Lactic Acid in Muscle and its Effects on Meat Quality

Transcription

1 Lactic Acid in Muscle and its Effects on Meat Quality Eero Puolanne Department of Food Technology/ Meat Technology University of Helsinki Helsinki, Finland

2 Thank you! It is an honor and extraordinary pleasure to me to be the recipient of AMSA international Award, and also to have this presentation for you. I do appreciate this very much indeed! I also thank the organizers for an excellently organized meeting, fine meeting venue, good food, and finally, for the cozy casual atmosphere! I and my wife as well, we are so happy to visit this great country again and meet old friends and make new.

3 Warning! English is not my native language This PowerPoint presentation and my talk may cause pain and distress for native English-speaking individuals In case of headache, ask earplugs and painkillers from Rachel Hamilton, and close your eyes!

4 The research group Department of Food Technology Department of Basic Veterinary Sciences /biochemistry Prof. Eero Puolanne PhD Marita Ruusunen MSc Maria Kylä-Puhju Prof. Reeta Pösö MSc Katri Sepponen

5 Motto Life is to detain the entropy of solar energy All species try to proliferate and fill their ecological niche The strength of the molecules of an organism are tested billions of times per second The functions of molecules and the whole biological system is also tested constantly In nature, the organisms are optimally balanced The safety margins decrease the ability to compete Farm animals, do they have a harmonic growth and balanced physiology?

6 * Red fiber * White fiber * Intermediate fiber Porcine M. masseter

7 Femur crossection areas of 165 d old pigs Smallest, 447 mm 2 Largest, 715 mm 2 Unpublished results! Thinnest, 3.9 mm Thickest, 5.7 mm

8 ph is the Key Factor - Animal Welfare/Meat Quality A vast flow of information constantly flowing in the the bodies of fauna Consciousness is the capability of neural network to pick up relevant information, slow it down and put light on it If something is causing a danger to the life of the individual, pain and/or distress will occur. Our duty is to provide to the animals circumstances, where there is no unnecessary pain/distress In a living muscle the ph is high stress/low ph causes pain and distress

9 ph is the Key Factor - Animal Welfare/Meat Quality In a postmortal muscle the ph is Perimortal effects (time, temperature, ph) on tenderness development, color, and drip and waterbinding the effects of ph u on rigor/post rigor meat tenderness, color, water-binding (salt-phosphate effects)! ph differences of about units cause large differences originating! from mmol/kg lactic acid or! 5-10 mmol/kg glucose, only

10 WBC NaCl w/o NaCl ph

11 The Glycogen Glycogen, I.8% or 100 mmol/kg ( mmol/kg) expressed as glucose equivalents Serves as energy reservoir that is independent of blood supply Would provide energy for 20 s to 15 min??? depending very many factors, e.g. oxygen status, type of exercise and/or stress

12 The Glycogen Molecule Glycogenin, MW 37,300 Branched chains of 13 glucose units 1,4-bonds, or 1,6-bond at fourth and eighth units minimizes volume, maximizes non-reducing ends and available glucose

13 1,6-branch Glycogenin (PhD Kaisa Immonen, 2000)

14 Prolycogen and macroglycogen! Proglycogen, 6-7 tiers, MW 300,000 to 575,000, non-reducing ends, ca 10% protein! Macroglycogen, tiers, MW 5-10 mo 1000 to 2000 non-reducing ends, ca. 0.4% protein! phosphorylase tetramers (or dimers)/molecule

15 Lactic acid Glycolytic potential (PhD Kaisa Immonen, 2000)

16 (PhD Kaisa Immonen, 2000)

17 I II III (PhD Kaisa Immonen, 2000)

18 P.M. glycogenolysis Where is ATP needed? Calcium from sarcoplasmic reticulum --> shortening reactions --> ATP is consumed The key enzyme of glycolysis (phosphofructokinase) is inhibited by low ph Other reasons? Pre mortem post mortem? PSE? ph is a pivot point onset of rigor mortis, if not earlier the rigor is eventually different (stronger) when setting on at low ph?

19 Phosphorylase 2% of the total protein in muscles dimers or tetramers/glycogen molecule Each one is bound at two points to a (unbranched) A-chain One is active and the other is a regulatory binding site Cleaves glucose-1-p from the non-reducing end VERY quickly, until the fourth unit from the branching point! When the uppermost tier (whatever it is) is full, theoretically, 34.6% glucose of the glycogen molecule will be available

20 Phosphorylase (PhD Kaisa Immonen, 2000)

21 Phosphorylase! When glycogen content is 100 mmol/kg, ca 35 mmol/kg glucose-1-p -->70 mmol/kg lactic acid, which means more than 1 unit drop in ph in living muscle this enough for fast energy needs! Macroglycogen is used more in aerobic stress, proglycogen more in anaerobic stress and post mortem The mechanisms may involve the allosteric activation of phosphorylase b kinase by Ca ++ (breakdown of PG) and phosphorylase b by AMP (MG)

22 The Glycogen Debranching Enzyme Bifunctional: transferase and glucosidase! Transferase moves three glucose units to a B chain! 1,6-glucosidase cleaves 1,6-glucosyl unit as free glucose (8% of the residues) activity about 10% of that of phosphorylase! may limit the breakdown rate? Effects of ph and temperature? Living muscle/ post mortem muscle? Residual glycogen mmol/kg?

23 Phosphorylase Transferase 1,6-glucosidase (PhD Kaisa Immonen, 2000)

24 The Formation of Lactic Acid Glucose --> 2 lactic acid ATP The anaerobic production of energy Stimulated by mental (adrenalin) or physical (Ca ++, AMP) stress or both 0 (?) to 80% of energy production aerobically 2 to 40 mmol ATP/kg*min anaerobically 0 to 250 mmol ATP/kg*min! E.g. in pigs produce constantly?

25 Lactate Dehydrogenases Tetramers of muscle type (M) and/or heart-type (H) H 4 (LDH-1) H 3 M (LDH-2) M 4 (LDH-5)! At a high rate of glycolysis LDH-5 favors the reduction of pyruvate to lactic acid, especially in light muscles In heart and! in dark muscles LDH-1 converts lactic acid to pyruvate LDH-activity, M/H percentages

26 Monocarboxylate Transporters!!! Prof. Reeta Pösö s group pk a of lactic acid is 3.86, i.e. at physiological ph <0.1% undissociated acid! Only undissociated form is able to cross the membranes The key enzyme of glycolysis (phosphofructokinase) is inhibited by high concentrations of protons

27 *

28 Monocarboxylate Transporters Monocarboxylate transporters (MCT) transport lactate to the direction of lactate gradient Nine types found as far, and the most studied are MCT1 (in aerobic muscles) MCT2 ( low K m and V max ) MCT4 (high K m and V max, in anaerobic muscles)! Prof. Reeta Pösö s group has been able to show a high content of MCT2 in porcine muscles.! May be a house-keeping protein for continuous production of lactate?

29 High capillarization qnd oxygen supply Low capillarization and oxygen supply *

30 Lactate is utilized aerobically!!!!!! Lactic acid is converted to pyruvate in muscles, heart and non-contracting tissues (especially in liver and in red blood cells) LA contains over 90%of the energy of the glucose LA is consumed (--> CO 2 ) or stored as glucose (glycogen)! This takes place aerobically, only! The formation of glycogen in liver uses ATP that has been produced aerobically

31 (AEROBIC!) Glycogen *

32 The Buffering Capacity β = da/dph (A = the amount of acid/base) The buffering capacity is usually ca 50 (40-60) mequivalents/kg*ph In light muscles higher than in dark muscles Myofibrillar proteins ca me/kg*ph (ph >) Soluble components 10 (ph 5.5) to 30 (ph 6.8) me/kg*ph Lactic acid relevant only in low phs (below 5)! In muscles, mmol LA/kg needed to lower the ph from 7.2 to 5.5 (3 µmol/kg in water!)

33 (PhD Riitta Kivikari, 1996)

34 (PhD Riitta Kivikari, 1996)

35 The Rate of ATP Consumption and Production ATP content in the living muscle is ca 8-10 mmol/kg The consumption varies from 2-3 (at rest) to (maximal) mmol/kg*min aerobically 2 to 40 mmol ATP/kg*min anaerobically 0 to 250 mmol ATP/kg*min! Theoretically, the drop in ph from 7.2 to 6.2 would require ca 60 mmol/kg LA, i.e. to produce 60 to 90 mmol/kg ATP, which is consumed in 15 to 30 seconds

36 Sources of ATP Glycolysis mmol/kg Pork myoglobin mmol/kg Beef mmol/kg 1 mmol O > 6 mmol ATP ---> mmol/kg ATP Hemoglobin? Creatine phosphate mmol/kg I.e. up to 1/3 of that of glycolysis Delay the onset of glycolysis Slow down the rate of glycolysis Difference pigs and broilers / cattle and sheep Cooling rate: drip and cold shortening

37 What have we done? The liver cannot possibly receive and convert 150 to 900 g lactic acid in a short period of time Neither does the blood Therefore, lactic acid is used in muscles aerobically, preferably in red and intermediate fibers, sooner or later Beef animals and sheep may not have problems! In (many) porcine and poultry muscles there now are very little (10 0%) red and intermediate fibers and very poor capillarization to get oxygen! Cardio-respiratory capacity is also low! What else we should expect than PSE-like meat?! What about animal welfare?

38 Research needs " More quantitative physiological data on carbohydrate metabolism needed " The fate of lactic acid: where does it go in stress, how it is utilized/stored? " MCTs? " Proglycogen, macroglycogen vs. phosphorylase? " Buffering capacity? " Muscle status in stress and at death? " Muscle and species differences! Dead or alive: lactic acid is there!!