PALATABILITY, CONSUMER ACCEPTANCE, AND ULTRASTRUCTURAL CHANGES OF HOT-PROCESSED PORK INFUSED WITH PHOSPHATE, SALT, GLUCOSE, AND POTASSIUM CHLORIDE

by CHIH KAN6 WU, B.S., M.B.A., M.S. A DISSERTATION IN ANIMAL SCIENCE Submitted to the Graduate Faculty of Texas Tech University in Partial Fulfillment of the Requirements for the Degree of DOCTOR OF PHILOSOPHY Approved

May, 1992 $0 ' /1.-(3 -^J 7

I ^^K. ACKNOWLEDGMENTS

Chang, Hsein-Chi, a famous 16th century Chinese philosopher once said, "Luckiness is you have time to study." I feel more than lucky for myself for not only having time to study but also for having a wonderful advisor, committee chairman and friend. Dr. C. Boyd Ramsey. If not for his patience, encouragement and advice, this dissertation will never reach the graduate school. I am also in debt to Dr. Markus F. Miller, Dr. Leslie D. Thompson and Dr. David Wester for their kindly assistance and advice in completing this work. I would especially like to express my appreciation to all the Meat Laboratory personnel and my fellow graduate students for the greatest friendship and help I have experienced in the past 6 years. Finally and most importantly, I am deeply thankful for the love and support of my parents, Ju-Chain and Pi-Lan Wu, and my beloved wife during the years I have studied in the United States.

11 TABLE OF CONTENTS

ACKNOWLEDGMENTS i i

ABSTRACT v

LIST OF TABLES vii

LIST OF FIGURES viii

CHAPTER I. INTRODUCTION 1

II. REVIEW OF LITERATURE 5 Pork as a Dietary Component 5 Hot-processing Pork 6 Preslaughter Factor Affecting Yield and Palatability 7 Environment 9 Slaughter and Early Postmortem Procedures 2 2 Fabrication and Processing 2 4 Later Processing Procedures 29 Curing Procedures and Equipment 3 4 Other Processes 3 6 Preservation 40 Distribution and Display 4 5 Pork Cookery 4 7 Sodium Chloride and Polyphosphates on Water Holding Capacity 52 Calcium-Activated Proteases 53

111 III. EFFECTS OF INFUSING CALCIUM CARBONATE, POLYPHOSPHATES, SALT, GLUCOSE, AND POTASSIUM CHLORIDE ON HOT-BONED PORK SEMIMEMBRANOSUS MUSCLE 55 Abstract 55 Introduction 56 Materials and Methods 58 Results and Discussion 60 Conclusions 65 Implications 66

IV. ULTRASTRUCTURAL CHANGES, PALATABILITY AND CONSUMER ACCEPTANCE OF COLD- OR HOT-BONED PORK INFUSED WITH GLUCOSE, NACL, PYROPHOSPHATE AND KCL 67 Abstract 67 Introduction 68 Materials and Methods 70 Results and Discussion 7 5 Conclusions 104 Implications 106

LITERATURE CITED 107

APPENDICES A. MUSCLE SAMPLE PREPARATION FOR TRANSMISSION ELECTRON MICROSCOPY 134

B. PREPARATION OF MUSCLE SAMPLE FOR SDS-PAGE 137 C. DATA SHEET FOR COOKING CHARACTERISTICS 139

D. TRAINED SENSORY PANEL EVALUATION FORM 141

E. MEMORANDUM FOR RECRUITING CONSUMER PANEL 14 3

F. RETURN FORM FOR CONSUMER PANEL 14 6 iv G. INSTRUCTION FORM FOR SAMPLE PREPARATION AT CONSUMER'S HOME 14 8 H. PORK CHOP EVALUATION FORM FOR CONSUMER PANEL 150 ABSTRACT

Two studies were conducted to determine the effect and the feasibility of infusing an aqueous solution containing phosphates, glucose, sodium and potassium chlorides, and calcium carbonate into hot-boned pork muscle to produce a precooked, low sodium, convenience-type pork entree. The first study showed the infusion of 4% glucose, 4% sodium, 3% sodium chloride and 2% potassium chloride (GNPK) tenderized the hot-boned (HB) semimembranosus muscle cooked 2 h postmortem. Adding calcium carbonate to the GNPK solution increased the calcium content of treated muscle but had no tenderization effect on the hot-boned muscle. The results from the second study showed pork loins treated with the GNPK solution were lower in Hunter color measurements than the chops infused with distilled water (DW) or the control (NO). HB-GNPK was lowest in Hunter L, a, b values and cooking losses among the groups. SDS-PAGE electrophoretograms showed that the 30,000 dalton component, a consistent indicator of tenderness of muscle, appeared in HB-GNPK and all the CB treatments but did not appear in HB-DW or HB-NO muscles. Electron microscopy showed that the Z-lines of sarcomeres of muscles treated with GNPK were degraded, which should have made them more tender. Sensory panel rating and Warner-Bratzler shear

vi force showed that the HB- and cold-boned (CB)-GNPK infused roasts were about 3 0% more tender than the CB-DW and CB-NO groups; they were about 43% more tender than the HB-DW and HB-NO roasts. GNPK infused groups also were rated higher in juiciness, flavor intensity and salt intensity than DW or NO groups. The HP-GNPK group contained more fat and less protein than the CB groups. Both CB- and HB-GNPK chops were rated more tender, juicier and more desirable than the other chops. CB-DW and CB-NO groups were rated not different from each other but more tender, juicier and more desirable than the HB-NO and HB-DW chops. Most (> 92.5%) consumers who evaluated the CB- or HB-GNPK chops would purchase the product if available. This study showed that the increased tenderness of HB-GNPK muscle may be due to the accelerated degradation of myofibrillar proteins and that it is feasible to infuse HB pork muscle with glucose, NaCl, pyrophosphates and KCl to produce a hot-boned, precooked, low salt, microwave ready, convenience-type pork product that is superior to the cold-boned products and accepted by consumers.

vii LIST OF TABLES

1. Treatment Descriptions 59 2. Means And Standard Errors For Cold-Processed Controls And Infusion Treatments 61 3. Carcass Traits Of The Pigs 71 4 . Demographics Of The Consumer Panel 75 5. Pearson Correlation Coefficients Between Sensory Panel Ratings 98 6. Means Of Consumer Responses To Purchase And Pricing Questions About Tested Pork Loin Chops....104

Vlll LIST OF FIGURES

1. Electron micrographs of GNPK treated (a) and control muscle samples 7 6

2. SDS-PAGE electrophoretogram of the effects of processing and the infusion of GNPK, DW and NO of the hot- and cold-boned pork muscles on myofibrillar proteins 78 3. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on Hunter L, a and b values of the cold- and hot-boned pork muscle 7 9 4. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on thawing loss of the cold- and hot-boned pork muscles 81 5. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on cooking losses of the cold- and hot-boned pork muscles 82 6. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on cooking time of the cold- and hot-boned pork muscles 8 6

7. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on juiciness of the cold- and hot-boned pork muscles 87

8. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on tenderness rating and Warner-Bratzler shear value of the cold- and hot-boned pork muscles 88

9. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on sensory panel ratings of salty flavor intensity of the cold- and hot-boned pork muscles 91

IX 10. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on sensory panel ratings of sweet flavor intensity of the cold- and hot-boned pork muscles 92 11. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on flavor intensity of the cold- and hot-boned pork muscles 94 12. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on overall palatability of the cold- and hot-boned pork muscles 96 13. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on moisture, fat, protein and ash content of the cooked cold- and hot-boned pork muscles 98 14. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on consumer ratings of tenderness of the cold- and hot-boned pork muscles 100 15. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on consumer ratings of juiciness of the cold- and hot-boned pork muscles 101 16. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on consumer ratings of overall desirability of the cold- and hot-boned pork muscles 102 17. Means for the effects of processing and the infusion of GNPK, DW and NO on consumer decisions of price they would pay for the cold- and hot-boned pork muscle product 105 CHAPTER I INTRODUCTION

The advantages of developing a hot-processing system for pork include superior functional properties of pre- rigor muscle for further product manufacturing, reduced cooler space requirements, decreased energy requirement for chilling, reduced labor requirements, shortened in-plant holding time, improved shelf life of the product, more uniform quality of the retail cuts, and reduced cooler shrinkage and level of purge (Reagan, 1983). All of these advantages increase the efficiency and are beneficial to the pork industry. However, these advantages are accompanied by several obstacles such as developing an efficient chilling system for hot-processed cuts, how to maintain product tenderness, controlling cut distortion and developing packaging systems that can adequately protect the hot-processed products (Reagan, 1983). Phosphates are the compounds prepared from phosphoric acid in which the acid has been partially or fully neutralized with alkali metal ions, predominately sodium, potassium or calcium. Depending on their basic structures, phosphates can be categorized into orthophosphates and condensed phosphates. Both groups of phosphates are extensively used in food industries (Dziezak, 1990). When they are used in red meat products, phosphates influence water binding, color, texture, coagulation, emulsification and microbial growth as a result of their chemical components (Barbut, 1989). Glucose is the most important biological sugar molecule; it serves as the major energy source for all the living cells (Stryer, 1988). In muscle cells, a glucose molecule enters the glycolysis pathway to be broken down to two molecules of acetyl-coenzyme A (Acetyl-CoA) and releases two molecules of adenosine triphosphate (ATP) high energy compound (Stryer, 1988). When the oxygen supply is abundant in muscle cells, the acetyl-CoA will enter the mitochondria and further break down to CO2 and water, releasing 36 ATP. On the other hand, if the oxygen supply is diminished, the acetyl-CoA will be converted to lactic acid and deposited in the muscle. The two ATP released through glycolysis are the only energy source for muscle to contract. After all the ATP is depleted, the muscle will stay in the contracted stage because no ATP is provided to release the actomyosin complex (Greaser, 1986). This stage is known as rigor mortis. Sodium chloride has been used as a safe preservative for food. Both sodium and chloride ions are major electrolytes in human body water balance (Vender et al., 1986). Overconsumption of sodium increases water intake and water retention to maintain homeostasis of the body. Therefore, high sodium intake has been identified as one possible contributor to the development of hypertension, which is a condition that occurs in 10 to 20% of the U.S. population (Pearson and Wolzak, 1982). Average Americans consume 10 to 12 g of salt per day, which is equivalent to 3,900 to 4,000 mg sodium (FASEB, 1979). This intake is 20 to 25 times greater than the minimum adult requirement (IFT, 1980). Medical and public opinion is inclining to the view that a general reduction of salt intake is desirable as a health maintenance measure and for treatment of hypertension and other cardiovascular diseases (Sebranek, et al., 1983). Recommendations to reduce dietary sodium levels were made by the USDA (1990) as a part of their dietary guidelines for Americans. Hoes et al. (1985) injected a solution containing 8% sodium chloride (NaCl), 2% potassium chloride (KCl) and 3% polyphosphate (PP) to 110% of fresh weight into pork m. longissimus muscles and produced more tender and juicy pork loin chops than the hot- or cold-processed controls. Wu (1990) reported the infusion of a glucose, NaCl, polyphosphate and KCl based solution to hot-processed pork and concluded that 4% NaCl can be replaced by glucose. The substitution of glucose for salt increased tenderness and juiciness of precooked products. However, the interactions between glucose, polyphosphates and NaCl were not clear. The objectives of the following studies were to determine: (1) the ultrastructural changes of the prerigor muscle fibers in response to the treatment with NaCl, polyphosphates, glucose, and the combination of those materials; and (2) palatability and consumer acceptance of the precooked, hot-processed pork loin chops infused with these ingredients. CHAPTER II LITERATURE REVIEW

Pork As A Dietary Component The image of meat as a desirable component of the human diet has been tarnished. A recent Gallup Survey revealed that two-thirds of Americans choose foods based on "good" or "bad" perceptions (National Livestock and Meat Board, 1990). A number of health problems, especially coronary heart disease, have been linked to high-fat, high- cholesterol diets. Because meat is a source of both fat and cholesterol and it is a major component of the U.S. diet, many people have questioned meat's role as a possible risk factor for the disease. Therefore, they perceive meat as a "bad" choice. Nevertheless, what we eat is not the only risk factor associated with coronary heart disease. Several factors have been identified: cigarette smoking, high blood pressure, obesity, lack of exercise, elevated blood cholesterol and, most importantly, genetic constitution of the person (Krause and Mahan, 1984).

Lean meat is a naturally advantageous option for use in nutrient-dense, yet low calorie, diets. It is a nutrient dense food, supplying a large amount of essential nutrients in relation to the calories it provides. Meat protein is high in both quality and quantity. It contains heme iron, which is much more easily absorbed by the body than nonheme iron. Meat is also a good source of highly available zinc and other minerals and vitamins such as phosphorus and Vitamins Bi, B2, Bg, B12 sirid niacin. Meats can be part of a heart-healthy diet when eaten in moderation. Choose lean cuts and prepare them in a way that does not add additional fat (American Heart Association, 1989). Of the commonly-consumed red meat species, pork is equal or superior to beef or lamb in tenderness, sometimes is more flavorful than beef or lamb, less juicy than lamb but as juicy as beef, and equal to beef and lamb in overall desirability. In all cases, pork is superior to goat and horse meat (Smith et al., 1974a). Pork is similar to other red meats in nutritive value. Although lower in iron (due to lower myoglobin content) than beef or lamb, pork is higher in thiamin and Vitamin Bg (Paul and Southgate, 1978). If meat is desirable as a dietary component and pork is as nutritious and palatable as other lean red meats, then pork is a desirable dietary component.

Hot-processing Pork Hot processing conserves about 4 0% of the energy needed for processing (Hoes et al., 1980) because inedible parts do not need to be chilled, and further heat-treated cuts do not need to be chilled and reheated to normal processing temperatures. Processing times are reduced (Mandigo, 1966b), so products in inventory (and interest on their value) are reduced also. Combined with processing yield and quality advantages (Trautman, 1964; Mandigo, 1966b; Moore et al., 1966; Weiner et al., 1966; Mandigo et al., 1979), these cost reductions make hot processing advantageous. Unfortunately, not all of the problems associated with hot processing have been resolved. Loin tenderness can be a problem (Marsh et al., 1966; Cagle and Henrickson, 1969; Marsh et al., 1972; Hinnergardt et al., 1973a, b). Hot processing affects the biochemistry of muscle tissue and its resulting yield and palatability traits. Many other factors also affect these traits.

Preslaughter Factors Affecting Yield and Palatability Genetics, environment, genetic-environmental interactions and age and sex at slaughter of pigs can alter the yield and palatability of pork. The largest source of variation in most studies (Weir, 1953; Alsmeyer et al., 1965; Rust et al., 1972a; Jones et al., 1980; Hines et al., 1980; DeVol et al., 1988) is variation among animals. Genetics. The genetic makeup of an animal, especially breed, affects the quantity and quality of the meat the animal will produce and how the animal responds to 7 environmental changes and slaughter conditions. In an excellent review. Bray (1968) discussed the effect of breed on pork quantity and quality traits. Quantity. Average backfat thickness and area of the loineye (m. longissimus) muscle are used as predictors of pork yield (Brown et al., 1951; Whiteman et al., 1953; USDA, 1970; Davis et al., 1975; Fjelkner-Moding and Pearson, 1986; DeVol et al., 1988). Of 805 carcasses from quality pork contests, Hampshires had significantly less backfat and larger loineyes than nine other breeds. Poland China hogs had significantly larger loineyes than all breeds but Hampshire. In an unpublished study from Illinois (Kauffman as quoted by Bray, 1966), Berkshires and Rolands were less fat, as measured by backfat, and Chester Whites had larger loineyes. In each of these studies, the magnitude of differences was not large—.36 cm of fat or 6.2 cm^ of loineye, and percentage lean cuts varied less than 3%. Ouality. In the contest hogs mentioned above, Duroc hogs had significantly more marbling (intramuscular fat). Kauffman et al. (1964b) reported that more marbling improved the flavor, tenderness and especially juiciness of fresh pork. Muscles from Spotted Poland China and Poland China hogs also tended to have high marbling amounts (Judge et al., 1959).

8 Color is another quality trait that has been related to palatability (Judge et al., 1960; Davis et al., 1975; Hodgson et al., 1991). As color lightened, tenderness increased, juiciness decreased, and firmness and pH of the raw loin chops decreased. Lean from hogs of Poland China and Landrace breeds tended to have a paler meat color, while Berkshire and Yorkshire lean had a darker color. In the 1962 National Barrow Show, 2 5% of the Hampshire and Poland China hogs yielded pale, soft and exudative (PSE) meat, but none of the Berkshires' muscles were PSE. Hendricks et al. (1971) showed muscles from Poland China hogs exhibited faster muscle fiber shortening (indicating more white fibers) and larger fibers than those from Hampshire hogs. In general, no one breed is superior in quality and quantity traits simultaneously, and superior individuals are present in every breed.

Environment Environmental factors can have large effects on the yield and palatability of pork. Feed source and amount and antemortem stress not only affect feed conversion, but also affect quantity and quality of the pork produced. Feeding. In a study by Greer et al. (1965), restricted feeding of a corn-soy ration improved percentage ham and loin, reduced backfat and slightly reduced marbling compared to self-feeding. Handlin et al. (1961) replaced corn with barley (less energy-dense) and noted cutability improvements in the barley-fed hogs. Feed conversion decreased with the feeding of less dietary energy. Dahl (1958, 1960) varied the ratio of saturated to unsaturated fatty acids in several feeds and found the hogs acquired fatty acid profiles very near the profile of their feed. Soft, oily carcasses resulted from feeding high levels of oils (high in unsaturated fatty acids); hard, firm carcasses resulted from feeding fats high in saturated fatty acids. Stress. Exposure of hogs to stressful situations causes yield and quality deterioration, especially if the animals are genetically stress-susceptible. Temperature changes, exhaustive exercise and handling (particularly electrical prodding) are stressors of swine (Lewis et al., 1962). Because of genetically controlled biochemical differences, some animals have higher ultimate pH and myoglobin concentrations in postmortem muscle than others (Scaife, 1955; Boles et al., 1991a, b). That pale, soft pork could be caused by transportation, fighting, heat or excitability was noted as early as 1914 in Germany (Wismer-Pedersen and Riemann, 1960). Excitability, according to Wismer-Pedersen (1959) , can induce more rapid pH fall postmortem compared to exercise or normal handling and cause the PSE condition.

10 PSE is related to a genetically controlled trait, porcine stress syndrome (PSS). PSS hogs have higher initial glycogen levels and lower myoglobin levels in muscle and generally are more excitable than normal hogs (Briskey et al., 1960a). PSE is not confined to one or several muscles; strong interrelationships between muscles of an animal in pH and color exist (Briskey et al., 1960a). Judge et al. (1959, 1960) reported another condition associated with PSS—dark, firm and dry (DFD) muscles with higher pH and lower free water than PSE muscles. Results of stress. Bendall and Wismer-Pedersen (1962) found that PSE pork was due to low water retention and sarcoplasmic protein solubility at low (physiological) ionic strengths and low myofibrillar protein solubility at higher ionic strengths. Kauffman et al. (1964a) compared pale, normal and dark pork for pH and yield properties. As muscle pH decreased, color score decreased (muscles became lighter) and expressible juice increased. PSE and DFD are caused by variations from normal pH. Loin chops from DFD pork were more juicy and tender and shrank less than those from PSE pork; after curing, the hams were equal in palatability. Deethardt and Tuma (1971b) found DFD pork less tender and more juicy than normal and PSE pork. Hams with higher pH yielded a greater percentage of their weight as cured

11 and smoked product. The pH after chilling (24 h postmortem) was highly associated with pH after aging (7 d) and cooking (Kauffman et al., 1964a). In another study (Kauffman et al., 1964b), increased marbling was more prevalent in normal than PSE or DFD pork. This finding indicated a basic difference in metabolism between normal and PSS-type hogs. This metabolic difference was shown to be due to the ratio of red (oxidative, lipolytic) fibers to white (anaerobic, glycolytic) fibers in the individual muscles. The ratio varies between muscles based on location or function, but animals with higher than normal red:white fiber ratios in one muscle also have correspondingly higher than normal red:white fiber ratios in other muscles (Beecher et al., 1965). Schmidt et al. (1970) reported a shorter rigor time course and a greater incidence of PSE in high white:red fiber ratio hogs. Elimination of stress and struggle at slaughter reduced postmortem glycolytic rates. Muscles from animals injected with curare, a neuromuscular blocker, suffered a decreased rate of pH drop (glycolysis) compared to muscles from hogs not treated with the drug (Bendall, 1966). Briskey et al. (1966) found that animals with high glycolytic rates had high glucose-6-phosphate (G-6-P) and glucose levels and low ATP, creatine phosphate (CP) and fructose diphosphate levels compared to normal animals.

12 Bodwell et al. (1966) studied Poland China and Hampshire hogs postmortem and found a stoichiometric relationship between glycogen reduction and lactic acid production in the former breed and a less clear relationship in the latter. High temperature (37^0) treatment of carcasses immediately after slaughter caused greater expression of PSE, while a low temperature (-29^0) caused rapid chilling and no PSE. Lower pH resulted in decreased protein solubility and water-binding capacity. Sink et al. (1967) found no difference in total fat content or fatty acid composition between DFD and PSE pork. These findings agreed with Kauffman et al. (1964b). The sarcoplasmic protein profiles between normal and PSE pork also did not differ (Borchert et al., 1964b). The best predictor of water-binding capacity (WBC) is final pH, according to Herring et al. (1971), and loin color is related to both pH and WBC. Cured ham purge was greatest in PSE semimembranosus and gracilis muscles and least in PSE semitendinosus and biceps femoris muscles. DFD and normal muscles had less purge than pale muscles (Topel et al., 1972). Oven-broiled PSE chops, in a study by Bennett et al. (1973), cooked slower, had higher losses during cooking, and were more tender and less juicy than DFD chops; noirmal chops were intermediate.

13 Flynn and Bramblett (1975) noted that, of PSE, normal and DFD pork, PSE chops had the highest 2-thiobarbituric acid (TBA) values after frozen storage (which indicated the development of more rancidity) and lowest juiciness scores. DFD chops were the opposite. Flavor and tenderness did not differ. In a large survey of British hogs, Kempster and Cuthbertson (1975) found no relationship between muscle pH and carcass weight or muscle temperature. High correlations between m. longissimus and semimembranosus pH and pH at 1 h or 24 h postmortem were found. Mean pH at 1 and 24 h was 6.5 and 5.8, respectively. Investigating a possible cause of PSE, Cheah and Cheah (1976) discovered an initial (within 1 h postmortem) rapid calcium efflux into the sarcoplasm, which caused rapid glycolysis early postmortem. Consumers preferred normal to light or dark chops and dark to light chops (Topel et al., 1976). Normal chops had more marbling; pale chops had very unstable color, turning gray/green after 2 to 3 d of display. Pale chops had more cooking losses (as in previous studies) and lower organoleptic acceptability. PSE lean had more drip during display, more protein, less fat, and did not differ from lean in normal chops in dry matter, ash, Na, K, Ca, Mg, P, Fe, Zn, free amino acids or total amino acids (Ewan et al., 1979). Storage and cooking losses were greater in PSE

14 compared to normal chops. Normal chops were less tender and had higher shear force values (SFV) than PSE chops. Color desirability decreased faster during display for PSE lean than for normal lean. Overcoming stress. Briskey et al. (1959a) used exercise and feeding treatments to investigate pork characteristics. Exercise (about 5 km of running) of pigs, with or without a 72-h fast, caused higher 40-min and 24-h postmortem muscle pH values and depleted glycogen stores. Fasting only did not cause differences from the control (full-fed). Water content was correlated to pH. In another study by Briskey et al. (1959b) on stress-susceptible pigs, exercised hogs were dark and firm and sucrose-fed hogs were pale and soft. Carbohydrate (glycogen) levels were depleted by exercise and restored by sucrose feeding (Briskey et al., 1960b); glycogen was converted to lactate, which lowered the muscle pH. Lewis et al. (1967) stressed normal and stress- susceptible hogs and froze the carcasses immediately after slaughter. Stress increased meat aroma, flavor, texture, tenderness, juiciness and overall satisfaction compared to unstressed controls. Freezing decreased aroma, texture, tenderness and overall scores and lactic acid production; it increased cooking losses, glycogen and ATP levels.

15 stress combined with freezing raised the pH of some muscles. Exercise also raised muscle pH in Duroc and Poland China hogs (Bowers et al., 1968); sugar feeding increased muscle glycogen, reducing sugars and browning of broiled chops. The Durocs, as previously mentioned, had higher marbling and quality (color) scores than the Poland Chinas. The reducing sugars and fat participated in the browning reactions. Hogs slaughtered in cool weather possessed darker loin color scores and more marbling than hogs slaughtered in warm weather (Judge et al., 1959). Sayre et al. (1961) immersed hogs in 5OC water for 30 to 40 min before slaughter, which caused reduced muscle glycogen levels, lactate production and muscle acidity compared to untreated hogs. Color scores increased (muscles darkened) due to treatment, but WBC was not consistently affected. At moderate humidity, hogs alternated from 32 to 21^0 exhibited more glycolysis (PSE) than hogs held at 270c before slaughter; at low humidity, no effect of temperature treatment was found (Howe et al., 1968). In a similar study (Aberle et al., 1969), hogs held at 32°C had more rapid glycolysis and pH decline than hogs held at 21^0. The latter had increased ATP and CP and decreased G-6-P compared to the former.

16 Lewis et al. (1981) used periodic electrical stimulation to reduce pH drop compared to untreated animals. Exogenous acid infusion postmortem lowered pH, while base infusion raised it in animals' muscles. Stress-susceptible Pietrain pigs, in work by Warris and Lister (1981), were improved in quality scores by antemortem treatment with a beta-receptor drug. The effects of levels of circulating catecholamines (epinephrine and norepinephrine) in the susceptible hogs were reduced by the blockers. The drug had no effect on stress-resistant (Large White) pigs, which had normal quality regardless of treatment. These results indicated that excitability and stress-susceptibility were hormonally as well as neurally related (Bendall, 1966). Epinephrine could well be the cause of Ca efflux noted by Greaser et al. (1969). Proximate composition. Besides these biochemical effects, moisture, fat and protein composition of muscle affects pork yield and palatability. Karmas et al. (1961) noted a constant (about 4:1) relationship between moisture and protein, independent of fat level. The percentage lean in a cut could be calculated as 1.5 times the moisture percentage less 11.5%. Kauffman et al., (1961) demonstrated that consumers preferred to buy cuts high in lean and low in intramuscular fat, although the well-marbled chops were preferred after tasting the chops. Increased marbling was

17 associated with higher juiciness, flavor and tenderness scores and cooked yields (Kauffman et al., 1964b; Murphy and Carlin, 1961; Davis et al., 1975). Since marbling increased as carcass backfat increased (Judge et al., 1959; Murphy and Carlin, 1961), selection of animals for leanness may result in reduced palatability (Kauffman, 1960) . Bray (1966) identified color, texture, firmness and marbling as the best indicators of quality in pork. Prediction equations to estimate expected palatability have been based primarily on marbling scores, color and firmness (Davis et al., 1975; DeVol et al., 1988; Hodgson et al., 1991) . The equations were successful in separating pork loins into three groups of expected outcome of palatability (superior, acceptable and inferior). The selection factors included biochemical traits and compositional traits. When simultaneously selecting for expected cutout and quality in pork loins. Rust and co-workers (1972b) were not as successful in stratifying loins into palatability groups. Loins that were selected for 25.8 cm^ or larger loineyes, uniform color, small degree or more marbling, and some cutability traits were less tender and juicy than randomly selected loins. In this study, marbling was not correlated to juiciness and only slightly correlated to tenderness.

18 Age and sex. Age of the animal at slaughter can be used to account for some of the differences in palatability of the pork. Carcasses from older animals have coarser and thicker collagen fibers but the same amount of total connective tissue in their muscles as carcasses from younger animals (Carpenter et al., 1963a). Animals up to 15 mo old did not differ in braised loin chop tenderness, but meat from the oldest group (36 to 42-mo-old packer sows) was tougher than that from younger groups. As animal age or weight increased, bacon tenderness decreased (Carpenter et al., 1963b). Regardless of age, lean bacon shrank less than fat bacon. In another study (Kauffman et al., 1964c), moisture content and degree of fat unsaturation decreased as age of the animal increased. Higher moisture and degree of unsaturation (softer fat) were associated with lower palatability scores. Thornton et al. (1968) reported a significant negative correlation between animal age and meat tenderness as age increased from 85 to 392 days. Sex condition of swine also affects palatability and yield. In general, barrows are fatter and have more marbling and less protein and water in the meat compared to the meat of gilts finished under the same conditions. Boars gained faster and their meat was significantly leaner and less palatable than meat from barrows, primarily due to

19 decreased aroma and flavor intensity scores and secondarily due to tenderness (Teague et al., 1964). Implantation of boars with diethylstilbestrol (DES) increased subcutaneous, intermuscular and intramuscular fat deposition; improved color scores, flavor and tenderness; and reduced or eliminated condemnations due to sex (boar) odor compared to untreated boars. Flavor and aroma scores for implanted boars were intermediate to those for barrows and untreated boars (Ockerman et al., 1981). Beta-agonists. Beta-agonists are a group of synthetic chemicals that simulate the function of epinephrine and norepinephrine. Feeding beta-agonists to growing and finishing pigs improves the feed efficiency of meat production (Ricks et al., 1984); also, they can reduce the fat content of meat and meat products. Several researchers reported that the ultimate pH of the m. longissimus muscle was not affected by feeding clenbuterol (Bekaert et al.,1987; VanWeerden 1987), or salbutamol (Cole et al., 1987). Moreover, feeding pigs ractopamine (Hancock et al., 1987; Watkins, 1990), L- 644,696 (Convey et al., 1987), clenbuterol (Ricks et al., 1984), cimaterol (Bekaert et al., 1987), and salbutamol (Cole et al., 1987; Wood et al., 1987) did not affect the muscle color of the m. longissimus muscle. Marbling scores of the m. longissimus muscle have not been affected by

20 feeding cimaterol (Jones et al., 1985) or ractopamine (Hancock et al., 1987; Merkel et al., 1988; Watkins et al., 1990). Nevertheless, Moser et al. (1986) and Ricks et al. (1984) observed a linear increase in marbling scores of pigs fed cimaterol or clenbuterol. Wallace et al. (1987) reported a nonsignificant trend toward a decrease in marbling in pigs fed the agonist L-644,969. Tenderness (shear force values) of pork loin chops was not adversely affected by feeding beta-agonists to pigs in most studies (Ricks et al., 1984; Van Weerden, 1987; Merkel et al., 1988; McKeith et al., 1988). Other researchers (Jones et al., 1985; Yen and Mersman, 1988) reported that the shear force values were significantly increased; however, the values were still within the acceptable tenderness range. McKeith et al. (1988) reported no significant effect of ractopamine on panel juiciness, tenderness, flavor intensity, off-flavor intensity and overall desirability scores of fresh pork loin chops. In summary, feeding growing and finishing pigs beta- adrenergic agonists markedly improve growth performance and carcass composition. They have little or no effect on qualitative properties and palatability of pork muscles.

21 Slaughter and Early Postmortem Procedures Treatments of animals during the slaughter procedure also can affect pork yield and palatability. Stunning and dehairing. The method used to render hogs unconscious (as required by the Humane Slaughter Act) affected the rate and extent of postmortem glycolysis (Overstreet, 1975). Of six treatments, blood and muscle pH levels remained higher in unstunned, unrestrained hogs, making them less likely to be PSE. Electrically stunned (290 V AC for 2 s) and unrestrained, unstunned hogs had higher postmortem CP levels at 1 h postmortem, which also indicated slower glycolysis and ATP hydrolysis. Use of an atmosphere of 75% carbon dioxide and 25% air required a longer stunning time and caused lower blood pH and higher blood CO2 than other treatments, but muscle traits were not affected. Restrained and unstunned, electrically-stunned (90 V AC for 10 s), and captive bolt-stunned hogs were similar to each other in blood and muscle characteristics. Captive-bold stunning was not recommended because it resulted in excessive leg contraction and struggling, which made hoisting and shackling difficult and dangerous. Atmosphere-stunned hogs struggled least. Sticking as soon after stunning as possible was recommended to assure postmortem quality (Overstreet et al., 1975) and prevent occurrence of blood-splashed lean.

22 Hair removal can be accomplished either by scalding carcasses in 60Oc water followed by scraping to pull the hairs from the follicles or by removal of the skin as well as the hair. Scalding damages the value of the skin for leather (Mowafy and Cassens, 1975) and requires hot water, equipment and labor to remove the hair (Judge et al., 1978). Skinning also requires much labor and equipment, and the technology for skinning pork has not been developed as well as in beef. Jabaay (1977) successfully skinned hogs using fine adjustments to dehider knives, correct skin temperature and some hand skinning before attachment to a mechanical hide puller. Ayres (1955) hypothesized that skinning hogs would decrease microbial loads compared to scalding because carcasses would not be exposed to feces and contamination in the scalding vat. Salm et al. (1978) found fewer bacteria on the shoulders of skinned hogs, but hams of scalded hogs had more bacteria. Carcass yields were similar, but skinned carcasses shrank less than scalded ones. Scalded carcasses were warmer before chilling, but chilled faster than skinned carcasses. Vattakavanich et al. (1981) found that hot-processed, precooked chops from scalded hogs were more tender and acceptable in flavor and overall satisfaction than chops from skinned hogs.

23 Fabrication and Processing After hair removal, evisceration, splitting and washing, carcasses are normally chilled below 3^0 before being cut into wholesale cuts. Hot processing. Weiner et al. (1966), Moore et al. (1966) and Mandigo (1966a, b) described processes in which hog carcasses were cut into wholesale cuts about 1 h postmortem. Loins frozen prerigor had greater SFV's and expressible juice percentages than normal or unfrozen prerigor loins, but prerigor-cured hams and loins were more tender than the controls (Weiner et al., 1966). Moore et al. (1966) found no difference attributable to hot or cold processing method in fresh loin tenderness. In each instance, hot processing resulted in considerable time savings. Marsh et al. (1972) noticed a 25% increase compared to controls in fresh pork SFV's caused by removal of the m. longissimus from the bone before the completion of rigor mortis and rapid cooling to 0°C. Cold shortening similar to that in beef and lamb muscle and rabbit red muscle apparently was the cause for decreased tenderness. Calcium chelators (ethylene glycol bis(beta-aminoethyl ether)- N,N,N',N'-tetraacetate, EGTA; ethylenediaminetetraacetate, EDTA) inhibited shortening and improved tenderness of hot- boned muscle (Weiner and Pearson, 1969).

24 Phosphate infusion improved juiciness and tenderness of both hot- and cold-processed loins (Hoes et al., 1980). In another study (Hinnergardt et al., 1973a, b), fresh- freezing and freeze-drying of hot pork loins resulted in decreased tenderness compared to conventional chops. Color, aroma, flavor, appearance, proximate analysis, percentage rehydration and cooking losses, WBC and pH were not affected by processing. Montgomery et al. (1977) used precooking to save energy and to attempt to prevent cold shortening in hot-boned pork m. longissimus muscle. Microwave- and conventionally- precooked, hot-processed chops were less tender than cold- processed chops. Microwave precooking caused increased cooking losses and decreased tenderness in hot- and cold- processed loin chops and roasts. Precooked roasts were less susceptible to heat-induced shortening and toughening during precooking. In a follow-up study, Sigler (1977) precooked hot loins as roasts then rewarmed them after cutting them into chops. Loins so treated were more tender than cold-processed loins similarly treated. Conventional reheating was superior to microwave reheating in all of the palatability traits of the chops. In all cases, an improvement in juiciness was needed for the process to be acceptable.

25 For fresh cuts yield, Mandigo et al. (1979) reported no differences between hot and cold processing after chilling; but before chilling, hot sides produced more rough ham and Board of Trade ham, skin and fat, less picnic shoulder, and more picnic trim. Hoes et al. (1980) similarly found that hot sides produced more fresh ham and less picnic shoulder than chilled sides, but also more spareribs and total wholesale cuts, which did not differ in the previous study. Cured and retail cut yields did not differ except for smoked bacon and total retail cuts yield (.48% and 1.58% more for hot than cold sides, respectively). Prerigor ground and seasoned fresh pork sausage had fewer aerobic mesophilic bacteria and lipolytic bacteria, higher juiciness and overall acceptance scores, and lower cooking losses than sausage prepared 6 d postmortem (Lin et al., 1979). Huffman and Cordray (1979) manufactured restructured chops from hot and cold pork. All restructured chops, whether from hot or cold pork, had more desirable eating qualities than intact boneless chops. Restructured chops with salt added were superior to other chops in all traits measured except color. Prerigor chops with salt and pre­ rigor or post-rigor chops with salt and sodium tripolyphosphate lost less weight during cooking than other chops.

26 Electrical stimulation. Forrest et al. (1966) used electrical stimulation to investigate methods to predict postmortem glycolytic rate, ultimate morphology and post- rigor pH. Excised muscles stimulated within 10 min of bleeding had responses that highly correlated with the above traits. Strength and duration of contraction was greatest in muscles low in glycolytic rate and of normal color.

For bacon production, Ockerman and Dowercial (1980) improved cure (salt and sodium nitrite) distribution by electrically stimulating carcasses before rigor. Tumbling had a similar effect. Neither method affected the total amount of cure absorbed. For normal and stress-susceptible hogs fasted for short and long time periods, electrical stimulation increased tenderness and overall palatability in chops from normal, long-fasted hogs compared to other treatments. Ham temperatures, color scores and firmness scores were lowered by electrical stimulation of stress- susceptible, short-fasted hog carcasses (Johnson et al., 1982). Overall, electrical stimulation had minimal effects on fresh meat quality, weight loss or cooked meat quality. Delayed chilling. Kastner and Russell (1975) and West (1979) reported that high-temperature conditioning, such as holding prerigor carcasses at elevated (16 to 35^0) temperatures for several hours, would speed up lysosomal

27 proteolysis and prevent cold shortening in beef and lamb. For pork, however, delayed chilling (1 h at 370c, then 24 h at 3OC) induced more rapid pH declines and more soft and exudative pork (Wismer-Pedersen and Briskey, 1961). More research, especially with stress-resistant pigs, is needed in this area. Rapid chilling. In the Wismer-Pedersen and Briskey (1961) study, carcasses chilled at -20C until a S^C internal temperature was reached had the slowest pH decline, highest WBC and improved color; chilling for 24 h at 3°C was intermediate to delay chilling and rapid chilling in pH decline and muscle quality. In excised muscles, though, chilling below 15°C caused Ca release and cold-contracture in the m. longissimus (predominantly white fibers), and all red fiber muscles studied (Bendall, 1975) similar to beef (Marsh and Leet, 1966). At 2°C, two phases of contraction were noted. The first was pH and load dependent, characterized by variable extensibility until the pH reached 6.7 or less; the second was called rigor contraction. Extensibility loss was constant, independent of pH and load. The ATP depletion delay was the cause of the first phase. Inability to relax actomyosin using ATP, which was being depleted, was the cause of the second phase.

28 Later Processing Procedures After processing into wholesale cuts, most (about 70% in the United States) pork is further processed, usually into cured or fully-cooked cuts. Curing. Curing involves the addition of salt, nitrates or nitrites and other ingredients to enhance the organoleptic and keeping quality of the meat. Curing ingredients. Historically, salt was added in great amounts to meat to preserve it by lowering water activity (Pearson, 1982) . Flavor changes resulted. We, as consumers, have adapted our taste preferences or thresholds toward salt at some level, although not as high as was necessary for preservation. Salt is added to sausages and restructured products to increase water binding (Sherman, 1961a; Wierbicki et al., 1976), fat emulsification (Sherman, 1961a, b) and particle cohesion (Huffman and Cordray, 1979; Theno et al., 1978). These effects are due primarily to the ionic strength of salt solutions causing myosin and actomyosin to solubilize. Wierbicki et al. (1976) found 1 to 3% salt solutions reduced meat shrinkage as salt level increased, no change in shrink from 3 to 5% salt, and increased shrink from 5 to 10% salt. Salt does have a detrimental effect on pork. During freezer storage, salt acted as a pro-oxidant in pork muscle in a study by Ellis et al. (1968). Salt accelerated

29 monocarbonyl production (free fatty acids and malonaldehyde) but not peroxide formation. Concerns about a possible relationship between salt intake and hypertension have led either to salt reductions or substitutions by potassium analogs of sodium-containing ingredients in processed meat products. Terrell and Olson (1981) also voiced concerns about sodium in the diet and its possible role in hypertension development. Salt is necessary for the processing and sensory characteristics of processed meat products, but consumers and the government will likely cause lowering of NaCl use in processed meats (Olson and Terrell, 1981). Because adaptation to salt flavor thresholds occurs and is reversible, thresholds for sensory perception can be moved up or down. Similarly, KCl thresholds can be moved (Terrell and Olson, 1981) or NaCl can be combined with KCl to mask flavors (Frank and Mickelsen, 1969). However, salt is added not only for flavor, but for shelf-life, yield and stability. Changes in NaCl level will change all of these factors. Perhaps calcium, magnesium or potassium chlorides can be used to replace some of the sodium chloride in processed meat (Olson and Terrell, 1981). Recently, the USDA has approved the use of these chloride salts as "tenderizers." Mickelsen et al. (1977) replaced one-half of the salt in hams with KCl and produced

30 an acceptable product. Bologna made with 2.25% NaCl had a slightly better flavor, texture and acceptability than bologna made with .75% NaCl and 1% KCl. Fresh pork sausage made with KCl was judged more bland and less acceptable than the all-salt product (Lesiak, 1980). Tolerance to and preference for KCl developed with time; again, adaptations occurred and thresholds moved. Hand et al. (1982) made restructured pork roasts with NaCl or KCl. After 1 or 4 wk of frozen storage, KCl caused more off-flavors than NaCl. Nitrogen-containing ingredients such as sodium or potassium nitrate or nitrite are thought to have been chance contaminants of curing salt used for prehistoric and early historic preservation of meat. Nonetheless, beneficial effects from inclusion have been noted. Nitrates and nitrites reduced or prevented botulism toxin formation (Hornsey, 1957), high TBA values (Freeman et al., 1982), warmed-over flavor (Hadden et al., 1975) and caused cured meat flavor (Cho and Bratzler, 1970; Hadden et al., 1975) and color (Hornsey, 1957; Taylor and Walters, 1967; Eakes and Blumer, 1975). Color development occurred regardless of the form (nitrate or nitrite) used. Mitochondrial enzymes present in the meat or in bacteria contaminating it can reduce nitrate to nitrite and nitrite to nitric oxide, the active form (Taylor and Walters, 1967). Nitric oxide reacted with hemes from myoglobin and hemoglobin to form

31 nitrosohemochrome, the pink pigment in cooked, cured meats (Hornsey, 1957). Other antioxidants (butylated hydroxyanisole and citric acid) reduced off-flavors, but not as well as nitrite and did not give cured flavor (MacDonald et al., 1980a, b). Berry and Blumer (1981) investigated replacement of nitrite with potassium sorbate in bacon because of possible carcinogenic properties of nitrite-cured meat. Sorbate was less able to prevent microbial spoilage flavors and caused chemical flavors and prickly mouthfeel. Bacon with no curing ingredients used was unacceptable in flavor. Consumers preferred nitrite- cured bacon to salt-only cured bacon even if warned of nitrite presence (DuBose et al., 1981). Polyphosphates are added to cured meats, especially hams, to improve texture and yield. Bendall (1954) noted swelling (moisture absorption) in meat of 10 to 30% caused by orthophosphate (1 P per molecule), pyrophosphate (2 P's), Calgon (4-15 P's) and phosphate glass (20-1,000 P's) A 1% salt and .5% Na pyrophosphate (SPP) solution resulted in 95% yield of cooked meat. WBC of beef (Swift and Ellis, 1956) and pork (Siegel and Schmidt, 1979) increased with additions of NaCl and SPP. The ionic strength of the solutions dissolved actomyosin and myosin and caused a spongy network of fibers. Sodium metaphosphate (SMP) improved yield of cured hams, but not color uniformity

32 compared to hams without phosphate (Mullins et al., 1958). Prerigor infusion of hams with SMP resulted in higher pH, darker color and higher glycogen levels, as well as more tender meat than the control (Kamstra and Saffie, 1959); addition of lactic acid with the phosphate improved color by lowering pH to more nearly normal. At the time of infusion, massive contraction occurred followed by relaxation. Water-infused hams contracted continuously until rigor onset. Sherman (1961b) emulsified fat with polyphosphates by forming soaps. Alkaline phosphates were more effective than more acidic ones. Salt addition broke the emulsions. Above .40 ionic strength and pH 6, water-binding linearly increased as ionic strength and pH increased. Calcium at concentrations normally present in tap water did not affect cooked ham yield (Hegarty et al., 1970). Until recently, only sodium acid pyrophosphate was permitted by federal regulations in sausages. Its use improved some flavors (salt, beef, smoke and seasoning), decreased pork flavor and aroma, fat flavor and fat mouth- feel, and had no effect on cooked yield or proximate analysis of frankfurters (Hargett et al., 1980). In all cases, phosphates improved yield and sensory attributes of country hams; at .5% use level a mixture of 5% SMP and 9 5%

33 STP was most effective of eight treatments in increasing yield (Vollmar and Melton, 1981). Sodium ascorbate and its isomer, erythorbate, are used to accelerate curing reactions (Pearson, 1982) . Display stability, but not color uniformity, is aided by its use (Mullins et al., 1958).

Curing Procedures and Eguipment Methods of incorporation of these curing ingredients varies depending on the type of product produced. Country hams are cured by rubbing the dry ingredients over the intact or skinned cut and allowing diffusion to distribute the cure (Montgomery et al., 1976). Injection of a water solution of the curing ingredients allows more rapid curing and a juicier product (Kramlich et al., 1973). Robertson and Beery (1976) pumped hams with liquid jets through .254- mm orifices at 296 kg/cm^ to 130% of their weight with no detectable difference from multineedle injection of the cure. Increased pressure of injection raised sensory panel impression of saltiness and flavor scores and total salt content of bacon compared to normal pressures (Nusbaum and Rust, 1978). Multineedle injection, vacuum tumbling while pumping, vacuum pumping and injection followed by tumbling or vacuum tumbling were not different in their effects on bacon properties (Motycka et al., 1981).

34 Tumbling and massaging. Tumbling (mixing and falling of meat pieces) is used to increase yield and cure diffusion (Ockerman et al., 1978) by causing fiber (cell membrane) disruption. Intermittent tumbling causes more disruption than continuous tumbling (Cassidy et al., 1978). Massaging is similar to tumbling in that cured meats are agitated, but is less severe than tumbling. Massaging at 5^0 at 4 rpm for 30 min forward and 30 min in reverse was positively correlated to binding, color uniformity, color intensity and moisture retention of sectioned and formed cured ham (Gillett et al., 1981, 1982). Prerigor vacuum tumbled hams underwent more complete and uniform salt distribution than untumbled or post-rigor muscles used for sectioned hams (Solomon et al., 1980). Tumbling at 0°C improved color, yield (3.45%), cohesiveness and tenderness compared to control muscles; at 25°C, yield decreased but the other factors increased above 0°C levels (Knipe et al., 1981). STP increased appearance, color, sliceability, taste, aroma, and yield of tumbled hams in a study by Krause et al. (1978a). Cure migration increased with tumbling and/or STP use (Krause et al., 1978b). Massaging and phosphates had similar effects; phosphates increased yields more than massaging (Siegel et al., 1978b). Optimal levels for extracting proteins to bind

35 chunks of muscle were 3% salt and .5% polyphosphate (Siegel et al., 1978a; Theno et al., 1978).

Other Processes Bacterial preservation. Gilliland (1980) and Bacus and Brown (1981) suggested the use of microbial starter cultures to extend the shelf-life of meat. These beneficial bacteria compete with and eliminate possible pathogens and reduce spoilage organisms. Ideally the culture would not be detrimental to meat quality. Lactobacilli inhibit other bacteria by producing hydrogen peroxide, which is lethal to bacteria such as Staphylococcus aureus, Clostridium perfringens and some Pseudomonas species that do not have a catalase for H2O2 (Gilliland, 1980). These bacteria (coagulase negative staphylococci or micrococci) aid in nitrate reduction and sugar fermentation to lactic acid to allow pH drop and drying of semidry and dry sausages (Bacus and Brown,1981). Prerigor processed products. Since Trautman (1964) reported that prerigor meat allowed better salt-soluble, heat-coagulable protein extraction than post-rigor meat, and that salt-insoluble protein (connective tissue) and water-soluble (stromal) protein emulsified little fat, interest in using prerigor meat for sausage processing has grown. Mandigo and Henrickson (1966a, b) found similar or

36 slightly increased cured ham and bacon yields and quality from prerigor processing. Acton and Saffle (1969) combined preblending (early addition of salt) and prerigor meat to produce sausage emulsions superior to controls. Hot pork used for wiener production lost more gel/water and less fat than conventional pork, but no difference in emulsifying capacity or stability was noted (Stilwell et al., 1978). Prerigor ground or ground and salted meat for processed products were less susceptible to oxidative rancidity than post-rigor meats (Judge and Aberle, 1980). Inhibition of oxidation was thought to be due to higher ultimate pH in prerigor compared to post-rigor meat and fresh pork sausage (Brerup et al., 1981). Arganosa and Henrickson (1969) reported faster cure diffusion and nitroso pigment development in prechill muscle compared to post-chill muscle. Hot-cured hams did not differ from cold- cured hams in color stability (Mandigo and Kunert, 1973). Hot hams had less "in process" contamination and anaerobic bacteria than cold hams (Cornish and Mandigo, 1974). Work by Mandigo et al. (1977) confirmed earlier work (Mandigo and Henrickson, 1966a, b) that accelerated processing had no effect on cured product yields. Prerigor salt infusion with tumbling produced superior tenderness in restructured pork, no-nitrite hams (Judge and

37 Cioch, 1979). Flavor was found to be more like fresh pork roast than cured ham. Prerigor-cured bacon had greater pigment conversion and salt residual than fresh post-rigor or frozen bacon (Amundson et al., 1981). Further technological developments in these areas are needed. Restructured products. Restructured products have been developed so that low value, less tender or less uniform carcass parts could be converted to more desirable products. Schwartz and Mandigo (1976) blended pork shoulder pieces, salt and STP to produce restructured pork chops. Salt increased TBA values, packaging loss, cooked color, aroma, flavor and eating texture compared to similar no- salt products. STP decreased cooking losses and increased TBA values, packaging loss, raw color and juiciness. Salt and STP were synergistic in action. Recommendations were made to use .75% salt and .125% STP for best effect. Campbell et al. (1977) made restructured pork patties; the yields increased as thickness of patties increased. Patties 1.9 cm thick cooked at 177^0 were highest yielding and easiest to predict final shape. Flaked patties were more cohesive and acceptable to the sensory panel than ground patties (Chesney et al., 1975). Prerigor meat retained more protein and cohesiveness than post-rigor meat. For flaked, restructured products, several particle sizes and adequate blending made the best product (Popenhagen and Mandigo,

38 1978) . Campbell and Mandigo (1978) recommended convection heating for cooking and reheating restructured patties. Hot-boned and cold-boned pork rolls were more tender with added salt than without, and were not different from each other (Furumoto and Stadelman, 1980). For frozen restructured products, Huffman and Cordray (1979) suggested use of salt levels between .5 and 1.0% to optimize palatability and structural properties. Salt from 2.25 to 3.0% and STP from .25 to .37% were optimal for most cured, flaked and formed products (Neer and Mandigo, 1977). Tsai and Ockerman (1982) introduced emulsion coating as a method to reduce time and labor associated with tumbling and massaging sectioned and formed products. Emulsion- coated product was superior in flavor and textural appeal and inferior in cohesiveness to tumbled product. Other traits such as yield, sliceability and overall acceptability were similar. Mechanical deboning. Mechanical deboning added value to pork carcasses. Bones that would require more hand labor to remove the meat on them than the meat is worth were pressed through a sieve or plate by a screw auger or hydraulic press. Softer meat passed through the sieve and most of the bone passed through another outlet. Up to 10% mechanically processed pork product (MPPP) was acceptable in frankfurters (Marshall et al., 1973); 25% or more caused

39 excessive deformation or damage in the franks and bone was more easily recognized. MPPP from a Beehive auger machine had a protein efficiency ratio of 2.0 to 2.6 compared to 3.0 for casein (Field et al., 1979). Franks prepared with up to 30% MPPP from a hydraulic machine were similar to franks made with hand-boned product (McMillan et al., 1980). More than 30% m PPP made the franks too soft and allowed bone detection by consumers. Smoked sausage with MPPP was darker and less desirable in tenderness than normal product (Ockerman et al., 1981a). Improvements in recovery and use of this product potentially could save millions of pounds of meat (Ockerman et al., 1981a).

Preservation Adequate means of preservation increases the value of any perishable product because flexibility in sales and transportation is increased. Several methods of pork preservation besides curing have been developed. Refrigeration. Refrigeration of fresh cuts markedly improves shelf life of pork compared to room temperature storage, but appearance of fresh loin chops deteriorates rapidly after several days (Smith and Carpenter, 1977). Aging of intact pork loins up to 12 d had a slight effect on palatability, but the benefits were offset by yield losses (Harrison et al., 1969). Kastner et al. (1970) used

40 chilled oil to cool pork cuts, which reduced the moisture loss to chilled air experienced in normal chilling. The authors did not mention the effect of the oil on palatability or appearance of the cuts. Hypobaric refrigeration tripled storage time compared to normal storage for long-distance shipping of pork (Jamieson, 1980).

Freezing. Freezer storage at -16 to -22°C extended pork shelf-life to several months (Noble and Hardy, 1945). Fat flavor and aroma levels decreased as storage time increased. Colder temperatures were not beneficial. Berry et al. (1971) noted deleterious effects on chop yield due to freezing. Bannister et al. (1971) froze pork chops by three at-home methods and with liquid nitrogen vapor. The cryogenically-frozen chops appeared more desirable but were equal in cooked "quality" to home-frozen chops. Blast freezing at -29^0 or below for pork patties was sufficient to kill trichinae (Trichinella spiralis). Carbon dioxide was equivalent to nitrogen in freezing ability for these thin patties. For small cuts, Ayres (1964) successfully increased the storage period of pork chops packaged by a hot-melt clear plastic bag compared to wrapped by foil. For larger cuts and longer storage times, freezer burn was a problem. Blast freezing, water glazing and poly-bagging of pallets of

41 bellies is the best combination to reduce shrinkage, trim losses and freezer burn compared to forced air or still air freezing and poly-bagging or glazing alone (Ashby and James, 1974). Frozen storage from 1 to 37 wk linearly reduced functional properties such as fat- and water- binding of pork for sausage production (Miller et al., 1980).

For pigs treated with adrenaline before slaughter, muscle pH remained higher than in untreated pigs, and oxidative deterioration during frozen storage was slower. Low pH catalyzed oxidative rancidity (Owen and Lawrie, 1975). Radappertization. Since World War II, interest in using ionizing radiation to sterilize meat has grown, especially for military use, because no refrigeration is needed. For meat, extended storage after exposure to radiation (radappertization) must be accompanied by heat inactivation of nascent proteolytic enzymes. Usually 60 to 77°C is sufficient for enzyme inactivation (Chiambalero et al., 1959) . Salt and phosphates (STP) improved both consumer and trained sensory scores for both irradiated and control restructured products (Shults et al., 1976). Increased fat level improved tenderness and increased radiation dosages to 60 kJ/kg lowered product sensory ratings (Shults et al., 1977) . Cured products were better suited to

42 radappertization because nitrite reduced the "wet dog" or other off-flavors and aromas associated with radappertized cured meats. In radappertized meat, nitrite is not necessary to prevent botulism; however, relatively high levels are necessary to prevent or reduce cured color fading during storage (Kamarei et al., 1981). Nitrate did not prevent fading because the meat or bacterial enzymes necessary to reduce the nitrate to nitrite were inactivated. Canning. Canning and retorting of meat has been practiced since the Napoleonic Wars. The high temperatures used in retorting render the meat "commercially sterile" as long as the outer package remains intact. New technology in canning has been limited to the package; the product and process have not changed much in the last century. Canning and pasteurization result in extended, but not indefinite, shelf-life at refrigerated temperatures. Freeze-drying and air drying. Lowering water activity by removing water instead of adding salt increased shelf- life of pork. Frozen meat heated on a plate under vacuum to lyophilize the water did not cause freezer burn. Drying plate temperatures above 54°C caused surface browning (Tuomy and Felder, 1964). Addition of magnesium and calcium ions before freezing increased rehydration of homogenized freeze-dried muscle fibrils and reduced rehydration of

43 diffusion-rehydrated fibrils. Increased pH (by added NaOH) before freezing was associated with increased water binding regardless of the presence of cations (Wismer-Pedersen, 1965a). Pyrophosphate produced increased swelling (water absorption) of wetted areas and EDTA induced better wetting of dry areas in the rehydration of fibrils if added before freezing and freeze drying. Only phosphate in the rehydration solution was effective in increasing water- binding in untreated freeze-dried fibrils of pork muscle (Wismer-Pedersen, 1965b). Storage of freeze-dried pork allowed considerable weight reduction compared to canned product, but 3 mo is the maximum shelf-life at 39 or 48^0 for raw freeze-dried pork. Cooked pork deteriorated almost immediately unless held at -40^0 (Townsend et al., 1978). Judge et al. (1981) compared long-distance transport of fresh-frozen versus freeze-dried pork. Freeze-dried pork was more shelf-stable, easier to handle, weighed less and was more uniform, but was less economical than frozen pork because of energy and equipment costs unless long-time storage/long-distance transport was required. Beef often is sold in the cured, dried form (dried beef and jerky), but pork seldom is sold in this form. Ockerman and Kuo (1982) described a process common in China where pork is dry

44 cured, tumbled, cooked to 63°C, thin-sliced, soaked in soy sauce, and forced-air dried to 45% of the original weight. The product was shelf-stable, especially if vacuum packaged. Demand could be increased by consumer education. Technology already developed for production of intermediate-moisture meat for pet food could be used for producing human food and merits further research.

Distribution and Display Once slaughtered and processed into wholesale or retail cuts, pork must be distributed to retail outlets and effectively displayed to induce purchase. Consumers use color to judge the acceptability of fresh and cured pork (Kropf, 1980), so the goal of distribution and display systems is to optimize color and reduce shrinkage. Smith and Carpenter (1974b, 1977) compared several pork handling systems. At a 9-d storage and shipping time, vacuum packaging and carbon dioxide (CO2) atmosphere packaging treatments of fresh loins and Boston shoulders were superior to parchment wraps in preserving meat color and reducing shrink. Carbon dioxide chilling (crust-freezing) and polyvinyl chloride wrap induced bacterial growth, off odors and lower color scores compared to vacuum or CO2 packaging but lower losses than parchment wraps.

45 Callow (1932) used CO2 atmospheres to maintain color of bacon. A mixture of 70% nitrogen, 25% CO2 and 5% oxygen reduced bacterial growth (compared to air) on fresh, boneless pork chops (Adams and Huffman, 1972). Packaging in CO2 had a reducing effect on bacterial growth during shipping, and had a residual effect on growth during subsequent fresh display (Silliker et al., 1977). In the same study, carbon monoxide (CO) reduced oxidative rancidity and greatly improved color, but concerns about the safety of CO and spoilage without color deterioration (being, in effect, adulteration) preclude its use. Huffman (1974) studied the effects of several gas mixtures on microbial growth on pork chops. High CO2 concentrations inhibited growth, air and O2 supported aerobic growth, and N2 had no effect on growth. Vacuum packaging also reduced shrinkage and prevented mold growth during aging and display of country-cured hams in a study by Kemp et al. (1981). Discoloration. Besides packaging and handling systems, display systems affect pork visual acceptance. Display, however, can be the most damaging step in distribution. Gould (1963) stated that high intensity incandescent lighting raised surface temperature of meat, causing more rapid color deterioration. Lighting that did not cause a temperature rise had no effect on pork loin chop color

46 compared to meat displayed under dark conditions. Wachholz et al. (1978) displayed normal, PSE and DFD chops to determine consumer preferences. Of the consumers purchasing chops, one-half chose normal-colored chops; one-half chose either PSE or DFD chops. Some consumers may prefer pale or dark to normal color, or else do not know the difference. Nonetheless, any research treatment that affects the color of pork should have results measured with color scores and/or hedonic scores. Kropf (1980) recommended display with 75 ft candles of light incident on meat surfaces. Fluorescent wide-spectrum or balanced lighting was preferred to reflect true colors (no washing out or excess of reds).

Pork Cookery Of all treatments given to meat to ensure palatability, cooking method can have the largest effects. Tender, juicy chops can be toughened and dried by excessive heat or inappropriate cooking methods. Tougher, drier cuts can be made tender and juicy by using appropriate cooking methods. Meat is cooked primarily to improve palatability. Again, we have adapted our tastes to prefer the flavor of cooked meat to that of raw or rare meat. Cooking also solubilizes collagen, a connective tissue protein (Deethardt and Tuma, 1971a). Cuts high in collagen are

47 tough, and are made more tender by cooking correctly. Cuts low in collagen are made less tender by cooking because the solubilization of collagen is offset by coagulation of myofibrillar proteins. In m. longissimus muscles of 68-, 82- and 95-kg hogs, meat cooked to an internal temperature of 38°C was more tender than meat cooked to 85^0, and meat at 66^0 was the least tender. At the intermediate temperature, myofibrillar coagulation occurred, but collagen solubilization had not. Hogs weighing 109 kg had highest SFV's at 85^0. Reticular fibers were greatly different in amount between animals; collagen and elastin levels were fairly constant (Deethardt and Tuma, 1971a). Cooking methods. Causey et al. (1954) compared microwave cooking, pan broiling and oven baking (roasting) of ground pork patties and loaves thawed before or during cooking. Microwave-cooked patties were least desirable in color and flavor; pan broiled patties were most desirable. Thawing during cooking also increased acceptability compared to thawing before cooking. For loaves, microwave cooking reduced yield (by forming dried crusts) and palatability because no browning occurred. Thawed loaves retained more thiamin than loaves cooked from the frozen state or else the thawed loaves lost more water which concentrated the remaining thiamin.

48 Deethardt and Tuma (1971b) tested four cooking methods for conventional roasting of pork loin roasts: open pans in a 177^C oven to ll^C internal meat temperature; covered pans in a 177^0 oven to 77^0 internally; open pans to 520c internally, then covered to 77°C internally; and covered pans to 52°C internally, then uncovered to 77°C internally. Open pans during all or the last one-half of cooking made the roasts more juicy. Open pans during all or the first one-half of cooking required less cooking time. Covered, then open cooking reduced cooking losses. Bennett et al. (1973) oven-broiled and deep-fried normal, pale and dark chops. Oven broiling produced more tender chops but required longer cooking times than deep frying. Muscle differences inherent in the animal were more apparent in oven-broiled chops. Microwaved chops in a study by Bowers et al. (1974) required less cooking time, sustained more cooking losses and retained less moisture than conventionally-cooked chops. Vitamin Bg levels did not differ due to treatment. Oven-broiled chops were more flavorful, tender and acceptable than deep fat-fried chops, but had more rancidity (Flynn and Bramblett, 1975). Chambers et al. (1982) cooked beef and pork muscles by dry or moist heat in microwave and conventional ovens. Sarcomere length, fiber diameter, percentage fat and

49 percentage connective tissue were not useful in predicting sensory attributes, especially tenderness. Safety. Meat also is cooked to kill disease-causing organisms. Carlin et al. (1969) showed conclusively that the thermal death temperature for Trichinella spiralis larvae in experimentally-infected pork was between 57 and 60Oc if roasts were conventionally cooked. Microwave-cooked roasts cooked to 75^0 in the center had remaining viable larvae, especially if loaves were long and narrow rather than round or oblong. Many underdone areas were noticed, which were the sources of the viable larvae. Many overdone areas were the cause of excessive cooking losses. Cooking temperature. After pork is made noninfective by cooking, concerns about the effect of cooking temperature on palatability and yield can be addressed. Time- temperature interaction can be extremely important. Pork loin roasts cooked in a 177°C oven to 85, 74 or 66^0 or held at 660c for 1 h varied in palatability and cooking losses (Webb et al., 1961). As final internal temperature increased, tenderness and cooking time decreased and flavor, drip losses and total cooking losses increased. Sherman (1961c) cooked salt- and phosphate-treated meat to temperatures from 25 to lOO^C. Maximum fluid retention was reached at 50OC with 2% salt plus .5% STP. This

50 temperature was higher than the temperature at which untreated meat had maximum fluid retention. Weir (1960) cooked pork roasts at 150 and 156°C. Roasts at the lower temperature cooked more slowly, were more tender and juicy, and yielded less cooked product than roasts at the higher temperature. Small pieces of pork cooked in cylinders at 60, 66, 71, 77, 82, 88, 93 and 99^0 decreased in tenderness from 60 to 66°C, then increased in tenderness with temperature (Tuomy and Lechnir, 1964). Browning and burning occurred at the higher temperatures. Time at 60^0 was not a factor in determining tenderness of the meat.

Jones et al. (1980) employed 98, 121, 149 and 163^0 ovens to cook frozen and thawed pork roasts. When cooked at 121°C, cooking losses were minimal and palatability was not different from meat cooked at the higher temperatures. At 93°C, the roasts were more tender, but were the least juicy and had the greatest cooking losses compared to meat cooked in ovens at the other temperatures. Cooking time decreased as oven or meat beginning internal temperature increased. Meat thawed during cooking was more juicy than meat thawed before cooking. Anatomical location. Position of a cut (chop within the loin) or location of the sample within the cut also affected palatability and yield. Center chops were less

51 tender than either anterior or posterior end chops (Weir, 1953) . SFV, press fluid and evaporation loss also varied from end to end of pork loins; anterior ends were superior to other locations in these traits. Allowances for these differences should be made by using the appropriate experimental designs for research studies (Mackey and Oliver, 1954). Harrison et al. (1967) found the center section to be less tender than the ends, but the center lost less fluid during cooking and had more expressible juice. The anterior ends took more time to cook because overlying muscles and fat made the cuts thicker. Alsmeyer et al. (1965) compared the position effects of dorsal, medial and lateral on palatability within chops. If measured by Warner-Bratzler shear machine, lateral samples were less tender; if measured by the STE instrument, medial cores were less tender; dorsal pieces were more tender in all cases. Within the center loin section (tenth rib to fourth lumbar vertebra), loin chops did not differ in palatability (Rust et al., 1972a). Within chops, proximal (dorsal) cores were more tender than distal (lateral) cores.

Sodium Chloride and Polyphosphates Effects on Water Holding Capacity of Meat Sodium chloride has a dramatic effect on water-holding capacity of meat. When a piece of meat block is placed in a 52 large volume of approximately 1 M NaCl, it increases volume by the uptake of both water and NaCl. It also releases proteins and smaller molecules into the NaCl solution (Offer and Knight, 1988). NaCl acts as a mild structure- breaker, and in moderate concentration tends to promote dissociation of protein assemblies by weakening the secondary bonds between molecules. Although polyphosphates usually are used in conjunction with NaCl in the curing process, they can affect the water- holding capacity of meat by themselves. Thus, raw meat slices dipped briefly into 10% solutions of polyphosphate showed a reduced rate of drip loss (Offer and Knight, 1988). Bendall (1954) showed that, with as little as .17 M NaCl, .5% pyrophosphate at a constant pH of 5.7 caused a marked increase in the volume of minced raw meat. The increase was greater than that obtained with NaCl of the same ionic strength, and greater than the sum of the volume increase obtained with .17 M NaCl or .5% pyrophosphate used separately. So a synergistic effect on swelling had occurred.

Calcium-activated Proteases Calcium-activated, neutral proteolytic enzyme activity was first reported by Huston and Kerbs (1968), and was isolated and purified from porcine skeletal muscle by

53 Dayton et al. (1976a, b). Purified calcium activated factor (CAF) from skeletal muscle retains the maximum activity in neutral pH (Suzuki et al., 1982; Calkins and Seidemann, 1988; Olson et al., 1977; Koohmaraie et al., 1984, 1986, 1988a, b), and causes partial degradation of myofibrils, attacking the Z-disk and hydrolyzing troponin-T, troponin- I, C-protein and tropomyosin (Dayton et al., 1975). Two types of CAF were isolated, based on the requirement of calcium concentration in enzyme activities (Koohmaraie et al., 1986): low-calcium-requiring CAF (CAF-I) requires only /iM of Ca 2+ (Goll et al., 1983; Koohmaraie et al., 1986), and high-calcium-requiring CAF (CAF-II) that requires mM of Ca 2+ for maximum activities. It is suggested CAF-I may be more important in living muscle cells because calcium concentration will never reach 1 mM in normal physiological condition (Goll et al., 1983). However, both CAF's have similar functions in degrading postmortem muscle tissues.

54 CHAPTER III EFFECTS OF INFUSING CALCIUM CARBONATE, POLYPHOSPHATES, SALT, GLUCOSE, AND POTASSIUM CHLORIDE ON HOT-BONED PORK SEMIMEMBRANOSUS MUSCLE

Abstract Eight crossbred 96-kg pigs were slaughtered to determine the effects of infusing 10% of an aqueous solution of 4 % glucose, 4% NaCl, 3% polyphosphates, 2% KCl (GNPK),and 3% calcium carbonate (CC) into hams via the femoral artery 1 h postmortem. Opposite side hams were cold processed (CP) after 72 h at 3°C, were not infused, and served as controls. At 2 h postmortem, semimembranosus muscles were hot boned (HB) from infused hams and four 1.5-cm chops were broiled to 77^0. Because differences (P < .05) among animals were found for proximate analysis variables, the CP ham values were used in a covariant analysis to adjust for animal differences. Infusion of solution GNPK produced more (P < .05) tenderness, moisture and sodium in the cooked muscle than solutions containing CC or only distilled water (DW). Adding CC to solution GNPK affected only the calcium content of the muscle and had no beneficial effect on tenderness. However, the calcium content of meat receiving GNPK + CC was more than 3-fold lower (P < .01) than CC because the polyphosphates bound the calcium ions and

55 precipitated them from the muscle tissue. Muscles infused with either CC or DW differed only in calcium content and cooking losses with DW being lower. Addition of calcium carbonate to infusion solutions for pork did not produce beneficial effects over a solution containing NaCl, KCl, polyphosphates and glucose.

Introduction Current economic conditions and new consumer demands, coupled with the challenge of competing with other items in the market place, make it necessary to adept a more efficient processing system in the pork industry. Hot- processing of pork and fabricating the carcass into wholesale or retail cuts can be a proper candidate to satisfy this demand. Hot processing can reduce the refrigeration, storage and transportation energy requirements (Hoes et al., 1980; Seidemann and Cross, 1982; Reagan, 1983), reduce cooler space requirements by 80% (Cuthbertson, 1980; Henrickson, 1981; Reagan, 1983), reduce "in-plant" holding times (Cuthbertson, 1980; Bowling, 1981; Reagan, 1983), increase rendering efficiency (Bowling, 1981), increase product yield (Seidemann and Cross, 1982; Reagan, 1983), and shelf life (Reagan, 1983), and improve binding quality for further processing. However, several problems associated with hot boning keep the system from

56 being widely applied by the industry — reduced tenderness, distortion of the cuts, not suitable for existing packaging systems, and the belly not suitable for bacon production (Reagan, 1983). Hot-boning of pork loins, followed by infusing a phosphate-based solution into the muscle produces a more tender, juicy, and flavorful loin chop than hot- or cold- boned controls (Hoes, 1980; Harmon et al., 1990; Wu et al., 1990). Koohmaraie et al. (1988, 1989, 1990) reported that infusing calcium chloride into whole or part of lamb carcasses improved meat tenderness dramatically at 1 d postmortem. Wheeler et al. (1991) reported the same result with hot-boned, beef round muscles. The improvement of tenderness in these studies was believed to be the activation by exogenous calcium of calpains (CDP-I and CDP- II). However, based on one preliminary study conducted in our lab, the metallic flavor of the calcium chloride was unacceptable to consumers. Calcium carbonate, the major ingredient of antacids and the calcium supplement of choice by many physicians, generally is considered safe to ingest by humans (Govoni and Hayes, 1985). It provides enough calcium ions for calpains to function in muscle tissue but has a weaker metallic flavor than calcium chloride. It may be a potential substitute for the calcium chloride.

57 The objective of this study was to determine the effects of artery pumping of a solution of 4% glucose, 4% NaCl, 3% polyphosphates and 2% KCl; a 3% calcium carbonate solution; the combination of these two solutions; or distilled water on tenderness and composition of hot-boned, pork semimembranosus muscle that was cooked immediately to produce a convenience type, precooked meat entree.

Materials and Methods Hot-boning. Eight 96-kg crossbred barrows were slaughtered and dressed by conventional procedures at the Meat Laboratory. Immediately after splitting of the carcasses (about 45 min postmortem), the ham from one randomly selected side was removed from each carcass. Each ham was randomly assigned to one of the four infusion treatments (Table 1) and infused to 110% of its fresh weight via the femoral artery. After pumping, the ham was held at 7°C for 1 h. At 2 h postmortem, semimembranosus muscles were hot processed from the infused hams and sectioned into 1.5-cm chops perpendicular to the muscle fiber direction. Semimembranosus muscles of the opposite sides were removed after the carcasses were cold processed (CP)—chilled at l^C for 72 h without any other treatment.

58 Table 1. Treatment Descriptions

Treatment Description CP Cold processed control — no infusion GNPK^ Aqueous solution of 4% glucose (w/v), 4% NaCl, 3% polyphosphates and 2% KCl CC^ Aqueous solution of 3% CaC03 (w/v) GNPK + CC^ Aqueous solution of 4% glucose (w/v), 4% NaCl, 3% polyphosphates, 2% KCl and 3% CaC03 DW^ Distilled water only

^Infused to 110% of beginning weight. Cooking. Four chops from each muscle were broiled to a 77^C internal temperature on a Farberware Open Hearth electric broiler. The chops were turned after reaching 4 0°C. The temperature was determined by inserting a .3 mm thermocouple to the geometric center of the chop. Cooking losses were recorded. Warner-Bratzler shear value. Tenderness was determined by averaging the Warner-Bratzler shear (WBS) force values of four 1.3-cm cores taken from each chop. Each core was sheared twice. Proximate analysis. The remainder of the cooked chops was cut into small pieces, frozen in liquid nitrogen and thoroughly homogenized in a Waring blender (Borchert and Briskey, 1965). Moisture was determined in duplicate by the vacuum oven procedure, total lipid was determined in

59 duplicate by ether extraction with the Soxhlet apparatus, and protein content was determined in duplicate by the Kjeldahl crude nitrogen procedure (AOAC, 1980). Mineral contents. Sodium, potassium and calcium contents were determined by atomic absorption spectrophotometry (AOAC, 1980) in duplicate on the cooked muscle. Data analysis. Data from the cold-boned sides were analyzed as random effects to determine whether differences among pigs were significant. If differences were detected, data of the hot-boned sides were analyzed as a completely randomized design with analysis of covariance using data from the cold-boned side of each hog as the covariant. Adjusted means were separated by Fisher's Least Significant Difference procedures (Steel and Torrie, 1980).

Results and Discussion Differences (P < .01) were detected among individual animals for the variables tested on the CP control muscles; therefore, data from the cold-boned hams were used to be the covariant for analysis of variances of hot-boned hams. Table 2 presents adjusted means and standard errors for the infusion treatments. Hams infused with distilled water (DW) or the calcium carbonate (CC) solution were rated less tender than hams infused with the GNPK and GNPK + CC

60 eg in o CO 00 CM o in CM CM O 4J C 0) 73 vo 0) v^ m <4-l •H u TJ o 10 Q) IS c^ I Q to <0 73 CM o CM o (0 H in H {/] n •P u S U ^ O U J3 <0 (0 (0 to ^ S CM CM (T> o w ^ o in in in CM 00 o a o - CM CO 00 n 00 CM o in to 12 0 C 0) Si i3 £1 Si u vo Si n vo o 0) u in in

0) CO oV> ap c»P :3 \ x: EH c 4-> CP CT 4J rH W O \ •H (0 O U CJ> -H 4J +J B EH O to • o U X 0> 0) •H B C ^ o <^ CO (0 in c •P a •H Q} o (0 (D Q) •H U w o •H xi to iH S • CQ o O 2 O 0 12 u M < U •- V o Si - 04 (0 ^ solutions. The effect of glucose on prerigor muscle is not well documented. However, previous work in our laboratory showed that replacing one-half of the sodium chloride in the GNPK solution with glucose produced a synergistic effect on tenderness and juiciness of hot-boned pork m. longissimus muscle and decreased the sodium content (Wu et al., 1990). Offer and Knight (1988) stated that an aqueous sodium chloride solution increased the water holding capacity and induced the swelling of meat by depolymerizing the myosin filament and weakening the M-line and Z-disc to cause myofibrillar expansion. These swelling mechanisms are further promoted by adding polyphosphates. Tenderness is positively related to the overall swelling of the meat. This swelling phenomenon probably enhanced the tenderness of the muscles infused with GNPK and GNPK + CC solutions. Adding calcium carbonate did not have a beneficial effect on tenderness of cooked muscles. The result was different from the reports of Koohmaraie (1988a, b, 1989, 1990) who injected CaCl2 into whole or part of beef and lamb carcasses and increased the tenderness dramatically 1 d postmortem. Wheeler et al. (1991) injected CaCl2 into hot-boned beef round muscles and reported an increase in tenderness of the injected muscle 1 d postmortem. They concluded that the improvement of tenderness was due to the activation of both CDP-I and CDP-II proteases by excess

62 calcium ions. However, the bitter metallic flavor caused by calcium chloride limited the usage of this technicjue. Calcium carbonate, on the other hand, is currently used as the main ingredient in antacids and generally is considered safe for human consumption. Another difference between this study and Wheeler's was that the meat was cooked at 2 h postmortem instead of 1 d, and very possibly that the time was not long enough for the proteases to fully function in the myofibril system. On the other hand, the increase of calcium ion concentration of prerigor muscle by adding CC may enhance the contraction of muscle because glycogen and other high energy compounds are not depleted. More muscle contraction would have a detrimental effect on tenderness because it increases the myofibril effects on toughness (Goll et al., 1974). Cooking losses, of great importance to meat processors and of concern to consumers, were lowest but did not differ (P > .05) from the GNPK + CC treatment for the muscles infused with the GNPK solution. However, when CC was used alone, cooking shrinkage was highest among the treatments but not statistically different from the GNPK + CC and the DW treatments. The polyphosphates and salts in the GNPK solution apparently enhanced water retention by the muscle, but this effect was lessened when calcium carbonate was present. The possible explanation of this phenomenon is

63 that increased calcium concentration in living muscle tissue actually induced the contraction of muscle cells and reduced the lattice spaces between muscle fibers and thus reduced the water-holding capacity. Another possibility is that the binding forces between calcium ions and the carboxyl groups of proteins is stronger than the hydrogen bonds between water and protein and thus expel the water from the muscle tissue (Stryer, 1988). These effects on water retention were confirmed by the moisture content of the cooked muscles—the GNPK treatment resulted in more moisture in the finished product than CC and DW treatments. Crude protein content of the muscles infused with calcium carbonate was lower than protein contents of the other groups (P < .05). This effect possibly is due to an increased denaturation of myofibrillar proteins and loss of the solubilized proteins from the tissues during cooking. The two treatments containing GNPK had lower ether extract percentages. However, the percentage of ether extract should have been affected by treatment only as the treatments varied the percentages of moisture in the

muscle. Sodium content of the cooked muscles was greatly increased by infusion of the solution (GNPK) containing sodium chloride. Other treatments did not affect sodium

64 level. This result was expected since the major source of sodium was the GNPK infusion solution. The muscle infused with the GNPK and GNPK + CC solutions had higher potassium content than the other treatments. This result was also expected due to the solutions contended potassium chloride to balance out the sodium chloride. Calcium contents of the muscles treated with CC and GNPK + CC were higher than in the muscles treated with GNPK and DW. These results was expected because these two treatments contained calcium carbonate. However, the calcium content was lower in the GNPK + CC group than in the CC group because polyphosphates bound to the calcium ions and precipitated them from the tissue.

Conclusions Hot-boned ham muscle infused with a solution containing 4% glucose, 4% sodium chloride, 3% polyphosphates and 2% potassium chloride had less cooking loss, was more tender, contained more water, less fat and more sodium than the ham muscles infused with 3% calcium carbonate or distilled water. Adding 3% calcium carbonate to the GNPK solution did not have a beneficial effect on tenderness over the GNPK solution alone. However, adding

65 calcium carbonate increased calcium content of the cooked products. Implications Results from this study indicate that the tenderness of hot-boned ham muscle can be improved by infusion an aqueous solution containing 4% glucose, 4% sodium chloride, 3% sodium tripolyphosphates and 2% potassium chloride. Muscles infused with GNPK solution had lower cooking loss and were more tender than the muscles infused with CC or DW. Adding calcium carbonate into the GNPK solution did not have a tenderization benefit on the hot-boned muscle; however, it increased the calcium content of the final product which may have a nutritional benefit to consumers in helping prevent osteoporosis.

66 CHAPTER IV ULTRASTRUCTURAL CHANGES, PALATABILITY AND CONSUMER ACCEPTANCE OF COLD- OR HOT- BONED PORK INFUSED WITH GLUCOSE, NACL, PYROPHOSPHATE AND KCL

Abstract The effect and consumer acceptance of infusing glucose, sodium chloride, sodium pyrophosphates and potassium chloride (GNPK) on the tenderness of cold- and hot-boned (CB and HB, respectively) m. longissimus muscle to produce a microwave-ready, precooked, convenient meat entree were studied. Electron microscopy showed that Z-lines of sarcomeres of muscle treated with GNPK were degraded. SDS- PAGE electrophoretograms showed that the 30,000 dalton component, a consistent indicator of tenderness of meat, appeared on HB-GNPK and all the CB treatments but did not appear in HB muscles infused with distilled water (DW) or not infused (NO). GNPK-treated roasts were lower in Hunter L, a and b values and cooking losses than the chops infused with DW or the control (NO). HB-GNPK was lowest in Hunter values. Sensory panel rating and Warner-Bratzler shear force showed that the HB- and CB-GNPK infused roasts were about 30% more tender than the CB-DW and CB-NO groups; they were about 43% more tender than the HB-DW and HB-NO roasts.

67 GNPK infused groups also were rated higher in juiciness, flavor intensity and salt intensity than DW or NO groups. The HP-GNPK group contained more fat and less protein. Both CB- and HB-GNPK chops were rated more tender, juicier and more desirable than the other chops. CB-DW and CB-NO groups were rated not different but both groups were more tender, juicier and more desirable than the HB-NO and HB-DW chops. Most (> 92.5%) consumers who evaluated the CB- or HB-GNPK chops would purchase the product if available. This study showed that the tenderization effect of HB-GNPK muscle was due to the accelerated degradation of myofibrillar proteins and that it is feasible to infuse hot-boned pork muscle with glucose, NaCl, pyrophosphates and KCl to produce a hot-boned, precooked, low salt, microwave ready, convenience-type pork product that is superior to the cold- boned products and accepted by consumers.

Introduction The advantages of developing a hot-processing system for pork include superior functional properties of pre­ rigor muscle for further product manufacturing, reduced cooler space requirement, decreased energy requirement for chilling, reduced labor requirements, shortened in-plant holding time, improved shelf life of the product, more uniform quality of the retail cuts, and reduced cooler

68 shrinkage and level of purge (Reagan, 1983). All of these effects increase the efficiency and are beneficial to the pork industry. However, accompanying these advantages are several problems such as developing an efficient chilling system for hot-processed cuts, how to maintain product tenderness, controlling cut distortion and a packaging system that can adequately protect the hot-processed products (Reagan, 1983). Dependent on their basic structures, polyphosphates can be categorized into orthophosphates and condensed phosphates. Both groups of the phosphates are extensively used in food industries (Ellinger, 1972; Dziezak, 1990). When they are used in red meat products, phosphates influence water binding, color, texture, coagulation, emulsification, and microbial growth as a result of their chemical characteristics (Ellinger, 1972; Barbut, 1989; Dziezak, 1990). Ellinger (1972) also proposed that pyrophosphate and polyphosphates can dissociate the actomyosin complexes into their major components, actin and myosin, resolve the rigor state and thus tenderize the meat. The objective of this study was to determine the effects and consumer acceptance of infusing a solution containing 4% glucose, 4% NaCl, 3% potassium pyrophosphates

69 and 2% KCl on palatability of hot-boned pork m. longissimus muscle.

Materials and Methods Infusion treatment. Twenty barrows were slaughtered at the Texas Tech Meat Laboratory in groups of four. Table 3 shows the carcass characteristics of the pigs. The carcass was completely split into two sides. Immediately after the split, the m. longissimus (loineye) muscle of one side was randomly assigned to be excised from the last lumbar vertebra to the third thoracic vertebra (hot boned, HB) and sectioned into three ecjual length roasts. The roasts were randomly assigned to one of three treatments: 1. Infusing a solution containing 4% glucose, 4% NaCl, 3% potassium pyrophosphates and 2% KCl (GNPK) to 110% of raw weight; 2. Infusing distilled water (DW) to 110% of raw weight; and 3. Control, no infusion (NO). The counterpart of the roasts on the other side of the carcass received the same treatment after being chilled 24 h at 20c (cold boned, CB). Electron microscopy. After infusion, the roasts were held at 7^0 for 1 h for the infusion solution to equilibrate. Five 2-mm diameter, 1-mm long cores were removed from each roast for transmission electron microscopy observation. The samples were fixed (primary), dehydrated and embedded in Spur resin according to

70 Table 3. Carcass Traits of The Pigs

Trait Mean ± SD Live weight, kg 95.5+2.95 Hot carcass weight, kg 62.0+2.26 Muscling score^ 5.6 + 1.00 First rib backfat thickness, cm 3.9 + .60 Last rib backfat thickness, cm 2.6 + .24 Last lumbar backfat thickness, cm 2.7 + .28 ^1 = thin, 5 = average, 8 = thick. Glauert (1975, Appendix A). Thin sections (less than 100 nm, reflected as silver color under fluorescent light) were placed on a 300-mesh, Formvar-Carbon coated grid, stained by methanolic uranyl acetate and lead citrate (Robinson et al., 1987), and observed under an Hitachi HS-9 Electron Microscope (Hitachi Instrument Co., Tokyo, Japan). Color measurement. After infusion, the roasts were held at 7°C for 1 h for the infusion solution to equilibrate and then were frozen at -40^0 for 2 months. The roasts were thawed at 20c 24 h before cooking, and the thawing losses were recorded. After thawing, a 1.5-cm chop was cut from each roast and held at 7^0 for 15 min for the muscle to bloom. Hunter L, a, and b values were measured with a Minolta CR-200b Chroma Meter (Minolta Camera Co., Ltd. 3- 13, 2-Chome, Azuchi-Mochi, Chuo-Ku, Osaka 541, Japan).

71 Cooking. The remainder of each roast was weighed and placed on a metal wire net to be cooked in a electric range oven pre-heated to 170^0 to a 71^0 internal temperature. The internal temperature was determined by inserting a .25- cm thermocouple to the geometric center of the roast. Cooking time per kilogram of sample was determined by the total time required for the roast to cook to the pre-set temperature divided by raw roast weight. Percentage cooking loss was calculated by using the difference between the weight before cooking and the weight after cooking divided by the weight before cooking (Appendix C). Warner-Bratzler shear force. After cooking, each roast was sliced into six chops 2 cm in thickness. Two 1.3-cm diameter cores were taken from the second and third chops of the anterior end of the roast to determine the Warner- Bratzler shear force (WBS). Each core was sheared twice and the WBS of each roast was the average of these four values. SDS-PAGE. About 500 mg of cooked muscle was removed from the sheared cores after the WBS was determined to perform the SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis). Samples were homogenized in a buffer solution containing .05 M Tris (pH 6.8), .7M 2- mercaptoethanol, 3% SDS and 10% glycerol according to Greaser et al. (1983, Appendix B) . The sample was run on 15% acrylamide gel under 20 mA using an SE-600 vertical

72 slab gel apparatus (Hoefer Scientific Instruments, 654 Minnesota Street, San Francisco, CA). Proximate composition. The remainder of the two chops used to determine WBS was cut into small pieces, frozen in liquid nitrogen and thoroughly homogenized in a Waring blender (Borchert and Briskey, 1965). Moisture was determined in duplicate by the vacuum oven procedure, total lipid was determined in duplicate by ether extraction with the Soxhlet apparatus, and protein content was determined by the Kjeldahl crude nitrogen procedure (AOAC, 1980). The remaining four chops were individually vacuum packaged and stored at -40°C until sensory panel evaluations were performed. Sensory panel evaluation. Two of the remaining chops from each roast were removed from the freezer and thawed at 3^C for 24 h before the panel session. At the session, chops were warmed in an Amana microwave oven (650 W) for 3 0 s per chop and served to an eight-member trained panel to determine tenderness, juiciness, salt intensity, sweet intensity and flavor intensity using an 8-point scale where 1 = extremely tough, dry and bland and 8 = extremely tender, juicy, salty, sweet, and flavorful (Appendix D). Although the overall palatability is not recommended to be evaluated by a trained sensory panel (American Meat Science Association, 1978), it can provide valuable information for

73 new products. In this study, the panelists were not trained to evaluate the overall palatability and were encouraged to evaluate the sample according to their subjective preferences. Consumer panel evaluation. The consumer panel of 120 people was selected using questionnaires (Appendices E and F) returned by Texas Tech University faculty, staff and their families. Table 4 shows the demographic data for the panel. The members varied widely in age, education and family income. Each consumer was given two randomly- selected, individually-wrapped pork chops and evaluation forms (Appendices G and H) to complete and return. The consumers were asked to eat the chops cold or reheat them in a microwave or conventional oven or on the stovetop, record the reheating method, and not add any seasonings. The pork chops were evaluated for tenderness, juiciness and overall desirability on 8-point scales arranged in a linear form that only 1 = extremely tough, dry and undesirable and 8 = extremely tender, juicy, and desirable were described at each end of the line. The consumers also were asked if they would purchase this product if it were available in the market and how much they would pay for the product. Statistical analysis. Data were analyzed by ANOVA with the GLM procedure of SAS (1990) for a randomized block design with a split-plot arrangement. Pigs were considered

74 Table 4. Demographics of the Consumer Panel

Trait N Mean Minimum Maximum Age, yr 120 37.7 14 82 Education, yr^ 120 15.2 6 22

Family income, $ 120 35,862. 7,500. 100,000.+ ^Years of formal schooling; 12 = high school graduate and 16 = college graduate. to be the block, the processing procedure was used as the whole plot, and the infusion treatment was used as the split plot. Error terms specified in the model included block X boning for the whole plot and block x boning x processing for the split plot. When main effects were significant, means were separated by the protected Least Significant Difference method with a preset experiment error rate of 5% (Steel and Torrie, 1980). The results of purchasing decisions of the consumer panelists were analyzed as category data using the FREQ procedure described in SAS (1990).

Results and Discussion Figure 1 presents typical electron micrographs of GNPK treated (a) and control (b) muscles. These micrographs show that the Z-lines of sarcomeres in GNPK-treated muscle were degraded.

75 a. GNPK treated muscle (96,400x)

b. Control muscle (78,000x) Figure 1. Electron micrograph of GNPK treated (a) and control (b) muscle.

76 Yu and Lee (1986) suggested that the degradation of Z-lines is a major factor in increasing the meat tenderness. The degradation of Z-lines is one of the major functions of calcium dependent proteases, (Dayton, 1975; Koohmaraie et al. , 1988a). The mechanism by which the GNPK solution degraded Z-lines in this study was not clear. Figure 2 shows a typical SDS-PAGE electrophoretogram of treated muscles. A distinct band for the 30,000 dalton component appears in GNPK-infused, hot- and cold-boned muscles and CB-DW and CB-NO muscles. It did not appear in HB-DW and HB-NO treated muscles. The appearance of the 30,000 dalton component is the most consistent change with postmortem aging of meat and is a good indicator of tenderness (MacBride and Parrish, 1977; Penny, 1980; Koohmaraie et al., 1988). In this study, the 30,000 dalton component appearing in all the cold-boned muscles but only in GNPK-infused hot-boned muscle indicates an accelerated postmortem myofibril degradation of hot-boned muscle after infusion. Similar results were reported by Koohmaraie et al. (1988) who infused calcium chloride into lamb carcasses and the 30,000 dalton component appeared at 24 h postmortem. Figure 3 shows that cold-boned muscle was brighter red and more yellow than the hot-boned muscle (P < .05). Infusion of GNPK further reduced all the color measurement

77 HOT-BONED COLD-BONED

- Titin - Nebulin - C-Protein - x-Actinin - Desimin

Actin T-Tropomyos in ^- Tropomyosin 30,000 dalton protein Myosin LC 1 Troponin-I Troponin-C Myosin LC 2 Myosin LC 3

GNPK DW NO GNPK DW NO

Figure 2. SDS-PAGE electrophoretogram of the effects of processing and the infusion of GNPK, DW and NO of the hot and cold-boned pork muscles on myofibrillar proteins. 78 ^999\ Hunter L l/X/A Hunter a Hunter b

O

Q < u 0^

GNPK DW NO GNPK DW NO TREATMENT

Figure 3. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on Hunter L, a and b values of the cold- and hot-boned pork muscle. Pooled SEM: L = .73, a = .22 and b = .22.

79 attributes in both cold- and hot-boned muscles (P < .05) below those of the muscles infused with DW or nothing. The reflection of the muscle's surface color is the combination effects of the amount and chemical condition of the color pigments and the absorption ability of the moisture of the meat (Cross et al., 1988). Sodium chloride and pyrophosphates have been reported to act as pro- oxidants that accelerate heme pigment oxidation and cause the meat to turn brown (Ellinger, 1972; Fox, 1987; Trout, 1989). Also, the increased water-holding capacity due to adding sodium chloride and pyrophosphate to the meat caused more absorbency of light and turned muscle darker (Fox, 1987) . Thus, the color reduction caused by GNPK infusion possibly was due to the combination of NaCl and polyphosphates increasing the protein solubility of the myofibrils and altering the light reflectance rate of the treated muscles. This result is consistent with the report by Trout (1989), who studied the effects of NaCl and sodium tripolyphosphates (STPP) on myoglobin denaturation and concluded that they increased the percentage myoglobin denaturation. DW-infused muscle produced much more thawing (Figure 4) and cooking losses (Figure 5) than GNPK infusion or no infusion in both hot- and cold-processed muscle (P < .05). Thawing losses by the GNPK-infused groups tended (P < .10)

80 ^ COLD-BONED y//X HOT-BONED

20

O 10

< H

GNPK DW CONTROL TREATMENT

Figure 4. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on thawing loss of the cold- and hot-boned pork muscles. Pooled SEM = .65. 81 ^999\ COLD-BONED y//A HOT-BONED

40 1

^ 30 --

T 1 O '/A \ X, \llCX^> v/ //.-^V '^ \ 0 20 - J// ^ F/ / / / / / ,^ yA ''7'/ 1 /// ^XXXX/ ////A 1 /// «^^000v' ^ / / yy o / / / '' / o u 10

i /// i /// i GNPK DW NO TREATMENT

Figure 5. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on cooking losses of the cold- and hot-boned pork muscles. Pooled SEM = .82. 82 to be lower than by the NO groups. The ability of muscle proteins to bind tissue and added water is very important for the quality of meat and processed meat products (Hamm, 1986; Offer And Knight, 1988; Wismer-Pedersen, 1986). Water is held in the muscle by the internal structure of cells (Offer and Knight, 1988). Only 4% of the total water in muscle is held by the myofibrillar proteins, and the majority of the water inside a muscle fiber is held by the myofibrils. Wismer-Pedersen (1986) further proposed that the water molecules are considered as located in three concentric regions around the protein molecules: (1) a primary hydration shell where the molecular orientations are dominated by the active groups on the protein surface; (2) a secondary hydration shell where these orientation effects are attenuated and which eventually merges into (3) the surrounding region of unperturbed water. He suggested that regions 1 and 2 around the protein contain 20 to 27% of the total water in meat, and there is constant interchange of water molecules between the regions. Hamm (1986) suggested that water is held in muscle by capillarity, mostly in the interfilamental spaces within the myofibrils but a substantial part in the space between the myofibrils in the extracellular space. All these hypotheses suggest that the structural arrangements of

83 myofibrils are important to the water holding ability of muscle tissue. Freezing and thawing cause extensive tissue damages through the formation and melting of ice crystals. Water holding capacity is affected by the process of freezing and thawing. Offer and Knight (1988) stated that the addition of sodium chloride and sodium tripolyphosphates drastically increased the water holding capacity of muscle. This increase of water holding capacity is the result of two effects of the added solution: (1) the binding of anion to the positively-charged amino groups of the protein and shifting of the isoelectric point of muscle proteins toward a lower pH; and (2) pyrophosphate resembles adenosine triphosphate (ATP) in the molecular arrangement of the phosphate groups in the molecule and splitting of the crossbridges between the myosin and actin filaments (Hamm, 1986; Fox, 1987; Offer and Knight, 1988). Both of the effects increase the space between myofibril lattices and capability to hold more water. On the other hand, the infusion of distilled water into the muscle without adding salt or phosphate to change the structure or chemical states of the myofibrils resulted in the highest thawing and cooking loss and lowest juiciness rating. These results showed that distilled water alone did

84 not have any effect on increasing the ionic strength of the myofibrillar proteins, and the water holding capacity did not increase with increased water content. When the muscle was frozen, the added water just stayed frozen between muscle bundles. During the thawing and cooking process these ice crystals simply melted and exited the muscle. Figure 6 showed that GNPK-infused samples recjuired about 12% less time to cook to the 71^0 internal temperature compared to the cold-boned control group, and the DW-infused cold-boned muscle took longest among the groups (P < .01). If time required to cook is an indicator of the efficiency of production, the GNPK treatment showed a potential for increasing cooking turnover rate and reducing the energy usage in producing the product. Figure 7 presents effects of treatments on juiciness ratings by the sensory panel. The thawing and cooking losses indicated that the GNPK-infused groups probably would be highest in juiciness rating and these results were found. GNPK-infused muscles were rated more tender by the consumer panel than DW-infused or NO muscles (P < .05) in both cold- and hot-boned groups (Figure 8). For DW- and NO- infused muscles, hot-boned muscles were rated tougher than the cold-boned muscles. These tenderness differences were supported by the WBS values, an objective tenderness

85 ?%?R1 COLD-BONED yyyx HOT-BONED

200

a

100 o o>^ o

GNPK DW NO TREATMENT

Figure 6. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on cooking time of the cold- and hot-boned pork muscles. Pooled SEM = 4. 86 ^9991 COLD-BONED yyy>< HOT-BONED

^ 5 -

<

09 PQ 3 -

2 -

1 -

GNPK DW NO TREATMENT

Figure 7. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on juiciness of the cold- and hot-boned pork muscles. Pooled SEM = .2. 87 5^%^ SENSORY YZZl WBS

8 «.00

7 o - 6.60 fi 6 -

0^ 5 - - 5.20 09 4 - C/9 3.80 3 Q 2 - H - 2.40

1.00 GNPK DW NO GNPK DW NO TREATMENT

Figure 8. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on tenderness rating and Warner-Bratzler shear value of the cold- and hot-boned pork muscles. Pooled SEM: tenderness rating = .18 and WBS = .17.

88 measure. The missing 30,000 dalton component in the SDS- PAGE electrophroretograms indicated that the two hot-boned treatments without GNPK infusion should have been tougher than other treatment groups. Ellinger (1972) discussed the tenderization effect of phosphates on fresh whole meat. He believed that the major effect of phosphate in tenderizing the fresh whole meat was not due to the changes in pH but due to the ability of phosphates to dissociate the actomyosin complex into actin and myosin. Fox (1987) specified that pyrophosphates and triphosphates have the ability to resemble the adenosine triphosphate in the molecular arrangement of the phosphate groups and split the actomyosin complex into actin and myosin. Knight and Parson (1988) demonstrated a progressive increase of A-band area and sarcomere length of an isolated myofibril treated with various concentrations of NaCl solution. When the same NaCl treated myofibril was exposed to pyrophosphate, the A-band material was completely lost. They concluded that a 1.0 M NaCl solution alone can cause swelling of the myofibril. When the pyrophosphate was added, the concentration of NaCl could be reduced to .5 M and still increase the sarcomere length and swelling of the myofibril. The increased tenderness of the roasts receiving the GNPK solution possibly was due to infused pyrophosphates

89 resembling adenosine triphosphate (ATP) in the molecular arrangement of the phosphate groups in the myofibril molecule. Thus, they are capable, like ATP, of splitting the actomyosin crossbridges between myosin and actin filaments (Offer and Knight, 1988). Therefore, the crossbridges between contracted myofibrils can be detached and increase tenderness. The salt intensity rating (Figure 9) was much higher (P < .05) in GNPK-infused muscles of both cold- and hot- boned groups and was expected. However, the mean ratings varied from "slightly bland" to "slightly intense." The final salt content of the GNPK muscle was about .4%. Converting to a 3-oz serving size of the product, the salt concentration would be .3 g (.12 g sodium) per serving. This amount of salt accounted for only 5% of the maximum of 6 g of salt intake per day recommended for healthy adults (NRC, 1989), is safe and enhances the flavor of the product. No differences were detected for the intensity of sweet flavor among the treated groups (Figure 10, P > .05). This result was expected because the threshold for sweetness is highest among the four basic tastes (Amerine et al., 1965). Also, glucose is rated least sweet compared to other commonly used sweeteners.

90 ^%^ COLD-BONED yyyx HOT-BONED

o H <

u % H

09

GNPK DW NO TREATMENT

Figure 9. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on sensory panel ratings of salty flavor intensity of the cold- and hot-boned pork muscles. Pooled SEM = .20.

91 ^999i COLD-BONED yyyx HOT-BONED

O

H < Pi

09 2 - U ^5 H W 1 - 09

0 GNPK DW NO TREATMENT

Figure 10. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on sensory panel ratings of sweet flavor intensity of the cold- and hot-boned pork muscles. Pooled SEM = .22.

92 The CB-GNPK chops were rated higher in flavor intensity than the HP-GNPK group (moderately intense and slightly intense, respectively), and both groups were rated higher than HB-DW and HB-NO groups (slightly bland; Figure 11, P < .05) .

As described by Hornstein and Wasserman (1987), components of meat (protein, fat, carbohydrate, non-protein nitrogen-containing compounds, minerals, trace amount of vitamins and other organic compounds) react to produce complex volatile mixtures that are characteristic of meat aroma. However, the ultimate flavor of meat may also be influenced by the compositional changes that take place in meat during postmortem storage and processing (Hornstein and Wasserman, 1987). The new substances released from the processing procedures also served as flavor precursors. In this study, several new ingredients were added to the meat and created other unknown substances within the meat. Certain reactions of these meat constituents that can occur will either enhance the flavor or cause flavor deterioration. The oxidative rancidity of the polyunsaturated fatty acids in the meat can lead to serious off-odors and off-flavors. Phosphates have been reported to prevent oxidative rancidity (Mann et al., 1989; Dziezek, 1990; Stoick et al., 1991). Ramsey and Watts (1963) reported an excellent protection of sodium

93 ^99S^ COLD-BONED yyyx HOT-BONED

5 - 1-1

< PC

o 3 - > < Pu 2 -

GNPK DW NO TREATMENT

Figure 11. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on flavor intensity of the cold- and hot-boned pork muscles. Pooled SEM = .21. 94 tripolyphosphates against the development of rancid odors and flavor of cooked sliced beef. Glucose in the infusion solution may also contribute to the flavor by reacting with the amino compounds through the Maillard reaction to form glycosylamines and act as one of the aroma constituents (Belitz and Grosh, 1987; Hornstein and Wasserman, 1987). The overall flavor intensity was a combination of many factors; however, the infusion of pyrophosphate, sodium chloride, glucose and potassium chloride and their reactions with the meat constituents may have had a positive influence on the flavor of the final product. The sensory ratings for overall palatability are presented in Figure 12. GNPK-infused groups were rated higher (moderately desirable, 6 on a 8-point scale) than the other groups (slightly undesirable, P < .05). Because panelists were not trained to evaluate the overall palatability, this rating was a subjective judgment of the panelists. Pearson coefficients showed all the palatability traits were correlated with the overall palatability (Table 5). However, tenderness rating alone accounted for over 50% of the variation (R2 = .53); tenderness, juiciness and flavor ratings simultaneously accounted for 72% of the variation in the overall palatability rating. The

95 5^5^ COLD-BONED yyy\ HOT-BONED

O 6 - % NN H 5 - <

4 -

2 3 - pq

2 -

GNPK DW NO TREATMENT

Figure 12. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on overall palatability of the cold- and hot-boned pork muscles. Pooled SEM = .2.

96 Table 5. Pearson Correlation Coefficients Between Sensory Panel Ratings Salt Sweet Flavor Tender­ Juici­ inten­ inten­ inten­ Trait ness ness sity sity sity Overall palatab. .73** .64** .53** .32** .65** Flavor intensity .41** .44** .42** .19** Sweet intensity .26** .28** .34** Salt intensity .45** .52** Juiciness .51**

** P < .01. three-variable model developed from stepwise regression predicting overall palatability is:

Rating = -.99 + .47 (Tenderness rating) + .24 (Juiciness rating) + .38 (Flavor rating) Moisture content of the CB-GNPK group was higher than for the other treatment groups (Figure 13, P < .05), and no differences were detected among the other groups. This result shows that the infusion had more effect on water holding capacity of cold-boned muscle than on hot-boned muscle. However, the fat content of the HP-GNPK chops was higher than all of the CB chops, but less than the HP-NO group (P < .05). Overall, HP chops had a higher fat content than CB chops. This high fat content was caused by a

97 55^ WATER yyyA cp FAT ASH

pa O <

U U U

GNPK DW NO GNPK DW NO TREATMENT

Figure 13. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on moisture, fat, protein and ash content of the cooked cold- and hot- boned pork muscles. Pooled SEM: moisture = .41, protein = .27, fat = .31 and ash = .07.

98 relative increase of percentage by the loss of protein in HB-GNPK muscle. The protein contents of CB- or HP-GNPK chops were lower than the DW and NO groups (P < .05). This result was expected since both NaCl and polyphosphates have the capacity to increase the solubility of myofibrillar proteins (Ladwig, 1989; Knipe et al., 1990; Knight and Parsons, 1988) and cause these proteins to be lost during thawing and cooking processes. However, data show that the GNPK solution had a more potent effect in HP muscle in solubilizing the protein than in CB muscle. This decrease of protein also contributed to the HB groups' higher fat percentage. Figure 14 shows that consumers rated CB- and HB-GNPK chops more tender (P < .01) than all other treatments. CB- DW and CB-NO chops were rated more tender (P < .01) than the HB-NO and HB-DW chops, and HB-NO chops were more tender than HB-DW chops (P < .05). These results agree with the trained sensory panel scoring. Figure 15 shows that consumers rated CB- and HB-GNPK chops juicier (P < .01) than other chops, followed by CB-DW and CB-NO chops. The HB-DW chops were rated driest (P < .01). These results also agree with the trained sensory panel ratings. Consumers also rated CB- and HB-GNPK chops more desirable (rating = 7 on a 8-point scale, P < .05)

99 ^999i COLD-BONED yyyA HOT-BONED

8

o £ 6 H < 0i 5

CO

^ 3 P ^ 2 H 1 -

GNPK DW NO TREATMENT

Figure 14. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on consumer ratings of tenderness of the cold- and hot-boned pork muscles. Pooled SEM = .13.

100 5?%^ COLD-BONED yyyx HOT-BONED

9 8

<

09 09

GNPK DW NO TREATMENT

Figure 15. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on consumer ratings of juiciness of the cold- and hot-boned pork muscles. Pooled SEM = .14.

101 ^999!i COLD-BONED yyyx HOT-BONED

0 z H <

H

<

PQ

GNPK DW NO TREATMENT

Figure 16. Least squares means for the effects of processing and the infusion of GNPK, DW and NO on consumer ratings of overall desirability of the cold- and hot-boned pork muscles. Pooled SEM = .14.

102 than CB-DW, CB-NO, HB-NO and HB-DW (rating = 5.5, 5.3, 4.0 and 3.2, respectively. Figure 16) chops. Responses to the question of whether consumers would purchase the product show that a overwhelming majority of the respondents (92.5%) would purchase the cold-boned products (Table 6). An additional 5% of the consumers would purchase the HB-GNPK chops. Just over a one-third (37.5%) of the consumers would purchase the HB-NO (37.5%)and only 7.5% would purchase the HB-DW chops. These results show consumers accepted the hot-boned, GNPK-infused pork. Figure 17 shows that consumers were willing to pay a higher price to purchase CB- and HB-GNPK chops than the other groups (P < .05). HB chops infused with DW or receiving no infusion (NO) demanded only < $2.00 and $2.40/lb, respectively. Also, table 6 showed that only a low proportion of the consumers would buy the HB-DW (7.5%) and HB-NO (37.5%) chops. These results indicate that the purchasing decision was made by consumers according to the palatability of the product. Therefore, it would benefit the industry to produce high quality food entrees to reach this market niche.

103 Table 6. Means of Consumer Responses to Purchasing and Pricing Questions About Tested Pork Loin Chops^

Cold-boned Hot-boned

Category GNPK DW NO GNPK DW NO

Question: Would you purchase this product if it was available?

No 7.5 7.5 7.5 2.5 92.5 62.5 Yes 92.5 92.5 92.5 97.5 7.5 37.5 Question: If your answer was "YES" on the previous question, how much per pound would you pay for the product?

$2 .00 - 0 2.7 8.1 2.6 100.0 76.0

$2 .00 - 2. 99 59.5 91.9 81.1 62.5 0 20.0

$3 .00 - 3. 99 29.7 5.4 10.8 23.1 0 4.0

$4 .00 - 4. 99 10.8 0 0 10.3 0 0

$5 .00 - 5. 99 ^Each consumer evaluated two chops. Means are percentages N = 40 chops per treatment.

Conclusions

Infusing an aqueous solution containing 4% sodium chloride,

4% glucose, 3% sodium pyrophosphate and 2% potassium

chloride into hot- or cold-boned pork loin muscle produce a

more palatable, precooked, convenient entree. The

increasein tenderness with GNPK infusion of hot-boned

104 COLD-BONED yyyx HOT-BONED

$5.99

S4.99

<

G $3.99 H 0 2 $3.00 Ai£i-,$2.92 $2.53 $2.56 $2.4

$2.00

$1.00 GNPK DW NO TREATMENT

Figure 17. Means for the effects of processing and the infusion of GNPK, DW and NO on consumer decisions of price they would pay for the cold- and hot-boned pork muscle product.

105 muscle may be due to the accelerated degradation of myofibrillar proteins by the GNPK solution. The ingredients in the infusion solution also increase the water holding capacity, juiciness rating and flavor of the final product.

Implications Results from this study indicate that infusing a solution containing 4% glucose, 4% sodium chloride, 3% pyrophosphate and 2% potassium chloride into hot-boned pork muscle can reduce the cooking time, enhance the palatability of the muscle and produce an acceptable product for consumers. Applying this process and the hot- boning technicjue will increase the processing efficiency, reduce energy and operation costs and thus increase the competitiveness of the pork industry.

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133 APPENDIX A: MUSCLE SAMPLE PREPARATION FOR TRANSMISSION ELECTRON MICROSCOPY

134 PREPARATION OF MUSCLE SAMPLE FOR ELECTRON MICROSCOPY

I. Sample preparation Excise 2-mm X 1-mm of muscle in 4% glutaldehyde in .1 M phosphate buffer pH 7.2. II. Primary fixation 4% glutaldehyde in .1 M phosphate buffer pH 7.2 for 4 h. Wash with 0.1 M phosphate buffer for 20 min. (repeat 3 times). III. Secondary fixation .1% OSO4 in .1 M phosphate buffer, pH 7.2, for 2 h. Wash with .1 M phosphate buffer, pH 7.2, for 20 min (repeat 3 times). IV. Dehydration 10% ethanol with CUSO4 for 20 min. 30% ethanol with CUSO4 for 20 min. 50% ethanol with CUSO4 for 20 min. 70% ethanol with CUSO4 for 20 min. 95% ethanol with CUSO4 for 20 min. 100% ethanol with CUSO4 for 1 h (repeat 3 times). V. Infiltration 3:1 ethanol : resin for 2 h. 1:1 ethanol : resin for 2 h.

135 1:3 ethanol : resin for 2 h. Pure resin for 4 h (repeat 2 times). VI. Embedding Embed in the mold, look for the orientation of the muscle VII. Polymerization Polymerize in 70°C for 36 to 48 h. SOLUTIONS 1. Phosphate buffer Sorensen's buffer Solution A: .1 M KH2PO4 (13.76g/l). Solution B: .1 M Na2HP04. 2H2O (18g/l). To make pH 7.2 buffer solution add Solution A 28.5 ml. Solution B 71.5 ml. 2. 4% Glutaldehyde in 0.1 M phosphate buffer Dilute 8% glutaraldehyde (acjueous) with ecjual amount of 0.2 M phosphate buffer. 3. Embed in Spur resin.

136 APPENDIX B: PREPARATION OF MUSCLE SAMPLE FOR SDS-PAGE

137 PREPARATION OF MUSCLE SAMPLE FOR SDS-PAGE

1. .5 g muscle sample, free of fat and connective tissue. 2. Homogenize for 30 s at full speed with a Polytron in 4.5 ml of .05 M Tris buffer (pH 7.0) 3. Centrifuge for 10 min, discarded the supernatant 4. Rehomogenize the pellet for 5 sec in 4.5 ml Tris buffer (pH 7.0). 5. Add 10 ml SDS sample buffer (4% SDS, 1.5% mercapentoethanol and.06 M Tris, pH 6.8) to the homogenate. 6. Put the test tube in hot water bath for 12 min. 7. Completely solubilize the pellet and cool to room temperature. 8. Dilute the solution with sample buffer to obtain a final protein concentration of 4 mg/ml.

138 APPENDIX C: DATA SHEET FOR COOKING CHARACTERISTICS

139 Panel No. Date / / Time AM PM

Sample No. 1 Code 1 1 Weight, g 1 Frozen Before cooking % Thaw loss After cooking 1 1 % Cooking loss ! i Time in Time out 1 Cooking time, min Internal Temperature

WBS, lb Core 1 \ 1 \ Core 2 1 1 1 i Core 3 1 I Core 4 Average WBS

140 APENDIX D: TRAINED SENSORY PANEL EVALUATION FORM

141 NAME PANEL SESSION DATE / / TIME_ AM PM

SAMPLE JUICI­ TENDER­ SALT SWEET FLAVOR OVERALL NO. NESS NESS INTEN­ INTEN­ INTEN­ PALATA­ SITY SITY SITY BILITY i 1 ,

JUICINESS TENDERNESS SALTY FLAVOR INTENSITY 8 EXTREMELY JUICY 8 EXTREMELY TENDER 8 EXTREMELY INTENSE 7 VERY JUICY 7 VERY TENDER 7 VERY INTENSE 6 MODERATELY JUICY 6 MODERATELY 6 MODERATELY TENDER INTENSE 5 SLIGHTLY JUICY 5 SLIGHTLY TENDER 5 SLIGHTLY INTENSE 4 SLIGHTLY DRY 4 SLIGHTLY TOUGH 4 SLIGHTLY BLAND 3 MODERATELY DRY 3 MODERATELY TOUGH 3 MODERATELY BLAND 2 VERY DRY 2 VERY TOUGH 2 VERY BLAND 1 EXTREMELY DRY 1 EXTREMELY TOUGH 1 EXTREMELY BLAND

SWEET FLAVOR FLAVOR INTENSITY OVERALL PALATABILITY 1 INTENSITY ! 8 EXTREMELY 8 EXTREMELY 8 EXTREMELY PALATABLE | INTENSE INTENSE 1 7 VERY INTENSE 7 VERY INTENSE 7 VERY PALATABLE 6 MODERATELY 6 MODERATELY 6 MODERATELY PALATABLE |

INTENSE INTENSE 1 5 SLIGHTLY INTENSE 5 SLIGHTLY 5 SLIGHTLY PALATABLE | INTENSE 4 SLIGHTLY BLAND 4 SLIGHTLY BLAND 4 SLIGHTLY NON- PALATABLE 3 MODERATELY BLAND 3 MODERATELY 3 MODERATELY NON- i BLAND PALATABLE 2 VERY BLAND 2 VERY BLAND 2 VERY NON-PALATABLE 1 EXTREMELY BLAND 1 EXTREMELY BLAND 1 EXTREMELY NON- PALATABLE

142 APPENDIX E: MEMORANDUM FOR RECRUITING CONSUMER PANEL

143 TEXAS TECH UNIVERSITY COLLEGE OF AGRICULTURAL SCIENCES DEPARTMENT OF ANIMAL SCIENCE

To: Texas Tech University Employees From: Kenny Wu, Meat Science Graduate Student Meat Laboratory Mail Stop 2162

I am conducting a project concerning with the quality of newly developed precooked pork chops on their tenderness, juiciness, saltiness, sweet intensity, and flavor desirability at the consumer's table. I need a panel of consumers to help evaluate these chops in their homes. If selected to be a member of this panel of consumers, you will be provided fully cooked pork loin chops for each member of your family over 15 years of age. These pork chops can be served cold or warm. Each chop must be evaluated for tenderness, juiciness, saltiness, sweet intensity, and flavor desirability on forms which will be provided. The frozen pork chops can be picked up from the retail counter of the Meat Laboratory at your convenience. If you are willing to participate in this study, please complete and return the form in the campus mail. If you are selected as part of this representative panel, you

144 will be notified of the time for you to pick up the pork chops. All information provided will be confidential and will be used only in calculating averages and in other statistical analysis.

145 APPENDIX F: RETURN FORM FOR CONSUMER PANEL

146 RETURN FORM FOR PARTICIPATING IN THE PORK CHOP QUALITY STUDY

To: Kenny Wu Meat Laboratory Mail Stop 2162

Please print.

Your Name (Last) (First) (Middle) Campus Address Home Address Home Phone No. Annual Family Income $ Please provide the following information for household members who are at least 15 years of age:

HIGHEST EDUCATION, FIRST INITIAL SEX AGE 12 = HIGH SCHOOL GRADUATE

147 APPENDIX G: INSTRUCTION FORM FOR SAMPLE PREPARATION AT CONSUMER'S HOME

148 Dear consumer: Thank you for participating in this pork cjuality project. The following guidances will help you to properly complete this study. Please read carefully before serving the samples. You have received:

1. Two frozen boneless pork chops for every family members with a random number coded on each chop. 2. One evaluation form for each chop for every family member participating in the study. Reheating: 1. The pork chops are fully cooked so they can be eaten cold or reheated. 2. Please do not salt or season the pork chops until evaluation forms have been completed because differences in seasoning can influence your evaluations. 3. If you prefer to reheat the chops, please follow the following instructions for the best results: a. Microwave oven: Set on "HIGH" for 30 seconds.

b. Conventional oven: Preheat the oven at 350 F and warm the chops for 5 to 8 minutes. c. Stovetop: Warm the chops for 5 to 10 minutes.

4. Please do not overheat the chops because overheating will dry them out. 5. Please record your reheating method on the evaluation form.

149 APPENDIX H: PORK CHOP EVALUATION FORM FOR CONSUMER PANEL

150 PORK CHOP EVALUATION FORM

Sample Number (3 digit number) 1. Please circle the reheating method of the pork chop served, if it was reheated. 1) Not reheated 2) Microwave 3) Conventional Oven 4) Stovetop 5) B-B-Q Grill 2. Please rate the tenderness of the chop (please circle the rating on the line): 1 2345678 Extremely Extremely tough tender 3. Please rate the juiciness of the chop (please circle the rating on the line): 1 2345678 Extremely Extremely dry juicy 4. Please rate the desirability of the chop (please circle the rating on the line): 1 2345678 Extremely Extremely undesirable desirable 5. Would you purchase this product if it was available? 1) YES 2) NO If your answer is YES on the previous cjuestion, how much would you pay for the product for each pound (circle the most close one)? 1) Below $ 2.00, please Specify 2) $ 2.00 - 2.99 3) $ 3.00 - 3.99 4) $ 4.00 - 4.99 5) $ 5.00 - 5.99

After you finish, please return to Kenny Wu Meat Lab, Mail Stop 2162

Thank You for Your Help!!1 151