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Effect of disaccharides on survival during storage of freeze dried probiotics

Effet des disaccharides sur la survie de probiotiques lyophilisés au cours du stockage

  • Original Article
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Dairy Science & Technology

Abstract

The aim of this study was to investigate the effects of protective media and different relative vapour pressures (RVPs) on the survival of probiotics during freeze drying and subsequent storage, to determine the optimal conditions for the production of freeze dried probiotics at industrial scale, ensuring a high survival rate. The effect of protective media i.e. reconstituted skimmed milk (RSM) or either of the cryoprotective disaccharides lactose, trehalose, sucrose, maltose, lactose + maltose and lactose + trehalose on the survival of a probiotic culture was assessed at five different RVP environments (0.0%, 11.4%, 33.2%, 44.1% and 76.1%) at room temperature in freeze-dried systems. RVP was shown to have a significant effect on the survival rates of the probiotic cultures Lactobacillus paracasei NFBC 338 and Lactobacillus rhamnosus GG following freeze drying in RSM. Interestingly, retention of cell viability was greatest for cells stored at 11.4% RVP, but was compromised at all other RVPs tested. However, an increased tolerance to freeze drying was observed for L. rhamnosus GG when dried in the presence of disaccharides in the order of trehalose = lactose + maltose ⩾ lactose + trehalose ⩾ maltose > lactose > sucrose. Survival studies over a 38–40 day storage period indicated that trehalose and lactose + maltose were the most effective cryoprotective additives, especially notable at 0.0 and 11.4% RVP. At all other RVPs tested, viability was compromised. Crystallisation of the disaccharides was observed to be a detrimental factor affecting the survival of Lactobacillus during storage at high RVP, where an inverse relationship was shown to exist between the % RVP and the glass transition temperature (Tg) of the disaccharides.

Abstract

(RSM) (0.0%, 11.4%, 33.2%, 44.1% ?76.1%) (Lactobacillus paracasei) NFBC338 (Lactobacillus rhamnosus) LGG 11.4% LGG 38–40 0.0% 11.4% (Lactobacillus) Tg

Résumé

Le but de cette étude était de rechercher les effets de milieux protecteurs et de différentes pressions de vapeur relatives (PVR) sur la survie de probiotiques au cours de la lyophilisation et du stockage ultérieur, afin de déterminer les conditions optimales pour la production de probiotiques lyophilisés à l’échelle industrielle assurant un taux de survie élevé. L’effet de milieux de protection, i.e. lait écrémé reconstitué ou l’un des disaccharides cryoprotecteurs suivants : lactose, tréhalose, saccharose, maltose, lactose + maltose ou lactose + tréhalose, sur la survie d’une culture de probiotiques a été étudié dans 5 environnements de PVR différents (0; 11,4; 33,2; 44,1 et 76,1 %) à température ambiante dans des systèmes lyophilisés. On a montré que la PVR avait un effet significatif sur les taux de survie des cultures de probiotiques Lactobacillus paracasei NFBC 338 et Lactobacillus rhamnosus GG après lyophilisation dans le lait écrémé reconstitué : le maintien de la viabilité cellulaire était plus élevé pour des cellules conservées à une PVR de 11,4 %, alors qu’il était compromis à toutes les autres PVR étudiées. Cependant, une tolérance accrue à la lyophilisation était observée pour L. rhamnosus GG en présence de disaccharides, dans l’ordre tréhalose = lactose + maltose ⩾ lactose + tréhalose ⩾ maltose > lactose > saccharose. Les études de survie sur 38–40 jours ont mis en évidence que le tréhalose et le lactose + maltose étaient les additifs cryoprotecteurs les plus efficaces, surtout à 0 et 11,4 % de PVR. Á toutes les autres PVR testées, la viabilité était compromise. On a observé que la cristallisation des disaccharides était préjudiciable à la survie de Lactobacillus au cours du stockage aux PVR élevées, une relation inverse ayant été montrée entre le % de PVR et la température de transition vitreuse (Tg) des disaccharides.

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References

  1. Ahola A.J., Yli-Knuuttila H., Suomalainen T., Poussa T., Ahlstrom A., Meurman J.H., Korpela R., Short-term consumption of probiotic-containing cheese and its effects on dental caries risk factors, Arch. Oral Biol. 47 (2002) 799–804.

    Article  CAS  Google Scholar 

  2. Ananta E., Knorr D., Evidence on the role of protein biosynthesis in the induction of heat tolerance of Lactobacillus rhamnosus GG by pressure pre-treatment, Int. J. Food Microbiol. 96 (2004) 307–313.

    Article  CAS  Google Scholar 

  3. Carpenter J.F., Crowe J.H., Modes of stabilization of a protein by organic solutes during dessication, Cryobiology 25 (1989) 459–470.

    Article  Google Scholar 

  4. Carvalho A.S., Silva J., Ho P., Teixeira P., Malcata F.X., Gibbs P., Effects of various sugars added to growth and drying media upon thermotolerance and survival throughout storage of freeze-dried Lactobacillus delbreuckii ssp. bulgaricus, Biotechnol. Prog. 20 (2004) 248–254.

    Article  CAS  Google Scholar 

  5. Champion D., Le Meste M., Simatos D., Towards an improved understanding of glass transition and relaxations in foods: molecular mobility in the glass transition range. Trends Food Sci. Technol. 11 (2000) 41–55.

    Article  CAS  Google Scholar 

  6. Chavarri F.J., De Paz M., Nuôez M., Cryoprotective agents for frozen concentrated starters from non-bitter Streptococcus lactis strains, Biotechnol. Lett. 10 (1988) 11–16.

    Article  CAS  Google Scholar 

  7. Codex standard for fermented milks, Codex Stan 243-2003.

  8. Conrad P.B., Miller D.P., Cielenski P.R., de Pablo J.J., Stabilisation and preservation of Lactobacillus acidophilus in saccharide matrices, Cryobiology 41 (2000) 17–24.

    Article  CAS  Google Scholar 

  9. Corcoran B.M., Ross R.P., Fitzgerald G.F., Dockery P., Stanton C., Enhanced survival of GroESL-overproducing Lactobacillus paracasei NFBC338 under stressful conditions induced by drying, Appl. Environ. Microbiol. 72 (2006) 5104–5107.

    Article  CAS  Google Scholar 

  10. Corcoran B.M., Ross R.P., Fitzgerald G.F., Stanton C., Comparative survival of probiotic lactobacilli spray-dried in the presence of probiotic substances, J. Appl. Microbiol. 96 (2004) 1024–1039.

    Article  CAS  Google Scholar 

  11. Crowe J.H., Carpenter J.F., Crowe L.M., The role of vitrification in anhydrobiosis, Ann. Rev. Physiol. 60 (1998) 73–103.

    Article  CAS  Google Scholar 

  12. Crowe J.H., Crowe L.M., Carpenter J.F., Rudolph A.S., Aurell Winstrom C., Spargo B.J., Anchordoguy T., Interactions of sugars with membranes, Biochim. Biophys. Acta 947 (1988) 367–384.

    CAS  Google Scholar 

  13. Crowe J.H., Crowe L.M., Jackson S.A., Preservation of structural and functional activity in lyophilised sarcoplasmin reticulum, Arch. Biochem. Biophys. 220 (1983) 477–484.

    Article  CAS  Google Scholar 

  14. Crowe J.H., Hoekstra F.A., Crowe L.M., Membrane phase transitions are responsible for imbibitional damage in dry pollen, Proc. Natl. Acad. Sci. USA 86 (1989) 520–523.

    Article  CAS  Google Scholar 

  15. de Antoni G.L., Pérez P., Abraham A., Anón M.C., Trehalose, a cryoprotectant for Lactobacillus bulgaricus, Cryobiology 26 (1989) 149–153.

    Article  Google Scholar 

  16. Desmond C., Fitzgerald G.F., Stanton C., Ross R.P., Improved stress tolerance of GroESL-overproducing Lactococcus lactis and probiotic Lactobacillus paracasei NFBC 338, Appl. Environ. Microbiol. 70 (2004) 5929–5936.

    Article  CAS  Google Scholar 

  17. Desmond C., Stanton C., Fitzgerald G.F., Collins K., Ross R.P., Environmental adaptation of probiotic lactobacilli towards improvement of performance during spray drying, Int. Dairy J. 11 (2001) 801–808.

    Article  Google Scholar 

  18. Erkkila S., Suihko M.L., Eerola S., Petaja E., Mattila-Sandholm T., Dry sausage fermented by Lactobacillus rhamnosus strains, Int. J. Food Microbiol. 64 (2001) 205–210.

    Article  CAS  Google Scholar 

  19. Gardiner G.E., O’Sullivan E., Kelly J., Auty M.A., Fitzgerald G.F., Collins J.K., Ross R.P., Stanton C., Comparative survival rates of human-derived probiotic Lactobacillus paracasei and L. salivarius strains during heat treatment and spray drying, Appl. Environ. Microbiol. 66 (2000) 2605–2612.

    Article  CAS  Google Scholar 

  20. Gatlin L.A., Nail S.L., Protein purification and process engineering. Freeze drying: a practical overview, Bioprocess Technol. 18 (1994) 317–367.

    CAS  Google Scholar 

  21. Green J.L., Angell C.A., Phase relations and vitrification in saccharide-water solutions and the trehalose anomaly, J. Phys. Chem. 93 (1989) 2880–2882.

    Article  CAS  Google Scholar 

  22. Greenspan L., Humidity fixed points of binary saturated aqueous solutions, J. Res. Nat. Bur. Stand. 81A (1977) 89–96.

    Google Scholar 

  23. Hatley R.H., The effective use of differential scanning calorimetry in the optimization of freeze drying processes and formulations, Dev. Biol. Stand. 74 (1992) 105–119.

    CAS  Google Scholar 

  24. Hoekstra F.A., Crowe L.M., Crowe J.H., Effect of sucrose on phase behaviour of membranes in intact pollen of Typha latifolia L., as measured with fourier transform infrared spectroscopy, Plant Physiol. 97 (1991) 1073–1079.

    Article  CAS  Google Scholar 

  25. Hubalek Z., Protectants used in the cryopreservation of microorgansisms, Cryobiology 46 (2003) 205–229.

    Article  CAS  Google Scholar 

  26. Jennings T.A., Lyophilisation-introduction and basic principles, CRC Press, Boca Raton, 1999.

    Google Scholar 

  27. Koster K.L., Leopold A.C., Sugars and dessication tolerance in seeds, Plant Physiol. 88 (1988) 829–832.

    Article  CAS  Google Scholar 

  28. Labuza T.P., Kaanane A., Chen J.Y., Effect of temperature on the moisture isotherms and water activity shift of two dehydrated foods, J. Food Sci. 50 (1985) 385–391.

    Article  CAS  Google Scholar 

  29. Leslie S.B., Israeli E., Lighthart B., Crowe J.H., Crowe L.M., Trehalose and sucrose protect both membranes and proteins in intact bacteria during drying, Appl. Environ. Microbiol. 61 (1995) 3592–3597.

    CAS  Google Scholar 

  30. Lievonen S.M., Laaksonen T.J., Roos Y.H., Glass transition and reaction rates: nonenzymatic browning in glassy and liquid systems, J. Agric. Food. Chem. 46 (1998) 2778–2784.

    Article  CAS  Google Scholar 

  31. Manley K.J., Fraenkel M.B., Mayall B.C., Power D.A., Probiotic treatment of vancomycin-resistant enterococci: a randomised controlled trial, Med. J. Aust. 186 (2007) 454–457.

    Google Scholar 

  32. Marteau P.R., de Vrese M., Cellier C.J., Schrezenmeir J., Protection from gastrointestinal diseases with the use of probiotics, Am. J. Clin. Nutr. 73 (2001) 430S-436S.

    CAS  Google Scholar 

  33. Oetjen G.W., Freeze-drying, Wiley-VCH, Weinheim, 1999.

    Book  Google Scholar 

  34. Paiva C.L., Panek A.D., Biotechnological applications of the disaccharide trehalose, Biotechnol. Annu. Rev. 2 (1996) 293–314.

    Article  CAS  Google Scholar 

  35. Patist A., Zoerb H., Preservation mechanisms of trehalose in food and biosystems, Colloids Surf. B 40 (2005) 107–113.

    Article  CAS  Google Scholar 

  36. Roos Y.H., Phase transition in foods, Academic Press Inc., San Diego, USA, 1995.

    Google Scholar 

  37. Schoug Å., Olsson J., Carlfors J., Schnürer J., Håkansson J., Freeze-drying of Lactobacillus coryniformis Si3-effects of sucrose concentration, cell density, and freezing rate on cell survival and thermophysical properties, Cryobiology 53 (2006) 119–127.

    Article  CAS  Google Scholar 

  38. Sudoma A.L., Stabilisation of dried bacteria extended in particulate carriers, U. S. Patent No. 4 956 295, 1990.

  39. Sun W.Q., Davidson P., Protein inactivation in amorphous sucrose and trehalose matrices: effects of phase separation and crystallization, Biochim. Biophys. Acta 1425 (1998) 235–244.

    CAS  Google Scholar 

  40. Tunnacliffe A., Garcia de Castro A., Manzanera M., Anhydrobiotic engineering of bacterial and mammalian cells; is intracellular trehalose sufficient? Cryobiology 43 (2001) 124–132.

    Article  CAS  Google Scholar 

Download references

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Correspondence to Catherine Stanton.

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Miao, S., Mills, S., Stanton, C. et al. Effect of disaccharides on survival during storage of freeze dried probiotics. Dairy Sci. Technol. 88, 19–30 (2008). https://doi.org/10.1051/dst:2007003

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