2024 Nouvelle approche de fractionnement des composés protéiques du babeurre à l'aide d'hydroxyapatite à des fins de valorisation

Authors:
Jean Lung

Journal:
Dissertation

Institute:

Université Laval

Abstract:

Buttermilk is the co-product obtainedfrom churning cream into butter. As butter consumption continues to rise in Quebec and Canada, significant quantities of buttermilk, produced in the same mass ratios as butter, need to be valorized. Although buttermilk shares a similar composition to skimmilk, its technological use is limited due to certain characteristics. Apart from its emulsifying power, its techno-functional properties are often inferior to skim milk. These differences are attributed, among other factors, to the release of milk fat globule membrane (MFGM)  fragments  into  buttermilk  during  cream  churning.  However,  MFGM  contains  several  high-value compounds,such as phospholipids and membrane proteins, possessing nutritional and health properties. Thus, separating MFGM from buttermilk could yield two fractions: a delipidated fraction usable as skim milk and a lipid-rich fraction containing MFGM phospholipids usable as a bioactive compound. However, separating buttermilk components  remains  a  significant  challenge,  especially  on  an  industrial  scale.  This  study  focuses  on implementing   a   technique   for   separating   buttermilk  components   using  their   affinity   for   an   adsorbent, hydroxyapatite (HA).

As part of this thesis, an initial study was conducted to determine if it was possible to adsorb MFGM fragments from pasteurized and non-pasteurized cream buttermilk onto HA. This study confirmed that both types of MFGM could interact with HA, with MFGM from pasteurized cream buttermilk showing greater affinity for HA than MFGM from raw cream. Moreover, MFGM proteins tended to adsorb onto the surface of HA, while phospholipids (PL) adsorbed  internally.  However,  they  adsorbed  in  equal  proportions  and  at  the same  rate,  indicating  complete adsorption  of  MFGM  fragments  initially.  Subsequently,  aggregation  of  HA  was  observed  during  MFGM adsorption, suggesting that MFGM fragments adsorbed in multilayers on HA surfaces. This aggregation created interparticle  spaceswhere  additional  MFGM  fragments  could  intercalate.  This  first  part  verified  that  MFGM, composed of proteins and PL, could interact with and adsorb onto HA. Furthermore, no proportion or adsorption rate differenceswere observed between PL and MFGM proteins, only in their adsorption location within HA.

In a second study, the affinitiesof major buttermilk proteins,such as caseins(CN), α-lac(α-lac), and β-lg(β-lg),and  MFGMfragmentsfor  HA were individually  determined  under  physicochemical  parameterschanges(pH, ionic strength, and temperature). It was found that caseins exhibited the highest affinity for HA, followed by α-lac and β-lg,  which  had  similar  affinities  for  HA,  and  then  MFGM.  Subsequently,  a  study  on  the  influence  of physicochemical parameters on the adsorption of each buttermilk component onto HA was conducted. Changes in  pH,  ionic  strength,  and  temperature  only  affected  casein (CN) adsorption.  Specifically,  MFGM  and  β-lgadsorbed completely regardless of the parameters, while α-lac  adsorbed  at  90%,irrespectiveof  the  studied conditions.

The  third  study  aimed  to  validate  these  observations  for  each  component  but  in  a  model  mixture.  This  time, changes  in  physicochemical parameters  influenced  the  adsorption  of  each  component  in the  mix,  resulting  in the predominant adsorption of CN(90%) and minor adsorption of whey proteins (11% α-lac and 37% β-lg) and MFGM  (7%).  It  was  then  proposed to  fix theCNvia  HA,  allowing  subsequent  separation  of  MFGM  from  the remaining  whey  proteins  based  on  their  solubility  differences  at  varying  pH  levels.  By  adjusting  the physicochemical  parameters  determined  in  the  model  mixture,  it  was  possible  to recover  90%  of the CNin  a diluted  buttermilk  solution through  HA  adsorption.  Subsequently,  MFGM  was  separated from the unadsorbed whey  proteins  by  selective  precipitation.  Adjusting  the  pH  to  approximately  4.0  led  to  complete  MFGM precipitation compared to whey proteins, which remained mostly soluble. Only 30% of β-lgwas co-precipitated, attributed to its interaction with MFGM following cream pasteurization. Thus, the majority of β-lg(70%) and α-lac (100%), being more thermally stable, were entirely recovered in the soluble phase. Ultimately, this sequence of processes (HA and selective precipitation) yielded three fractions from diluted buttermilk: one enriched in CN, another in MFGM, and a third in whey proteins and the remaining buttermilk components (lactose and minerals).

This thesis project has contributed new insights into the HA-based separation of buttermilk components and the influence  of  physicochemical  parameter  changes  on  their  interactions.  The  knowledge  gained  regarding  the behavior  of  buttermilk  proteins  and  MFGM  with  HA  particles,  combined  with  selective  precipitation of  MFGM, has  led to  the  development  of  a  novel  and  straightforward  technique for  fractionating buttermilk  components, generating  fractions  enriched  in CN,  whey  proteins,  and  notably  MFGM.  This  has  addressed  two  identified challenges in the literature: isolating MFGM from other buttermilk components to valorize these distinct fractions (MFGM  and  delipidated  buttermilk),  and  separating  MFGM  from  whey  proteins.  These  new  insights  suggest potential  applications  for  fixing  proteins  and  PL  via  HA  for  other  dairy  fluids,  as  well  as  using  selective precipitation by pH adjustment to separateMFGM from whey proteins.