J. Anim Sci.
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Rosenvold, K.
Right arrow Articles by Andersen, H. J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Rosenvold, K.
Right arrow Articles by Andersen, H. J.
J. Anim. Sci. 2003. 81:130-134
© 2003 American Society of Animal Science

Dietary manipulation of pro- and macroglycogen in porcine skeletal muscle1

K. Rosenvold*, B. Essén-Gustavsson{dagger} and H. J. Andersen*,2

* Department of Animal Product Quality, Danish Institute of Agricultural Sciences, P.O. Box 50, DK-8830 Tjele, and and {dagger} Department of Large Animal Clinical Sciences, Faculty of Veterinary Medicine, Swedish University of Agricultural Sciences, P.O. Box 7018, SE-750 07 Uppsala

2 Correspondence:
(phone: + 45 89 99 12 41; fax: + 45 89 99 15 64; E-mail:
henrik.andersen{at}agrsci.dk).


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 Implications
 Literature Cited
 
The aim of the study was to investigate how feeding-induced changes in muscle glycogen stores affect the ratio of between the glycogen pools, pro- and macroglycogen. Pro- and macroglycogen content were determined in longissumus muscle from slaughter pigs subjected to a feeding strategy known to reduce total glycogen stores. Furthermore, early postslaughter glycolysis of the two glycogen forms was determined. The feeding strategy involved a diet (GLYRED diet) with a low digestible carbohydrate (5%)/high fat (18%) content, which was fed to the pigs the last 3 wk before harvest. A control group was fed a standard pig diet (49% digestible carbohydrate/5% fat). Total glycogen was reduced by 48 µmol/g dry weight (d.w.) in GLYRED pigs during the 3-wk feeding period. This was mainly due to a reduction in macroglycogen of 42 µmol/g d.w. During postmortem glycolysis the proglycogen appeared to be degraded in favor of macroglycogen. Moreover, total glycogen was degraded to a larger extent in muscle from the control pigs compared with muscle from GLYRED pigs. This difference was due to a significantly greater degradation of proglycogen in the control pigs. In conclusion, the results support earlier studies suggesting that proglycogen and macroglycogen are different glycogen pools that have different functions. Furthermore, the results show that the muscle glycogen pools can be manipulated through diet and that proglycogen is degraded in favor of macroglycogen under the anaerobic conditions postmortem.

Key Words: Glycogen • Glycolysis • Meat Quality


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 Implications
 Literature Cited
 
Meat quality is related to the rate and the extent of the postmortem glycogen metabolism. Consequently, the muscle glycogen stores at the time of harvest are decisive for meat quality. More than half a decade ago it was recognized that muscle glycogen from rats could be partitioned into "free" and "bound" moieties, depending on the degree of "binding" with proteins (Willistätter and Rohdewald, 1934; Wajzeer, 1939; Meyer and Jeanloz, 1943). Later it was recognized that these glycogen pools could be characterized on the basis of their solubility in acid (Bloom et al., 1951). The ratio between these two glycogen pools has also been reported to be decisive for the rate of anaerobic glycogen metabolism (Wismer-Pedersen and Briskey, 1961; Charpentier, 1966).

The two glycogen forms are known as macroglycogen (approximately 104 kDa), which is acid-soluble and has a high ratio of carbohydrate to protein (approximately 0.4% protein) and proglycogen (smaller or equal to 400 kDa), which is acid-insoluble and has a low ratio of carbohydrate to protein (approximately 10% protein) (Lomako et al., 1991, 1993).

The macroglycogen fraction in humans and rats is found to increase with high muscle glycogen concentrations (Jansson, 1981). Both proglycogen and macroglycogen are suitable energy substrates during muscle contraction (Derave et al., 2000), with macroglycogen mainly being metabolized during aerobic exercise (Asp et al., 1999) and proglycogen mainly during anaerobic conditions (Graham et al., 2000).

In pigs, muscle glycogen stores at the time of harvest can be reduced through strategic finishing feeding (Rosenvold et al., 2001a, b; 2002). Moreover, such a diet-induced reduction in muscle glycogen stores was both found to reduce the rate of early postmorten glycogen metabolism in the muscle and affect the meat quality. The purpose of the present study was to explore possible changes in proglycogen and macroglycogen as a result of strategic feeding procedure compared with a commercial diet (control).


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 Implications
 Literature Cited
 
The experiment was approved by The Danish Inspectorate of Animal Experimentation, and the pigs were treated in accordance with the guidelines outlined by the same authority.

Animals and Management
The 16 female pigs used in the present study were reared at the experimental farm of the Research Centre Foulum, the Danish Institute of Agricultural Sciences. They were crossbreeds between Danish Landrace x Danish Yorkshire sows and Duroc boars, and they were all noncarriers of the Halothane gene.

Eight pigs were fed a standard grower-finishing diet (control diet) and eight pigs were fed a diet known to reduce muscle glycogen stores in pigs (GLYRED diet) (Rosenvold et al., 2001a, b; 2002). The feedstuff and diet compositions, analyzed as described in Rosenvold et al. (2001b), are presented in Table 1Go.


View this table:
[in this window]
[in a new window]
  		Table 1. Ingredients and chemical composition of control and GLYRED diets. Ingredients in percentage and chemical composition in percentage of (DM)
 
The pigs were housed in individual pens from a live weight of 25 kg and fed the standard grower-finishing diet until a live weight of approximately 80 kg, when the strategic feeding period was initiated. The pigs were fed partially ad libitum to a standard scale (feed units per day according to live weight of the pig) of Research Centre Foulum (Madsen et al., 1990a, b). The strategic feeding was initiated by a 1-wk acclimatization period (d 1 to d 7) gradually changing from the standard grower-finishing diet into 100% of the GLYRED diet. Control pigs were fed the control diet during the entire strategic feeding period (d 1 to d 22). The pigs were harvested on d 22 after feed withdrawal of 12 to 15 h.

Slaughter Procedure
On the day of harvest (d 22), the pigs were transported from the rearing house to the experimental slaughter plant (200 m). The pigs were stunned by 85% CO2 for 3 min, exsanguinated, scalded at 62°C for 3 min, cleaned, and eviscerated within 30 min.

Muscle Biopsy Sampling and Glycogen Determination
Biopsy samples for determination of proglycogen, macroglycogen, and total muscle glycogen were taken in longissimus muscle of all pigs using a spring-loaded biopsy instrument (Biotech, Slovakia) on d 1, 1 h after feeding, prior to the diet change in the strategically fed group and on d 21, 1 h after feeding, and on d 22 immediately prior to stunning. The biopsies were taken at the last rib on d 1 and at a site approximately 5 cm from the first biopsy in the caudal or cranial direction on d 21 and d 22. Moreover, 45 min postmortem biopsies were taken at the last rib. Immediately after sampling the biopsies were frozen in liquid nitrogen and stored at -80°C until further analysis. All biopsy samples were taken in the right-hand side muscle.

Proglycogen and macroglycogen were determined in freeze-dried muscle tissue dissected free of visible blood and connective tissue. One to two milligrams of muscle was extracted in 200 µL of ice-cold 1.5 M perchloric acid for 20 min. After centrifugation at 3,200 x g at 4°C for 10 min, 100 µL of the supernatant was removed and used for analysis of macroglycogen. An additional sample of the supernate was removed for analysis of free glucose. The remaining supernatant was discarded and the pellet was kept and used for analysis of proglycogen content. Glycogen was hydrolyzed by the addition of 1 M HCl to the proglycogen and macroglycogen sample tubes that were sealed and heated to 100°C in a water bath for 2 h. Glucose was then analyzed in all the samples with a fluorometric technique as described by Lowry and Passonneau (1972). The free glucose in the supernatant was subtracted from the boiled macroglycogen sample in order to get only the macroglycogen fraction. Total glycogen was calculated as the sum of proglycogen and macroglycogen.

Meat Quality Indicators and Attributes
The pH (pH45 min) and temperature (T45 min) were measured 45 min postmortem in the left-hand side longissimus muscle at the last rib. The temperature was measured with a Testo 110 thermometer (Testo, Germany) and the pH was measured with a pH meter (Radiometer PHM201, Denmark) equipped with a probe type glass electrode (Metrohm LL Glass Electrode WOC, Switzerland) calibrated in pH 4.01 and 7.00 IUPAC buffers (Radiometer, Denmark) at 35°C. After pH and temperature measurements the carcasses were placed in a chill room at 4°C. At 24 h postmortem, pH (pH24 h) was measured as described for measurements taken 45 min postmortem—except for calibration of the pH electrode, which was done at 4°C (carcass temperature).

Water-holding capacity was measured in a chop (longissimus muscle) taken 10 cm from the last rib in the cranial direction according to the bag method described by Honikel (1998). The meat chop with a weight of approximately 100 g was trimmed and weighed 24 h postmortem. Subsequently, the sample was placed in a net and then hung in an inflated plastic bag for 48 h at 4°C, after which it was weighed again. Drip loss was calculated as the difference in weight before and after hanging.

Data Analyses
The statistical analysis was carried out with Statistical Analysis System version 8.01 (SAS Institute, Cary, NC, USA). The MIXED procedure was applied when calculating the least squares means and standard errors of means of all the variables. Least squares means were considered significantly different if P < 0.05. A model, including the fixed effects of diet and sampling time as well as their interaction and the repeated effect of time with animal as subject, was applied for proglycogen, macroglycogen, and total glycogen and the change in proglycogen, macroglycogen, and total glycogen calculated for each period: d 1 to d 21, d 21 to d 22, and d 22 to 45 min postmortem. A model including the fixed effects of diet was applied for the pH and temperature measurements as well as water-holding capacity.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 Implications
 Literature Cited
 
The contents of proglycogen, macroglycogen, and total glycogen during the strategic feeding period (d 1, d 21, and d 22) and 45 min postmortem and the changes in glycogen between the different measurement points are shown in Table 2Go and Figure 1Go, respectively. There were no significant differences in muscle glycogen contents (P = 0.623, 0.119, and 0.576 for total glycogen, proglycogen, and macroglycogen, respectively) at the beginning of the feeding period (d 1) between the two dietary groups.


View this table:
[in this window]
[in a new window]
  		Table 2. Total glycogen, proglycogen, and macroglycogen (µmol/g dry weight) measured in longissimus muscle biopsies at the start of the feeding experiment (d 1), after 3 wk feeding the day prior to harvest (d 21), immediately prior to stunning (d 22) and 45 min postmortema
 


View larger version (16K):
[in this window]
[in a new window]
  		Figure 1. Change in proglycogen, macroglycogen and total glycogen calculated for the three periods. A) during the experimental feeding period (d 1 to d 21); B) during fasting, transport, and preharvest handling (d 21 to d 22); and C) from harvest to 45 min postmortem (d 22 to 45 min postmortem). Different letters for the individual glycogen forms indicate significant differences (P < 0.05). *P-value for {delta}macroglycogen d 1 to d 21 = 0.086.

 
Total muscle glycogen in samples from the GLYRED pigs was reduced by 48 µmol/g d.w. during the 3-wk feeding period (d 1 to d 21). In contrast, total glycogen did not change in the control group during the trial period (d 1 to d 21). Subsequent overnight fasting, transport, and preharvest handling (d 21 to d 22) induced no significant changes in total muscle glycogen in the two dietary groups (P = 0.410 and 0.617 for control and GLYRED pigs, respectively). Forty-five minutes postmortem the total glycogen content was reduced by 67µ mol/g d.w. in the control pigs, while a nonsignificant reduction (P = 0.081) of only 44 µmol/g d.w. was registered in the GLYRED pigs compared with the level prior to harvest.

The large reduction in total glycogen during the 3-wk strategic feeding period in the GLYRED pigs was caused by a reduction in the macroglycogen content of 42 µmol/g d.w. (P = 0.086). No significant changes in macroglycogen were observed during fasting, transport, and preharvest handling (d 21 to d 22) or the first 45 min postmortem. This was independent of applied feeding strategy and initial glycogen status.

In contrast to macroglycogen, the proglycogen content did not change during the strategic feeding period (d 1 to d 21) or during fasting, transport, and preharvest handling (d 21 to d 22) in any of the two experimental groups. However, the proglycogen content was significantly reduced 45 min postmortem (P < 0.001 and P = 0.001 for control and GLYRED pigs, respectively) with the reduction being significantly larger (57 µmol/g d.w.) in muscle from the control pigs compared with muscles from the GLYRED pigs (30 µmol/g d.w.) (P = 0.011).

The value pH45 min was numerically higher in muscle from GLYRED pigs compared with pH45 min in muscle from control pigs. Values for T45 min and pH24 h were identical in muscles from the two groups, while water-holding capacity was numerically lower in muscle from GLYRED pigs (Table 3Go).


View this table:
[in this window]
[in a new window]
  		Table 3. pH measured 45 min (pH45 min) and 24 h (pH24 h) postmortem, temperature measured 45 min (T45 min) postmortem, and water-holding capacity (WHC) of longissimus measured in control and GLYRED pigsa
 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
Implications
 Literature Cited
 
The purpose of this study was to explore possible changes in proglycogen and macroglycogen in longissimus muscle from pigs fed a muscle glycogen reducing diet (GLYRED diet) and compare this with muscle from pigs fed a standard diet. The GLYRED pigs had reduced total glycogen stores at the time of harvest, and this was mainly caused by a reduction in the macroglycogen pool. The results also showed that the rate of glycolysis was reduced in GLYRED pigs during the first 45 min postmortem primarily due to a slower metabolism in the proglycogen pool.

To the authors’ knowledge, this is the first study to show that the proglycogen and macroglycogen pools can be changed through dietary manipulation without prior glycogen depletion. It should be kept in mind that the pigs were physically inactive during the experimental period, as they were kept in single pens. In addition, the pigs were still growing and gained approximately 1 kg/d during the experimental period (from approximately 80 kg to 100 kg). The growth rate was identical in the two experimental groups and indicates that the reduction in muscle glycogen was not a result of fasting, but a direct result of the dietary manipulation (Rosenvold et al., 2002).

During the experimental period the GLYRED diet induced a reduction in the total muscle glycogen stores as also shown in earlier experiments (Rosenvold et al., 2001a, b; 2002). The reduction in total muscle glycogen was mainly caused by a reduction in the macroglycogen pool. This result is complementary to results in humans and rats showing that the macroglycogen fraction is decreased when total glycogen content is low (Jansson, 1981; Adamo and Graham, 1998; Hansen et al., 2000). Initial glycogen levels in pigs before being fed the GLYRED diet must be expected to be high as the control diet contains 49% digestible carbohydrate, and the biopsies were taken 1 h after feeding.

Total glycogen content was reduced 45 min postmortem in both dietary groups compared with the preharvest situation. The greater reduction in total glycogen in the control pigs was caused by a significant reduction in proglycogen. The fact that proglycogen was degraded in favor of macroglycogen is in agreement with previous results (Charpentier, 1966). In addition, proglycogen and total glycogen stores has been reported to be higher in pigs displaying rapid glycolysis (Wismer-Pedersen and Briskey, 1961). The reduced glycolytic rate in muscle from GLYRED pigs was reflected in higher pH45 min value (0.1 unit). This difference was not significant, which might be due to the limited number of animals. However, the differences were identical to those observed in studies including a larger number of pigs (Rosenvold et al., 2001a; 2002), where pH45 min was found to be significantly different and resulted in improved meat quality measured by water-holding capacity.

In conclusion, the data show that the muscle glycogen pools can be manipulated through diet and that proglycogen is degraded in favor of macroglycogen under the anaerobic conditions postmortem. Moreover, the results support earlier studies suggesting that proglycogen and macroglycogen are different glycogen pools with different functions. Finally, further studies of the location and the regulation of the pro- and macroglycogen pools in porcine skeletal muscle are needed to obtain a fundamental understanding of the significance of postmortem glycolysis for pork quality.


   Implications
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Implications
Literature Cited
 
Present data shows that diet composition not only affect the muscle glycogen pool, but also the ratio between the subpools, pro- and macroglycogen. It appears that the ratio between pro- and macroglycogen is of importance for the rate of postmortem glycolysis known to be critical for the meat quality. Consequently, future studies on the influence of diet composition on meat quality should include investigations on the effect on both total muscle glycogen and the ratio between pro- and macroglycogen to ensure optimal use of strategic feeding as a tool in meat quality control.


   Footnotes
 
1 The authors wish to thank Robert Jonasson, Camilla Bjerg Petersen, Ivan Nielsen, and Jens Askov Jensen for technical assistance. Moreover, the authors are grateful to the Danish Directorate of Developments for partly supporting H. J. Andersen (Product Quality Project). Acknowledgments are given to the Pig Levy Fund and the Ministry of Food, Agriculture, and Fisheries for financing this work. Back

Received for publication March 22, 2002. Accepted for publication August 6, 2002.


   Literature Cited
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Implications
 Literature Cited
 


Adamo, K. B., and T. E. Graham. 1998. Comparison of traditional measurements with macroglycogen and proglycogen analysis of muscle glycogen. J. Appl. Physiol. 84:908–913.[Abstract/Free Full Text]

Asp, S., J. R. Daugaard, T. Rohde, K. B. Adamo, and T. E. Graham. 1999. Muscle glycogen accumulation after a marathon: Roles of fiber type and pro- and macroglycogen. J. Appl. Physiol. 86:474–478.[Abstract/Free Full Text]

Bloom W. L., G. T. Lewis, M. Z. Schumpert, and T. M. Shen. 1951. Glycogen fractions of liver and muscle. J. Biol. Chem. 188:631–636.[Free Full Text]

Boisen, S., and J. A. Fernández. 1997. Prediction of the total tract digestibility of energy in feedstuffs and pig diets by in vitro analyses. Anim. Feed Sci. Technol. 68:277–286.

Briskey, E. J. 1964. Etiological status and associated studies of pale, soft, exudative porcine musculature. Adv. Food Res. 13:89–178.[Medline]

Charpentier, J. 1966. Relation entre la teneur du muscle de porc en desmo- et lyo-glycogènes et l’intensité de la glycogénolyse post mortem. Ann. Zootech. 15:367–371.

Derave, W., S. Gao, and E. A. Richter. 2000. Pro- and macroglycogenolysis in contracting rat skeletal muscle. Acta Physiol. Scand. 169:291–296.[Medline]

Graham, T. E., K. B. Adamo, J. Shearer, I. Marchand, and B. Saltin. 2001. Pro- and macroglycogenolysis: relationship with exercise intensity and duration. J. Appl. Physiol. 90:873–879.[Abstract/Free Full Text]

Hansen, B. F., W. Derave, P. Jensen, and E. A. Richter. 2000. No limiting role for glycogenin in determining maximal attainable glycogen levels in rat skeletal muscle. Am. J. Physiol. 278:E398–E404.

Honikel, K. O. 1998. Reference methods for the assessment of physical characteristics of meat. Meat Sci. 49:447–457.

Jansson, E. 1981. Acid soluble and insoluble glycogen in human skeletal muscle. Acta Physiol. Scand. 113:337–340.[Medline]

Lomako, J., W. M. Lomako, and W. J. Whelan. 1991. Proglycogen: A low-molecular-weight form of muscle glycogen. FEBS Letter 279:223–228.[Medline]

Lomako, J., W. M. Lomako, W. J. Whelan, R. S. Dombro, J. T. Neary, and M. D. Norenberg. 1993. Glycogen synthesis in the astrocyte: From glycogenin to proglycogen to glycogen. FASEB J. 7:1386–1393.[Abstract]

Lowry, O. H., and J. V. Passonneau. 1972. A Flexible System of Enzymatic Analyses. Academic Press, New York.

Madsen, A., R. Østerballe, H. P. Mortensen, and C. Bejerholm. 1990a. Meat Quality. The Influence of feeds on meat quality of growing pigs. Natl. Inst. Anim. Sci., Rep. 673, Copenhagen, Denmark.

Madsen, A., J. Petersen, and A. Søgaard. 1990b. Anatomic content of the female and castrated male pig fed according to scale or ad libitum and slaughtered at 20, 50, 80 and 110 kg liveweight. Natl. Inst. Anim. Sci., Rep. 769, Copenhagen, Denmark.

Meyer, K. H., and R. Jeanloz. 1943. Recherce sur l’amidon XXV. Le glycogène du muscle natif. Helvet. Chim. Acta 26:1784–1798.

Rosenvold, K., H. N. Lærke, S. K. Jensen, A. H. Karlsson, K. Lundström, and H. J. Andersen. 2001a. Strategic finishing feeding as a tool in the control of pork quality. Meat Sci. 59:397–406.

Rosenvold, K., H. N. Lærke, S. K. Jensen, A. Karlsson, L. Lundström, and H. J. Andersen. 2002. Manipulation of critical quality indicators and attributes in pork through vitamin E supplementation, muscle glycogen reducing finishing feeding and pre-slaughter stress. Meat Sci. (In press.)

Rosenvold, K., J. S. Petersen, H. N. Lærke, S. K. Jensen, M. Therkildsen, A. H. Karlsson, H. S. Møller, and H. J. Andersen. 2001b. Muscle glycogen stores and meat quality as affected by strategic finishing feeding of slaughter pigs. J. Anim. Sci. 79:382–391.[Abstract/Free Full Text]

Wajzeer, J. 1939. Sur la formation et la décomposition du glycogène-protéide en relation avec les mutations du glycogène libre. Bull. Chim. Biol. 21:1242–1253.

Willistätter, R., and M. Rohdewald. 1934. Über den Zustand des Glytrogens in der Leber, im Muskel und in Leukocyten. Hoppe-Seyl Z. Physiol. Chem. 225:103–124.

Wismer-Pedersen, J., and E. J. Briskey. 1961. Rate of anaerobic glycolysis versus structure in pork muscle. Nature (Lond.) 189:318–320.[Medline]


This article has been cited by other articles:


Home page
 J ANIM SCIHome page
G. Bee, C. Biolley, G. Guex, W. Herzog, S. M. Lonergan, and E. Huff-Lonergan
Effects of available dietary carbohydrate and preslaughter treatment on glycolytic potential, protein degradation, and quality traits of pig muscles
J Anim Sci, January 1, 2006; 84(1): 191 - 203.
[Abstract] [Full Text] [PDF]

Home page
 J ANIM SCIHome page
E. Hambrecht, J. J. Eissen, R. I. J. Nooijen, B. J. Ducro, C. H. M. Smits, L. A. den Hartog, and M. W. A. Verstegen
Preslaughter stress and muscle energy largely determine pork quality at two commercial processing plants
J Anim Sci, May 1, 2004; 82(5): 1401 - 1409.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Rosenvold, K.
Right arrow Articles by Andersen, H. J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Rosenvold, K.
Right arrow Articles by Andersen, H. J.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS