Search

EP-4739146-A2 - A METHOD TO PRODUCE CRYSTALLINE LACTOSE PRODUCED BY A BIOTECHNOLOGICAL PRODUCTION PROCESS

EP4739146A2EP 4739146 A2EP4739146 A2EP 4739146A2EP-4739146-A2

Abstract

The present invention relates to methods to produce crystalline lactose, more specifically lactose obtained from a biotechnological production process.

Inventors

  • BEAUPREZ, JOERI
  • VERHAEGHE, TOM

Assignees

  • Inbiose N.V.

Dates

Publication Date
20260513
Application Date
20240705

Claims (20)

  1. 1. A method to produce a crystalline lactose, wherein said lactose is produced via a biotechnological production process wherein said method comprises a step of drying and/or crystallisation of said lactose.
  2. 2. The method according to claim 1, wherein said lactose is purified by purification steps selected from a microfiltration step, a centrifugation step, an ultrafiltration step, a nanofiltration step, an ion exchange step, a Simulated Moving Bed step, a colour removal step, a step of removal of sucrose, glucose or trehalose by addition of yeast, a step of removal of trehalose by addition of trehalase, a step of removal of sucrose by addition of an invertase; and preferably further comprises a concentration step; preferable said ion exchange step is a cation and/or anion exchange step, and/or a mixed bed ion exchange step.
  3. 3. The method according to any one of the preceding claims, wherein said biotechnological production process is a fermentation process, a bioconversion process or an enzymatic process.
  4. 4. The method according to claim 3, wherein said fermentation or bioconversion is followed by a biomass removal step, preferably said biomass removal step comprises a step of centrifugation, microfiltration and/or ultrafiltration.
  5. 5. The method according to claim 3, wherein said biotechnological production process is an enzymatic process and wherein at least one enzyme is immobilised.
  6. 6. The method according to any one of claim 1 to 3 or 5, wherein said biotechnological production process is an enzymatic process and wherein the reaction is performed between 50°C and 80°C, preferably between 55°C and 70°C.
  7. 7. The method according to any one of claim 1 to 4, wherein said biotechnological production process is a fermentation process or a bioconversion process characterised in that enzymatic conversions are performed by at least one enzyme produced by a cell synthesizing said enzyme.
  8. 8. The method according to claim 7, wherein said fermentation process or bioconversion process is performed at a temperature between about 25°C and about 50°C, preferably between about 25°C and about 40°C.
  9. 9. The method according to any one of claim 1 to 4, 7 or 8, wherein said biotechnological production process is a fermentation process characterised in that said lactose is produced by a cell genetically engineered to produce all enzymes necessary for production of lactose.
  10. 10. The method according to any one of the previous claims, wherein said lactose is purified from the biotechnological production process resulting liquid comprising sucrose, glucose, trehalose, fructose, and/or glycerol.
  11. 11. The method according to any one claim 2 to 10, wherein the lactose purity before purification is < 70%, < 60%, < 50%, < 40%, < 30%, < 20%, < 10% on total solids, and/or the purity at the end of said method comprising purification is higher than > 80% on dry solids, preferably > 85% on dry solids, more preferably > 90% on dry solids, even more preferably > 95% on dry solids, even more preferably > 97% on dry solids, even more preferably > 98% on dry solids, most preferably > 99% on dry solids.
  12. 12. The method according to any one of claim 2, 3, 5, 6, 10 or 11, wherein said purification of an enzymatically produced lactose comprises a step of a microfiltration or ultrafiltration to recover the used enzymes.
  13. 13. The method according to any one of claim 2 to 4, 7 to 11 wherein cells removed from said fermentation or bioconversion are reused in a further fermentation or bioconversion process.
  14. 14. A method according to any of one of claim 1 to 13, wherein said biotechnological production process comprises i) a step of nanofiltration to remove/reduce monosaccharides and/or salts; and/or ii) a step of ion exchange to remove charged materials.
  15. 15. The method according to any one of claim 1 to 14, wherein said lactose has an ash content below 1% on total solid, preferably below 0,5% on total solid, preferably with one or more of: a) Lead content lower than 0,1 mg/kg solid, preferably lower than 0,02 mg/kg solid; b) Arsenic content lower than 0,2 mg/kg solid, preferably lower than 0,02 mg/kg solid; c) Cadmium content lower than 0,1 mg/kg solid, preferably lower than 0,01 mg/kg solid; or d) Mercury content was lower than 0,5 mg/kg solid, preferably lower than 0,1 mg/kg solid.
  16. 16. The method according to any one of claim 1 to 15, wherein said lactose has at least one or more of: i) a protein content below 100 mg per kg dry solid, ii) DNA content below 10 ng per gram dry solid and iii) endotoxin content below 10000 EU per gram dry solid.
  17. 17. The method according to any one of claim 1 to 16, wherein said lactose crystals are dried by band drying, belt drying, vacuum band drying, vacuum belt drying, drum drying, vacuum drum drying, roller drying, vacuum roller drying and other types of drying.
  18. 18. The method according to any one of claim 1 to 16, said step of drying comprises any one or more of spray drying, lyophilization, evaporation, precipitation, spray freeze drying, freeze spray drying, band drying, belt drying, vacuum band drying, vacuum belt drying, drum drying, roller drying, vacuum drum drying, vacuum roller drying, and agitated thin film drying.
  19. 19. A crystalline lactose product wherein said lactose product is derived from a biotechnological production process, preferably produced by a method as described in any of claims 1 to 18.
  20. 20. The lactose product of claims 19, wherein said lactose is lactose monohydrate.

Description

A method to produce crystalline lactose produced by a biotechnological production process The present invention relates to methods to produce crystalline lactose, more specifically lactose obtained from a biotechnological production process. Lactose as a disaccharide knows a wide variety of applications, in the food and pharmaceutical industry ( Hebbink GA, Dickhoff BHJ. Chapter 5 - Application of lactose in the pharmaceutical industry. In: Paques M, Lindner CBT-L, editors. Academic Press; 2019. p. 175-229. Available from: https://www.sciencedirect.com/science/article/pii/B9780128117200000052, Hettinga KA. Chapter 6 - Lactose in the dairy production chain. In: Paques M, Lindner CBT-L, editors. Academic Press; 2019. p. 231- 66. Available from: https://www.sciencedirect.com/science/article/pii/B9780128117200000064). Today it is isolated from mammalian milk, mainly cow's milk. Due to the environmental impact of this production process, there is a need for alternative synthesis methods that can compete with the well-established extraction methods from milk at low production cost. Mammals synthesize lactose by means of a lactose synthase, or beta-1,4 galactosyltransferase, transferring a galactose to glucose with UDP-galactose as a donor substrate and glucose as an acceptor substrate. This principle has been engineered in microbes, copying the mammalian biosynthesis pathway ( Mao Z, Shin H-D, Chen RR. Engineering the E. coli UDP-glucose synthesis pathway for oligosaccharide synthesis. Biotechnol Prog [Internet], Jan [cited 2016 Mar 13];22(2):369-74. Available from: http://www.ncbi.nlm.nih.gov/pubmed/16599548) and the synthesis enzymes have been applied in vitro before to synthesize lactose by the addition of the expensive UDP-galactose (Lau K, Thon V, Yu H, Ding L, Chen Y, Muthana MM, et al. Highly efficient chemoenzymatic synthesis of [small beta]l-4-linked galactosides with promiscuous bacterial [small beta]l-4-galactosyltransferases. Chem Commun [Internet], 2010;46(33):6066-8. Available from: http://dx.doi.org/10.1039/C0CC01381A) as donor. A challenge one encounters with these methodologies is, is the energy requirement for lactose synthesis. The synthesis of UDP-galactose over UDP-glucose requires UTP, which either has to be added to the reaction mixture or be synthesized by the microbial host. After lactose synthesis UDP is released which is either lost as a substrate or may be recycled into UTP by means of ATP, another costly biochemical intermediate. Furthermore, the addition or synthesis of galactose is required, which is currently still derived from lactose and no alternative synthesis methods have been identified. A challenge one encounters is to purify lactose from a biotechnological synthesis environment, which encompasses products (such as e.g. medium component, buffer component, substrates, precursors, cells, proteins, DNA, lipids, cell debris, enzymes, chemicals, antifoam agents,...) that are not naturally present in lactose isolated from mammalian milk. These are amongst others sugars, such as glucose, fructose, UDP-glucose, UDP-galactose, galactose, sucrose; salts, such as phosphate buffers, sulphates, ammonia, magnesium, calcium...; proteins, cells or cellular components , which typically interfere with the purification of the biotechnologically produced lactose. These impurities can inhibit nucleation and crystal growth, and impurities themselves can potentially be incorporated into crystals, thus affecting the purity of the final product. It is therefore far from obvious to purify lactose from a biotechnological process at purities that are acceptable for the market. Purely from an energy point and cost point of view, the current state of the art would not be able to compete with extractive production of lactose, even if it is more environmentally friendly. Summary of the invention It is an object of the present invention to provide for methods to produce a crystalline lactose, biotechnologically produced, which solves the problem of the isolation of such lactose at high purity and at a reasonable production cost. The present invention provides a new method for efficiently producing lactose wherein said lactose is preferably first separated from its biotechnological production environment and then crystallized. According to this invention this and other objects are achieved by providing a method to produce a crystalline lactose, wherein said lactose is produced via a biotechnological production process wherein said method comprises a step of drying and/or crystallization of said lactose. Further benefits of the teachings of this invention will be apparent to one skilled in the art from reading this invention. Definitions The words used in this specification to describe the invention and its various embodiments are to be understood not only in the sense of their commonly defined meanings, but to include by special definition in this specification structure, material or acts beyond the scope