjusted fat thickness, estimated percentage of kidney, pelvic and heart fat (KPH), and USDA marbling score. Yield grades were calculated from adjusted fat thickness, HCW, REA, and KPH. In addition, camera carcass data were collected from the BeefCam computer system. After carcass data were collected and electrical impedance measured, a section of longissimus was excised, vacuum-packaged, packed in coolers with ice, and transported back to SDSU Meat Lab. Upon arrival at SDSU, two 2.5-cm thick steaks were removed from each sample. The first steak was utilized for d 5 measurements of electrical impedance and Warner-Bratzler shear force, and the second steak was utilized for d 14 measurements. Electrical impedance measurements on steaks were in a linear arrangement with penetrating depths of 1.3 cm or 0 cm (Figure 2.1b,c). Warner-Bratzler Shear Force Determination Fresh longissimus samples were vacuumed packaged and stored at 4°C until appropriate aging length (d 5 or d 14). Samples were removed from vacuum packages and cut into 2.5-cm-thick steaks. Steaks were cooked on a belt-fed impingement oven (Model 1132-000-A, Lincoln Foodservice Products, Inc., Fort Wayne, IN). Peak internal cooked temperature measurements were recorded for each steak using a hand held thermometer (model 39658-K, Atkins Technical, Gainesville, FL). Cooked steaks were cooled for 24 hours at 4°C before removing 6 to 8 cores (1.27 cm in diameter) parallel to
  • 34. 24 the muscle fiber orientation (AMSA, 1995). A single, peak shear force measurement was obtained for each core using a Warner-Bratzler shear force machine (G-R Electric Manufacturing Company, Manhattan, KS). Peak shear force values for each core were averaged to obtain Warner-Bratzler shear force value for each steak. Statistical Analysis Data were analyzed using the PROC REG procedure of SAS (SAS Inst. Inc., Cary, NC). Warner-Bratzler shear force was predicted using the following regressor variables; electrical impedance, marbling, carcass traits, and camera color using both simple linear regression analysis (single regressor variable) and multiple linear regression analysis (all combinations of regressor variables) (Meyers, 1990). Phase II Carcass Selection & Electrical Impedance Phase II was performed in a commercial beef packing plant (Swift & Company, Cactus, Texas). Electrical impedance was measured on the exposed longissimus muscle of 300 randomly-chosen beef carcasses to define population variation. The four electrodes were attached to four separate probes that were in a linear arrangement and penetrated the exposed longissimus muscle 5 cm (Figure 2.1a). The same arrangement of electrodes and probe depth was used to select carcasses (n=300) to equally represent all quantiles of the population. One hundred carcasses were selected three consecutive days for a total of 300 carcasses. These carcasses were then moved to a separate rail for
  • 35. 25 further testing. Electrical impedance measurements on carcasses were measured with either penetrating (1.3 cm) or surface (0 cm) probes in a linear arrangement (Figure 2.1b,c). Impedance measurements of carcasses 1 to100 were recorded on three consecutive days at the plant to determine the effect of measurement day on Tenderness prediction ability. Impedance measurements of carcasses 101 to 300 were measured only one day at the plant. SDSU personnel recorded hot carcass weight (HCW), longissimus muscle area (REA), adjusted fat thickness, estimated percentage of kidney, pelvic and heart fat (KPH), and USDA marbling score. Yield grades were calculated from adjusted fat thickness, HCW, REA, and KPH. In addition, camera carcass data were collected from the BeefCam and NIR computer systems. After carcass data were collected and electrical impedance measured, a section of longissimus was excised, vacuum-packaged, packed in coolers with ice, and transported back to SDSU Meat Lab. Upon arrival at SDSU, the samples were stored under refrigeration until d 14 postmortem. On d 14, a 2.5-cm thick steak was removed from each sample. Electrical impedance was measured and the steaks were cooked for Warner-Bratzler shear force analysis Electrical impedance measurements on steaks were in a linear arrangement with penetrating depths of 1.3 cm or 0 cm (Figure 2.1b,c). Warner-Bratzler Shear Force Determination Fresh longissimus samples were vacuumed packaged and stored at 4°C until appropriate aging length (d 14). Samples were removed from vacuum packages and cut into 2.5-cm-thick steaks. Steaks were cooked on a belt-fed impingement oven (Model
  • 36. 26 1132-000-A, Lincoln Foodservice Products, Inc., Fort Wayne, IN). Peak internal cooked temperature measurements were recorded for each steak using a hand held thermometer (model 39658-K, Atkins Technical, Gainesville, FL). Cooked steaks were cooled for 24 hours at 4°C before removing 6 to 8 cores (1.27 cm in diameter) parallel to the muscle fiber orientation (AMSA, 1995). A single, peak shear force measurement was obtained for each core using a Warner-Bratzler shear force machine (G-R Electric Manufacturing Company, Manhattan, KS). Peak shear force values for each core were averaged to obtain Warner-Bratzler shear force value for each steak. Statistical Analysis Data were analyzed using the PROC REG procedure of SAS. Warner-Bratzler shear force was predicted using the following regressor variables; carcass traits, camera color, BeefCam, NIR, and electrical impedance using both simple linear regression analysis (single regressor variable) and multiple linear regression analysis (all combinations of regressor variables). Accuracy was calculated of each instrument to certify carcasses as “not tough,” where error rate equals percent of those certified carcasses that were, in fact, tough (WBS > 49.0 N). Accuracy was calculated for each instrument to certify carcasses as “very tender,” where accuracy rate equals percent of those certified carcasses that were, in fact, very tender (WBS < 34.3 N). These accuracies were calculated for various certifications rates. Pearson’s correlation coefficients were generated using the PROC CORR procedure of SAS to examine effect of day on impedance measurements.
  • 37. 27 Phase III Carcass Selection & Electrical Impedance Phase III was performed at the SDSU Meat Lab and utilized 14 steer carcasses of Bos taurus (n=7) and Bos indicus (n=7) breed type. Samples were removed from the longissimus muscle approximately 30 min after death and frozen for protein degradation analysis. After splitting the carcass down the backbone, the right side of the carcass (n=14) was hung from the Achilles tendon (normal suspension) and the left side of the carcass (n=14) was hung from the aitch bone (hip suspension). The front and rear limbs of the hip suspension carcasses were tied with a rope and pulled together until the two limbs were parallel. Carcasses were then chilled for 24 hours at 4ºC. Following carcass chilling, SDSU personnel recorded hot carcass weight (HCW), longissimus muscle area (REA), adjusted fat thickness, estimated percentage of kidney, pelvic and heart fat (KPH), and USDA marbling score. Yield grades were calculated from adjusted fat thickness, HCW, REA, and KPH. A section of longissimus was excised, cut into 2.5-cm thick steaks, vacuum-packaged, aged for 1, 4, 7, or 10 d postmortem and then frozen. Warner-Bratzler shear force was measured at 1 and 10 d postmortem. Sarcomere length was measured at 1, 4, 7, or 10 d postmortem. Rate of proteolysis was evaluated by the degradation of troponin T (TnT), at 0, 1, 4, 7, and 10 d postmortem (day 0 samples collected at death was used to quantify the amount of TnT degradation that occurred from 1 to 10 d postmortem). Electrical impedance was
  • 38. 28 measured on carcasses at the exposed longissimus from 1 to 7 d postmortem, with either penetrating (1.3 cm) or surface (0 cm) probes in a linear arrangement (Figure 2.1b,c). Warner-Bratzler Shear Force Determination Fresh longissimus samples were cut into 2.5-cm-thick steaks, vacuumed packaged and stored at 4°C until appropriate aging length (d 1 or d 10) then frozen. Samples were thawed 24 hours at 4°C and cooked on an electric clam shell grill (George Forman Indoor/Outdoor Grill, Model GGR62, Lake Forest, IL). Peak internal cooked temperature measurements were recorded for each steak using a hand held thermometer (model 39658-K, Atkins Technical, Gainesville, FL). Cooked steaks were cooled for 24 hours at 4°C before removing 6 to 8 cores (1.27 cm in diameter) parallel to the muscle fiber orientation (AMSA, 1995). A single, peak shear force measurement was obtained for each core using a WBS force machine (G-R Electric Manufacturing Company, Manhattan, KS). Peak shear force values for each core were averaged to obtain Warner- Bratzler shear force value for each steak. Myofibril Preparation Upon completion of appropriate aging lengths (1, 4, 7, or 10 d postmortem), myofibrils were purified according to a modified procedure of Swartz, Greaser, & Marsh, (1993). Approximately 2 g of muscle were minced with a knife and homogenized in 35 ml of rigor buffer (RB, 75 mM KCl, 10 mM imidazole, 2 mM MgCl 2 , 2 mM EGTA, 1 mM NaN 3 , pH 7.2) for two 15 sec bursts using a Ultra-Turrax T25 homogenizer (Janke & Kunkel GmbH & Co. KG.) at medium speed. The suspension was centrifuged at 1,000
  • 39. 29 x g for 10 min at 4˚C. Supernatant was decanted and the remaining pellet was homogenized in 35 ml of RB for two 15 sec bursts at medium speed. The suspension was centrifuged again at 1,000 x g for 10 min. Again the supernatant was decanted and the remaining pellet was re-suspended by shaking in 35 ml of RM and centrifuged at 1,000 x g for 10 min. Final myofibril pellets were re-suspended in 20 ml of RB plus 0.1 mM phenylmethylsulfonyl fluoride and were ready for sarcomere length determination. Sarcomere Determination Purified myofibrils were transferred onto a microscope slide using centrifugation (Cytofuge 2, Model M801-22, Westwood, MA). Samples were then incubated for 60 min at 37˚C with monoclonal anti-α-actinin (sarcomeric), (A7811 Sigma, St. Louis, MO) diluted 1:5000 in RB. Samples were washed three times in RB (2 min per wash) and then incubated with a donkey anti-mouse IgG, FITC (fluorescein isothiocyanate) conjugated secondary antibody diluted 1:500 in RB for 60 min at 37˚C. Samples were washed three times in RB (2 min per wash). Samples were mounted with 30 µl mounting media (75 mM KCl, 10 mM Tris, pH 8.5, 2 mM MgCl 2 , 2 mM EGTA, 1 mM NaN 3 , 1 mg/ml p- phenylenediamine, 75% v/v glycerol). Sarcomere length was measured directly using a microscope (Olympus AX70) equipped with a fluorescence filter (FITC) at 400X magnification. Myofibril images were captured using Olympus DP71 camera and sarcomere length was measured using Image Analysis Softeware (Image Pro-Plus 5.1). The average of five sarcomeres was determined across 20 myofibrils per sample.
  • 40. 30 Whole Muscle Protein Extraction Upon completion of appropriate aging lengths (0, 1, 4, 7, or 10 d postmortem), sample proteins were extracted as described by Huff-Lonergan, Mitsuhashi, Parrish, & Robson (1996) with slight modifications. Briefly, 0.2 g of muscle were minced and added to 25 volumes of homogenizing solution (10 mM sodium phosphate, 10% w/v volume SDS, pH 7.0). Samples were homogenized using a motor driven Dounce homogenizer and clarified by centrifugation at 1,500 X g for 15 min at 4°C. Protein concentration was determined using the RC/DC Protein assay (based on the Lowry assay) (Bio-Rad Laboratories, Hercules, CA) and samples were diluted with water to a final concentration of 2.5 mg/ml. One volume of each sample was added to five volumes of sample buffer (3mM EDTA, 3% w/v SDS, 30% v/v glycerol, 0.003% w/v pyronin Y, 30 mM Tris-HCl pH 8.0) and 0.1 vol of 2-mercaptoethanol for a final protein concentration of 1.56 mg/ml. Samples were immediately heated to 100°C for 5 min and stored at - 20°C. Gel Electrophoresis and Transfer conditions Muscle extracts were loaded on 15% polyacrylamide resolving gels (Ready Gel, Tris-HCl gels; Bio-Rad Laboratories, Hercules, CA). Gels were run on the Bio-Rad Criterion Cell system (Bio-Rad Laboratories, Hercules, CA) at constant 200 V for approximately 90 min at 4°C. The running buffer used in both the upper and lower chambers consisted of 25 mM Tris, 192 mM glycine, 2 mM EDTA, and 0.1% w/v SDS. Following electrophoresis, gels were equilibrated for 30 min at room temperature in
  • 41. 31 transfer buffer (25 mM Tris, 192 mM glycine, and 15% v/v methanol). Gels were transferred to polyvinylidene difluoride (PVDF) membranes at 4°C using a Criterion blotter (Bio-Rad Laboratories, Hercules, CA) at a constant 90 V for 45 min. Western blotting Following transfer, membranes were blocked in Odyssey Blocking Buffer (OBB) ( Licor, Lincoln, NE) for 60 min at room temperature. Blots were incubated for 60 min at room temperature with rabbit anti-Actin (20-33, Sigma, St. Louis, MO) and monoclonal anti-TnT (JLT-12, Sigma, St. Louis, MO), diluted 1:5000 in OBB and Phosphate buffered saline with Tween (PBST). Blots were washed four times with PBST (5 min per wash) and then incubated with conjugated secondary antibodies, goat anti-mouse and goat anti-rabbit (Licor, Lincoln, NE) diluted 1:25000 in PBST for 60 min at room temperature. Blots were washed four times in PBST and bands were visualized using a Licor Odyssey Software (Lincoln, NE). Immunoreactive TnT was identified and the disappearance of intact TnT was quantified using Licor Odyssey scanner (Lincoln, NE). To account for potential variation in protein loading, intensities of protein bands were expressed relative to the total intensity of actin bands within the lane. The relative intensity of bands was then normalized to the relative intensity of intact TnT at 0 h. The percent degradation of the first three bands of TnT was calculated as percent TnT degraded from day 0.
  • 42. 32 Total Collagen Content Total collagen content was calculated from hydroxyproline quantification similar to the methods by Bergman and Loxley (1961) and Hill (1966) with slight modifications. Approximately, 2 g of powdered meat was placed into a 50-mL centrifuge tube and 12 ml of ¼ strength Ringer’s solution was added. Tubes were heated for 63 min at 77°C in a shaking water bath. Tubes were then centrifuged at 3000 x g for 5 min. After centrifugation supernatant was transferred to a flat bottom flask for analysis of soluble collagen. The residue in the tube was washed with 8 mL of Ringer’s solution and centrifugation was performed again. After centrifugation supernatant was transferred to the same flat bottom flask. Soluble collagen was then hydrolyzed in 20 mL of 12N HCl. Pellet remaining in the tube was transferred using 40 mL of 6N HCL (four 10 mL rinses) to the flat bottom flask for analysis of insoluble collagen. Flat bottom flasks were placed under condensers and refluxed in a heated sand bath for 16 hours. After refluxing, pH of the samples were adjusted to 6.2 with 12 M NaOH. Sample and 1 g of charcoal were transferred to a 250 mL volumetric flask and brought up to volume with double-distilled H 2 0. After filtration, 1 mL of sample solution was pipetted into a 25 mL test tube. Two mL of isopropanol were added, vortexed and then immediately 1 mL of oxidant solution was added, vortexed and reacted for 4 min. Thirteen mL of Erhlich’s solution were added to the tubes, vortexed and heated for 25 min at 60°C in a water bath. Tubes were cooled for five min in cold water. Absorbance was read at 558 nm with a spectrophotometer (Shimudzu UV 2101 PC). Hydroxyproline was converted to collagen by multiplying by 7.25 (Goll et al., 1963). Values form soluble and insoluble collagen
  • 43. 33 were added together to obtain of total collagen. Data are expressed as percent of collagen per gram of raw muscle. Statistical Analysis Warner-Bratzler shear force, sarcomere length, troponin T degradation, total collagen, resistance, reactance, and phase angle were analyzed using the PROC GLM procedure of SAS, with animal as the experimental unit. Least squares means were calculated and separated using the PDIFF option. Fixed main effects in the model included breed (B), suspension (S), and day postmortem (D) and their interactions (B x S, S x D, B x D, B x S x D). Pearson’s correlation coefficients were generated using the PROC CORR procedure of SAS to examine relationship among dependent variables.
  • 44. 34 Results and Discussion Phase I Carcass Traits Means and standard deviations for hot carcass weight (HCW), adjusted fat thickness, ribeye area, calculated USDA yield grade, marbling score, and Warner- Bratzler shear (WBS) are presented in Table 2.1. Carcasses sampled in phase I had 11 kg higher HCW, 0.2 cm less adjusted fat thickness and 6.5 cm2 larger ribeye areas when compared to the average industry carcass (NBQA, 2005). Consequently, calculated USDA yield grade was 0.2 lower than industry average, but marbling score was similar. Overall, carcasses sampled were similar to the average industry carcass (NBQA, 2005). Warner-Bratzler shear force decreased 13.5% from d 5 to d 14 postmortem. Electrical Impedance Electrical impedance measurements on the exposed longissimus using 5 cm penetration depth on d 4 were more accurate at predicting WBS than measurements at other anatomical locations or using other electrode designs on d 4 (Table 2.2). In addition, impedance measures on steaks (d 5 & d 14) tended to be better predictors than measures on carcasses (d 4). Impedance measures from tagged (carcass side that sample was removed) and untagged sides (carcass side that sample was not removed) were similar in their ability to predict WBS force, and using the average of two measurements did not significantly improve accuracy compared to using only the tagged side (Table
  • 45. 35 2.3). Therefore, impedance could be measured on either side of the carcass without altering impedance’s ability to predict beef tenderness. Day 4 impedance measurements on the carcass were better at predicting d 14 WBS force than d 5 WBS force (Table 2.4). In addition, d 5 impedance was better at predicting d 5 WBS force versus d 14 WBS force, but d 14 impedance was similar at predicting both d 5 and d 14 WBS force. Marbling was the poorest predictor of WBS (Table 2.5), explaining only 4% of the variation in WBS force, similar to Wheeler, Cundiff, and Koch (1994) who reported marbling explained, at most, 5% of the variation in palatability traits, but different from results by Wulf and Page (2000) who concluded marbling score, by itself, explained 12% of the variation in beef palatability. In addition, marbling added little to nothing when combined with other predictors. Carcass traits, color, and impedance were similar in their prediction capability, and when added together were quite additive suggesting that each is explaining a different component of tenderness. Conclusion from phase I, was that electrical impedance was able to predict beef tenderness. Phase II Carcass Traits Means and standard deviations for hot carcass weight (HCW), adjusted fat thickness, ribeye area, calculated USDA yield grade, marbling score, and Warner- Bratzler shear (WBS) are presented in Table 2.6. Carcasses sampled in phase II had 13 kg higher HCW with similar adjusted fat thickness and 3.3 cm2 larger ribeye areas when compared to the average industry carcass (NBQA, 2005). Calculated USDA yield grade
  • 46. 36 was similar, but marbling score was 33 points lower than industry average. Overall, carcasses sampled were similar to the average industry carcass (NBQA, 2005). Electrical Impedance By themselves, NIR, Impedance, and BeefCam explained 7%, 5%, and 3% of the variation in 14 d WBS force, respectively (Table 2.7). When used together, NIR and Impedance explained 12% of the variation. Because they were additive, it appears that NIR and Impedance were measuring two different components of beef tenderness. In order for an instrument to “eliminate the tough cattle”, it must be able to certify a large percentage of the population while minimizing the percentage tough cattle within the certified group. The 70 to 90% certification rates would be of the most interest in this “eliminate-the-tough-cattle” scenario. The NIR system was the most effective instrument at eliminating the tough carcasses (Table 2.8). Neither marbling nor impedance were useful in trying to sort off the tough carcasses and certify a large percentage of the population. An instrument could be used to select a premium group of exceptional beef, if it were able to “identify the very tender cattle”. The 20 to 40% certification rates would be of the most interest in this “identify-the-very-tender-cattle” scenario. The Impedance system was the most effective instrument at identifying the very tender carcasses (Table 2.9). The NIR system was not useful in trying to sort off the very tender carcasses. Therefore, it appears that the reason that NIR and Impedance appeared additive in Table 2.7 was because NIR was the most effective at identifying the tough carcasses and Impedance was the most effective at identifying the very tender carcasses.
  • 47. 37 Another way to compare the technologies is to look at their effectiveness to sort into tenderness groups. Table 2.10 shows average d 14 WBS force for each fifth of the sample population sorted by predicted tenderness by each instrument. Each fifth contains 56 to 60 carcasses. The 1st fifth was predicted to be the most tender and the 5th fifth was predicted to be the least tender. Similar to the results in Tables 2.8 and 2.9, Impedance was the most effective technology at identifying the most tender carcasses, whereas NIR was the most effective technology at identifying the toughest carcasses. Impedance measurements on d 1 postmortem were not highly correlated with measurements on d 2 and d 3 postmortem (Table 2.11), however d 2 impedance measurements were highly correlated with d 3 impedance measurements. Therefore, impedance is a more accurate predictor of beef tenderness the more beef ages. Phase III Carcass Traits Means and standard deviations for hot carcass weight (HCW), adjusted fat thickness, ribeye area, calculated USDA yield grade, marbling score, Warner-Bratzler shear (WBS), and electrical impedance are presented in Table 2.12. Carcasses sampled in phase III had 25 kg lower HCW, 0.4 cm less adjusted fat thickness with similar ribeye areas when compared to the average industry carcass (NBQA, 2005). Consequently, calculated USDA yield grade was 0.4 lower along with marbling score 67 points lower than industry average. Overall, carcasses sampled were slightly smaller and leaner than
  • 48. 38 the average industry carcass (NBQA, 2005). Warner-Bratzler shear force decreased 21.1% from d 1 to d 10 postmortem. Warner-Bratzler Shear Force As expected, longissimus WBS values were lower for Bos taurus (43.6 N) than Bos indicus (57.0 N) (P < 0.01, Table 2.13) and similar to values reported by Crouse et al. (1989) for beef longissimus at d 7 postmortem. Additionally, HS resulted in lower WBS values (46.4 N) than NS (54.3 N) (P < 0.0001, Table 2.13). Previous research by Herring et al. (1965) and Hostetler et al. (1972), reported longissimus samples with longer sarcomeres via hip suspension resulted in lower shear force values. Mean WBS values decreased significantly (P < 0.0001) over the aging period from 56.9 N at d 1 to 43.7 N by d 10 postmortem (Table 2.14), indicating a significant improvement in tenderness. There was a significant breed x day interaction for WBS (P < 0.05, Table 2.14). At d 10 postmortem WBS values were lower in both Bos taurus and Bos indicus than at d 1 postmortem, but Bos taurus decreased WBS to a greater extent than Bos indicus (P < 0.05). Sarcomere Length Representative micrographs of myofibrils with normal (1.82 µm) and hip suspension (2.43 µm) sarcomeres are presented in Figure 2.4. Sarcomere length was longer for Bos taurus (2.19 µm) than for Bos indicus (2.06 µm) (P < 0.05, Table 2.14), which is different from previous research by Whipple et al., (1990) and Stolowski et al., (2006) who found no difference in sarcomere length between Bos taurus and Bos indicus.
  • 49. 39 Longer sarcomeres found in Bos taurus was a result of hip suspension as there was no difference in carcasses from normal suspension. As expected, HS resulted in longer sarcomeres than NS (P < 0.0001, Table 2.13) and is in agreement with previous research by Herring et al. (1965) and Hostetler et al. (1972). There was a significant breed x suspension interaction for sarcomere length (P < 0.001, Table 2.13). Hip suspension increased sarcomere length of both Bos taurus and Bos indicus, but HS improved sarcomere length of Bos taurus to a greater extent than Bos indicus (P < 0.001). This interaction could be a result of different skeletal structure found between the two species resulting in longer sarcomeres in the HS Bos taurus. Postmortem Proteolysis A representative Western blot of whole muscle protein extracts from bovine longissimus with normal and hip suspension is presented in Figure 2.5. Degradation of troponin T was utilized in this study to measure the rate of postmortem proteolysis. Troponin T is a regulatory protein not typically involved in the tenderization of meat, but has been considered a good marker for proteolysis (Negishi, Yamamoto, and Kuwata, 1996; Penny & Dransfiled, 1979). Bos taurus had more breakdown of troponin T than Bos indicus (P > 0.05, Table 2.13) and is agreement with past research by O’Connor et al., (1997) that reported Bos indicus cattle have an increased level of calpastatin and therefore aged at slower rate than Bos taurus. Hip suspension resulted in more breakdown of troponin T than NS, (P < 0.0001, Table 2.13) and confirmed past research by Weaver, Bowker, and Gerrard, (2008) that concluded troponin T proteolysis is
  • 50. 40 sarcomere length-dependent and an increase in sarcomere length resulted in more rapid protein degradation. As expected, there was increased degradation of intact troponin T during aging from d 1 to d 10 postmortem (P < 0.0001, Table 2.14). Collagen Content Collagen (soluble, insoluble, and total) was measured to determine the amount of connective tissue in each longissimus sample. The was no difference between Bos taurus and Bos indicus for collagen values (Table 2.13). Therefore, breed type had no effect on the amount of connective tissue found in the longissimus muscle in this study. Electrical Impedance Electrical impedance values, reactance and phase angle, were lower for Bos taurus than Bos indicus (P < 0.05, Table 2.13), indicating reactance and phase angle could be related to the level of calpastatin. Hip suspension resulted in increased resistance values than NS (P < 0.0001, Table 2.13), indicating that resistance was dependent upon sarcomere length. Hip suspension resulted in lower phase angle values than NS (P < 0.0001, Table 2.13), possibly because an increase in sarcomere length resulted in a weakened structure. The breed x suspension interaction was significant for resistance (P < 0.001, Table 2.13). Hip suspension resulted in higher resistance values than NS, but Bos indicus resulted in a greater increase in resistance values than Bos taurus, (P < 0.001). The breed x suspension interaction was significant for reactance (P < 0.0001, Table 2.13). Hip suspension had lower reactance values than NS for Bos taurus, but HS had higher reactance values than NS for Bos indicus, (P < 0.0001). The
  • 51. 41 breed x suspension interaction was significant for phase angle (P < 0.0001, Table 2.13). Hip suspension resulted in lower phase angle values than NS for Bos taurus, but no difference was found in phase angle for Bos indicus for suspension method, indicating sarcomere length did not influence phase angle values of Bos indicus cattle (P < 0.0001). Phase angle decreased from d 1 to d 10 postmortem (P < 0.01, Figure 2.2), indicating phase angle may be inversely related to postmortem proteolysis. Previous work by Lepetit et al., (2002) and Damez, Clerjon, Abouelkaram, and Lepetit, (2007) reported a decrease in electrical impedance during aging and suggested electrical impedance could potentially evaluate meat aging. In addition, Byrne et al., (2000) reported electrical measurements changed significantly between d 1 and d 14 postmortem and were significantly correlated to WBS. Correlations Correlations are presented in Table 2.15. Sarcomere length was negatively related to WBS (r = -0.38), indicating longer sarcomeres resulted in a lower WBS values. In addition, sarcomere length was related to total collagen (r = 0.57), indicating longer sarcomeres had more connective tissue. Postmortem proteolysis was inversely related to WBS (r = -0.59), indicating proteolysis of myofibril proteins is the reason meat tenderizes during postmortem aging. Reactance and sarcomere length were negatively correlated (r = -0.24), indicating increased sarcomere length resulted in greater breakdown of membranes. Phase angle was negatively correlated to sarcomere length (r = -0.38), and postmortem proteolysis (r = -0.25), confirming phase angle is inversely
  • 52. 42 related to postmortem proteolysis. In addition, phase angle was related to WBS (r = 0.51), indicating phase angle can predict beef tenderness.
  • 53. 43 Implications In conclusion, electrical impedance is weakly related to beef tenderness, but may be useful to sort out very tender beef carcasses. Phase angle was inversely related to extent of postmortem proteolysis. Therefore, electrical impedance may be useful to predict beef aging.
  • 54. 44 Table 2.1. Simple statistics for carcass and muscle traits for Phase I Trait n Mean SD Minimum Maximum Hot carcass weight, kg 92 371 39 203 462 Adjusted fat thickness, cm 92 1.10 0.48 0.30 2.54 Ribeye area, cm2 92 92.9 12.9 46.5 124.5 Calculated USDA yield grade 92 2.7 0.9 0.8 4.4 Marbling scorea 92 431 115 250 960 Warner-Bratzler shear, N Longissimus, d 5 postmortem 92 43.1 10.8 19.6 83.4 Longissimus, d 14 postmortem 92 37.3 8.2 14.7 66.7 a 300 = quot;Slight00,quot; 400 = quot;Small00,quot; 500 = quot;Modest00,quot; 600 = quot;Moderate00.quot;
  • 55. 45 Table 2.2. The ability of electrical impedance measurements from various electrode depth and placement combinations to predict shear force (average of d4 and d14) Measurement R-square R-square (impedance only) (with other carcass traits) None 0.00 0.34 Carcass Measurements (day 4) a Ribeye 5 cm puncture 0.14 0.42 a Ribeye 1.3 cm puncture 0.06 0.40 Ribeye surfacea 0.08 0.38 b Strip loin 5 cm puncture 0.05 0.37 c Strip to round 5 cm puncture 0.07 0.38 Steak Measurements (day 5) a Ribeye 1.3 cm puncture 0.19 0.45 a Ribeye surface 0.20 0.45 Steak Measurements (day 14) a Ribeye 1.3 cm puncture 0.28 0.49 a Ribeye surface 0.27 0.48 a Electrodes in a linear arrangement with 5 cm between each electrode b Electrodes in a linear arrangement with 5 cm between detection and electrical source over a 50 cm field c Electrodes placed in the strip loin and round with 5 cm between detection and electrical source
  • 56. 46 Table 2.3. Evaluating the ability of electrical impedance measurements from tagged (carcass side sample was removed) and untagged (carcass side sample was not removed) sides to predict shear force (average of d5 and d14) Measurement R-square R-square (impedance only) (with other carcass traits) None 0.00 0.34 Ribeye 5 cm puncturea Tagged side 0.14 0.42 Untagged side 0.12 0.41 Average of both 0.15 0.42 Ribeye 1.3 cm puncturea Tagged side 0.06 0.40 Untagged side 0.05 0.37 Average of both 0.05 0.39 Ribeye surfacea Tagged side 0.08 0.38 Untagged side 0.08 0.36 Average of both 0.11 0.37 a Electrodes in a linear arrangement with 5 cm between each electrode
  • 57. 47 Table 2.4. Predicting d5 and d14 shear force using electrical impedance measurements from d4, d5, and d14 d 5 WBS force d 14 WBS force Measurement R-square R-square R-square R-square (impedance only) (with other traits) (impedance only) (with other traits) None 0.00 0.34 0.00 0.30 Day 4 a Ribeye 5 cm puncture 0.10 0.39 0.15 0.38 a Ribeye 1.3 cm puncture 0.03 0.39 0.08 0.35 a Ribeye surface 0.07 0.38 0.10 0.34 Day 5 a Ribeye 1.3 cm puncture 0.23 0.45 0.11 0.37 a Ribeye surface 0.22 0.44 0.13 0.38 Day 14 a Ribeye 1.3 cm puncture 0.26 0.44 0.25 0.47 Ribeye surfacea 0.22 0.42 0.26 0.47 a Electrodes in a linear arrangement with 5 cm between each electrode
  • 58. 48 Table 2.5. R-squares for various prediction models to predict shear force (d14) Variables used in the prediction model R-square Marbling 0.04 Carcass traits (other than marbling) 0.19 Color (from camera) 0.14 Impedance (ribeye 5 cm puncture – day 4) 0.15 Marbling + Carcass traits 0.22 Marbling + Color 0.15 Marbling + Impedance 0.15 Carcass traits + Color 0.28 Carcass traits + Impedance 0.26 Color + Impedance 0.29 Marbling + Carcass traits + Color 0.30 Marbling + Carcass traits + Impedance 0.27 Marbling + Color + Impedance 0.29 Carcass traits + Color + Impedance 0.37 Marbling + Carcass traits + Color + Impedance 0.38
  • 59. 49 Table 2.6. Simple statistics for carcass and muscle traits for Phase II Trait n Mean SD Minimum Maximum Hot carcass weight, kg 300 373 33 269 476 Adjusted fat thickness, cm 300 1.27 0.42 0.41 3.56 Ribeye area, cm2 300 89.7 9.7 69.0 131.0 Calculated USDA yield grade 300 3.0 0.7 1.0 5.9 Marbling scorea 300 399 61 290 640 Warner-Bratzler shear, N Longissimus, d 14 postmortem 300 33.3 7.8 20.6 71.6 a 300 = quot;Slight00,quot; 400 = quot;Small00,quot; 500 = quot;Modest00,quot; 600 = quot;Moderate00.quot;
  • 60. 50 Table 2.7. R-squares for various prediction models to predict shear force (d14) Prediction Model R-square SDSU Carcass Data 0.07 Camera Carcass Data 0.06 Camera Color Data 0.05 BeefCam 0.03 NIR 0.07 Impedance 0.05 14-d Impedance 0.16 CamCarc + CamColor 0.11 CamCarc + BeefCam 0.07 CamCarc + NIR 0.13 CamCarc + Impedance 0.12 CamColor + BeefCam 0.05 CamColor + NIR 0.09 CamColor + Impedance 0.11 BeefCam + NIR 0.10 BeefCam + Impedance 0.07 NIR + Impedance 0.12 CamCarc + CamColor + BeefCam 0.11 CamCarc + CamColor + NIR 0.15 CamCarc + CamColor + Impedance 0.17 CamCarc + BeefCam + NIR 0.14 CamCarc + BeefCam + Impedance 0.12 CamCarc + NIR + Impedence 0.18 CamColor + BeefCam + NIR 0.11 CamColor + BeefCam + Impedance 0.12 CamColor + NIR + Impedance 0.16 BeefCam + NIR + Impedance 0.15 CamCarc + CamColor + BeefCam + NIR 0.16 CamCarc + CamColor + BeefCam + Impedance 0.17 CamCarc + CamColor + NIR + Impedance 0.21 CamCarc + BeefCam + NIR + Impedance 0.19 CamColor + BeefCam + NIR + Impedance 0.17 CamCarc + CamColor + BeefCam + NIR + Impedance 0.21
  • 61. Table 2.8. Percentage of tough (>49.0 N) carcasses by certification rate and instrument Certification Rate, % Marbling NIR BeefCam Impedance No System Perfect System Impedance d14 10 3.3 0.0 0.0 0.0 4.3 0.0 0.0 20 3.3 0.0 0.0 0.0 4.3 0.0 0.0 30 2.2 0.0 2.4 1.1 4.3 0.0 0.0 40 1.7 0.0 1.8 1.7 4.3 0.0 1.7 50 4.0 0.7 1.4 2.7 4.3 0.0 1.3 60 4.4 1.7 1.8 4.4 4.3 0.0 1.7 70 3.8 2.4 2.5 4.8 4.3 0.0 1.4 80 4.6 2.1 4.0 4.2 4.3 0.0 2.9 90 4.8 3.4 3.6 4.4 4.3 0.0 3.0 100 4.3 4.4 4.6 4.3 4.3 4.3 4.3 n 300 295 281 300 300 300 300 51
  • 62. Table 2.9. Percentage of very tender (<34.3 N) carcasses by certification rate and instrument Certification Rate, % Marbling NIR BeefCam Impedance No System Perfect System Impedance d14 10 46.7 36.7 42.9 46.7 30.3 100.0 43.3 20 43.3 33.9 46.4 48.3 30.3 100.0 43.3 30 40.0 33.7 40.5 46.7 30.3 100.0 37.8 40 37.5 35.6 42.0 45.0 30.3 75.8 35.0 50 36.7 33.1 38.0 41.3 30.3 60.7 35.3 60 33.3 33.3 38.5 38.3 30.3 50.6 35.0 70 33.3 32.9 36.5 35.7 30.3 43.3 33.3 80 30.8 32.2 34.2 33.3 30.3 37.9 32.5 90 30.7 31.6 32.8 32.6 30.3 33.7 33.0 100 30.3 30.5 31.7 30.3 30.3 30.3 30.3 n 300 295 281 300 300 300 300 52
  • 63. 53 Table 2.10. Average Warner-Bratzler shear force (N) for sort groups (fifths) by instrument Sort Group Marbling NIR BeefCam Impedance No System Perfect System Impedance d14 1st Fifth 31.5 31.7 31.5 30.2 33.4 24.8 30.8 2nd Fifth 32.9 32.0 32.5 32.2 33.4 29.1 33.2 3rd Fifth 35.0 33.7 32.8 35.2 33.4 32.1 32.1 4th Fifth 35.1 33.0 35.7 34.7 33.4 36.1 34.1 5th Fifth 32.9 37.1 35.2 35.2 33.4 45.2 37.2 n 300 295 281 300 300 300 300
  • 64. 54 Table 2.11. Effect of day postmortem on impedance measurements (n = 100) R Xc PA Means Day 1 47.2 31.1 33.3 Day 2 46.1 30.6 33.5 Day 3 42.6 27.1 32.3 Correlations Day 1&2 0.67 0.56 0.74 Day 1&3 0.76 0.59 0.68 Day 2&3 0.88 0.86 0.87 Correlation with WBS Day 1 -0.20 -0.18 -0.03 Day 2 -0.05 0.03 0.09 Day 3 -0.08 0.06 0.15
  • 65. 55 Table 2.12. Simple statistics for carcass and muscle traits for Phase III Trait n Mean SD Minimum Maximum Hot carcass weight, kg 14 335 21 303 384 Adjusted fat thickness, cm 14 0.91 0.50 0.20 1.83 2 Ribeye area, cm 14 83.0 8.5 67.7 99.4 Calculated USDA yield grade 14 2.5 0.7 1.5 4.2 a Marbling score 14 365 89 280 620 Warner-Bratzler shear, N Longissimus, d 1 postmortem 28 56.9 9.8 40.2 78.5 Longissimus, d 10 postmortem 28 44.1 11.8 26.5 9.6 Sarcomere length, µm 112 2.12 0.36 1.64 3.01 Troponin T degradation, % 112 44.0 22.0 86.0 11.0 Total collagen, % 14 0.39 0.04 0.31 0.46 Electrical impedance Resistance 196 62.64 6.98 49.00 80.00 Reactance 196 37.83 7.65 21.00 63.00 Phase Angle 196 30.94 4.44 18.69 38.93 a 300 = quot;Slight00,quot; 400 = quot;Small00,quot; 500 = quot;Modest00,quot; 600 = quot;Moderate00.quot;
  • 66. Table 2.13. Least square means of WBS force, sarcomere length, troponin T degradation, collagen, resistance, reactance, and phase angle (averaged across all aging periods) Breed Main Effects Suspension Main Effects Breed x Suspension Interaction TA IN Pooled TA IN SEM P<F NS HS SEM P<F NS HS NS HS SEM P<F WBS, N 43.6 57.0 2.20 0.0010 54.3 46.4 1.10 <0.0001 47.5 39.7 61.1 53.0 1.70 0.9131 a c a b Sarcomere length, µm 2.19 2.06 0.03 0.0155 1.82 2.43 0.02 <0.0001 1.83 2.55 1.81 2.30 0.03 0.0009 Troponin T degradation, % 46.0 42.0 1.00 0.0350 42.0 47.0 1.00 <0.0001 43.0 48.0 41.0 47.0 1.00 0.6090 Soluble Collagen, % 0.04 0.04 0.01 0.5091 - - - - - - - - - - Insoluble Collagen, % 0.35 0.34 0.01 0.4368 - - - - - - - - - - Total Collagen, % 0.39 0.38 0.02 0.8077 - - - - - - - - - - Resistance 61.16 64.12 2.15 0.3498 60.59 64.69 0.38 <0.0001 60.12a 62.20b 61.06ab 67.18c 0.54 0.0003 b a c d Reactance 33.82 41.84 1.29 0.0009 38.03 37.62 0.58 0.6190 35.90 31.73 40.16 43.51 0.43 <0.0001 Phase Angle 28.81 33.08 0.87 0.0047 31.96 29.92 0.30 <0.0001 30.69b 26.92a 33.23c 32.93c 0.43 <0.0001 a,b,c Means for the breed x suspension interaction within a row lacking a common superscript letter differ (P< 0.05). 56
  • 67. Table 2.14. Effect of day shear force, sarcomere length, and troponin T degradation Day Main Effects Breed x Day Interactions TA IN Pooled 1 4 7 10 SEM P<F 1 4 7 10 1 4 7 10 SEM P<F WBS, N 56.9 - - 43.7 1.10 <0.0001 52.1f - - 35.2e 61.7g - - 52.4f 1.60 0.0242 Sarcomere length, µm 2.13 2.14 2.11 2.12 0.03 0.8985 2.20 2.21 2.18 2.18 2.07 2.07 2.04 2.06 0.05 0.9941 d c b a Troponin T degradation, % 18.0 34.0 51.0 75.0 1.00 <0.0001 19.0 36.0 54.0 76.0 16.0 31.0 49.0 73.0 1.00 0.6912 a,b,c,d Means within a row lacking a common superscript letter differ (P< 0.05). e,f,g Means for the breed x day interaction within a row lacking a common superscript letter differ (P< 0.05). 57
  • 68. 58 Table 2.15. Correlations of Warner-Bratzler shear force, sarcomere length, proteolysis, collagen, resistance, reactance, and phase angle (* P < 0.05) Variablea SL PROT SC IC TC R Xc PA WBS -0.38* -0.59* -0.25 -0.25 -0.24 -0.12 0.34 0.51* SL - 0.10 0.31 0.64* 0.57* 0.13 -0.24* 0.38* PROT - - -0.17 0.07 -0.30 0.14 -0.14 0.25* SC - - - 0.58* 0.83* 0.24 0.08 -0.80 IC - - - - 0.93* 0.40 -0.14 -0.44 TC - - - - - 0.37 -0.06 -0.33 R - - - - - - 0.49* -0.08 Xc - - - - - - - 0.83* a WBS = Warner-Bratzler shear, SL= Sarcomere Length, PROT = Troponin T degradation, %, SC= Soluble Collagen, %, IC=Insoluble Collagen, %, TC= Total Collagen, %, R = Resistance, Xc = Reactance, PA = Phase Angle
  • 69. 59 Figure 2.1. Electrode arrangement and penetrating depths. a) 2.5 cm 5.0 cm 2.5 cm 5.0 cm b) 2.5 cm 5.0 cm 2.5 cm 1.3 cm c) 2.5 cm 5.0 cm 2.5 cm 0 cm
  • 70. 60 Figure 2.2. The effect of breed type on resistance, reactance, and phase angle on the exposed longissimus muscle from d 1 to d 7 postmortem (*P < 0.05)
  • 71. 61 Figure 2.3. The effect of suspension on resistance, reactance, and phase angle on the exposed longissimus muscle from d 1 to d 7 postmortem (*P < 0.05)
  • 72. 62 Figure 2.4. Representative micrographs of myofibrils with normal (1.82 µm, left) and hip suspension (2.43 µm, right) sarcomeres of longissimus muscle (Magnification 400X)
  • 73. 63 Figure 2.5. Representative western blot of whole muscle protein extracts from bovine longissimus muscle with normal (NS) and hip (HS) suspension and postmortem aging periods were 1, 4, 7, and 10 days (d0 was used for quantification) NS HS . 0 1 4 7 10 1 4 7 10 Band } 1-3 4&5 6 7
  • 74. 64 Literature Cited AMSA. Guildlines for Cookery and Sensory Evaluation of Meat. (1995). American Meat Science Association, Chicago, IL. Bergman, I. and R. Loxley. (1963). Two improved and simplified methods for the spectrophotmetric determination of hydroxyproline. Analytical Chemistry, 35, 1961-1965. Boleman, S. J., S. L. Boleman, R. K. Miller, J. F. Taylor, H. R. Cross, T. L. Wheeler, M. Koohmaraie, S. D. Shackelford, M. F. Miller, R. L. West, D. D. Johnson, and J. W. Savell. (1997). Consumer evaluation of beef of known categories of tenderness. Journal of Animal Science, 75, 1521–1524. Byrne, C. E., D. J. Troy, and D. J. Buckley. (2000). Postmortem changes in muscle electrical properties of bovine M. longissimus dorsi and their relationship to meat quality attributes and pH fall. Meat Science, 54, 23-34. Crouse, J. D., L. V. Cundiff, R. M. Koch, M. Koohmaraie and S. C. Seideman. (1989). Comparisons of Bos indicus and Bos taurus inheritance for carcass beef characteristics and meat palatability. Journal of Animal Science, 67, 2661-2668. Damez, J., S. Clerjon, S. Abouelkaram, J. Lepetit. (2007). Dilelectric behavior of beef meat in the 1-1500 kHz range: Simulation with the Fricke/Cole-Cole model. Meat Science, 77, 512-519. Goll, D. E., W. G. Hoekstra, and R. W. Bray. (1963). Age-associated changes in muscle composition. The isolation and properties of a collagenous residue from bovine muscle. Journal of Food Science, 28, 503–509 Gupta, D., C. A. Lammersfeld, J. L. Burrows, S. L. Dahlk, P. G. Vashi, J. F. Grutsch, S. Hoffman, and C. G. Lis. (2004). Bioelectrical impedance phase angle in clinical practice: implications for prognosis in advanced colorectal cancer. The American Journal of Clinical Nutrition, 80, 1634-1638. Herring, H. K., R. G. Cassens, and E. J. Briskey. (1965). Further studies on bovine muscle tenderness as influenced by carcass position, sarcomere length, and fiber diameter. Journal of Food Science, 30, 1049-1054. Hill, F. (1966). The solubility of intramuscular collagen in meat animals of various ages. Journal of Food Science, 31, 161-166.
  • 75. 65 Hostetler, R. L., B. A. Link, W. A. Landmanm, and H. A. Fitzhugh, Jr. (1972). Effect of carcass suspension on sarcomere length and shear force of some major bovine muscles. Journal of Food Science, 37, 132-135. Huff-Lonergan, E., T. Mitsuhashi, F. C. Parrish Jr., and R. M. Robson. (1996). Sodium dodecyl sulfate-polyacrylamide gel electrophroresis and western blotting comparisons of purified myofibrils and whole muscle preparations for evaluating titin and nebulin in postmortem bovine muscle. Journal of Animal Science, 74, 779-785. Lukaski, H. C. (1996). Biological indexes considered in the derivation of the bioelectrical impedance analysis. American Journal of Clinical Nutrition, 64, 397S-404S. Meyers, R. H. (1990). Classical and modern regression with applications. 2nd ed. Duxbury, Pacifica Grove, CA. pp. 8, 82. Miller, M. F., M. A. Carr, C. B. Ramsey, K. L. Crockett, and L. C. Hoover. (2001). Consumer thresholds for establishing the value of beef tenderness. Journal of Animal Science, 79, 3062–3068. Morgan, J. B., J. W. Savell, D. S. Hale, R. K. Miller, D. B. Griffin, H. R. Cross, and S. D. Shackelford. (1991). National Beef Tenderness Survey. Journal of Animal Science, 69, 3274-3283. Negishi, H., E. Yamamoto, and T. Kuwata. (1996). The orign of the 30 kDa component appearing during post-mortem aging of bovine muscle. Meat Science, 42, 289- 303. NBQA. (2005). National Beef Quality Audit-2005: Survey of targeted cattle and carcass characteristics related to quality, quantity, and value of fed steers and heifers. Journal of Animal Science, 86, 3533-3543. NCA. (1994). National Beef Tenderness Conference: Executive Summary. National Cattlemen’s Association, Englewood, CO. O’Connor, S. F., J. D. Tatum, D. M. Wulf, R. D. Green, and G. C. Smith. (1997). Genetic effects on beef tenderness in Bos indicus composite and Bos taurus cattle. Journal of Animal Science, 75, 1822-1830. Penny, I. F., and E. Dransfield. (1979). Relationship between toughness and troponin t in conditioned beef. Meat Science, 3, 135-141.
  • 76. 66 Shackelford, S. D., T. L. Wheeler, M. K. Meade, J. O. Reagan, B. L. Byrnes, and M. Koohmaraie. (2001). Consumer impressions of Tender Select beef. Journal of Animal Science, 79, 2605-2614. Stolowski, G. D., B. E. Baird, R. K. Miller, J. W. Savell, A. R. Smith, J. F. Taylor, J. O. Sanders, and S. B. Smith. (2006). Factors influencing the variation in tenderness of seven major beef muscles from three Angus and Brahman breed crosses. Meat Science, 73, 475-483. Swartz, D. R., M. L. Greaser, and B. B. Marsh. (1993). Structural studies of rigor bovine myofibrils using fluorescence microscopy. I. Procedures for purification and modification of bovine muscle proteins for use in fluorescence microscopy. Meat Science, 33, 139-155. Weaver, A. D., B. C. Bowker, and D. E. Gerrard. (2008). Sarcomere length influences postmortem proteolysis of excised bovine semitendinosus muscle. Journal of Animal Science, 86, 1925-1932. Wheeler, T. L., L. V. Cundiff, and R. M. Koch. (1994). Effect of marbling degree on beef palatability in Bos taurus and Bos indicus cattle. Journal of Animal Science, 72, 3145-3151. Whipple, G., M. Koohmaraie, M. E. Dikeman, J. D. Crouse, M. C. Hunt and R. D. Klemm. (1990). Evaluation of attributes that affect longissimus muscle tenderness in Bos taurus and Bos indicus cattle. Journal of Animal Science, 68, 2716-2728. Wulf, D. M. and J. K. Page. (2000). Using measurements of muscle color, pH, and electrical impedance to augment the current USDA beef quality grading standards and improve the accuracy and precision of sorting carcasses into palatability groups. Journal of Animal Science, 78, 2595–2607.
  • 77. 67 Appendix A. The effect of breed type on resistance, reactance, and phase angle impedance measurements of longissimus lumborum steaks from d 1 to d 21 postmortem
  • 78. 68 Appendix B. The effect of suspension on resistance, reactance, and phase angle impedance measurements of longissimus lumborum steaks from d 1 to d 21 postmortem
  • 79. 69 Appendix C. The effect of breed type on resistance, reactance, and phase angle impedance measurements of semitendinosus steaks from d 1 to d 10 postmortem
  • 80. 70 Appendix D. The effect of suspension on resistance, reactance, and phase angle impedance measurements of semitendinosus steaks from d 1 to d 10 postmortem
  • 81. 71 Appendix E. The effect of breed type on resistance, reactance, and phase angle impedance measurements of psoas major steaks from d 1 to d 10 postmortem
  • 82. 72 Appendix F. The effect of suspension on resistance, reactance, and phase angle impedance measurements of psoas major steaks from d 1 to d 10 postmortem

×