Effect of raw materials and production processes on dry powder media quality
Upcoming SlideShare
Loading in...5
×
 

Effect of raw materials and production processes on dry powder media quality

on

  • 702 views

Effect of raw materials and production processes on dry powder media quality

Effect of raw materials and production processes on dry powder media quality

Statistics

Views

Total Views
702
Views on SlideShare
702
Embed Views
0

Actions

Likes
0
Downloads
4
Comments
0

0 Embeds 0

No embeds

Accessibility

Upload Details

Uploaded via as Adobe PDF

Usage Rights

© All Rights Reserved

Report content

Flagged as inappropriate Flag as inappropriate
Flag as inappropriate

Select your reason for flagging this presentation as inappropriate.

Cancel
  • Full Name Full Name Comment goes here.
    Are you sure you want to
    Your message goes here
    Processing…
Post Comment
Edit your comment

Effect of raw materials and production processes on dry powder media quality Effect of raw materials and production processes on dry powder media quality Presentation Transcript

  • Effect of raw materials and production processes on dry powder media quality DMEM-F12 production: a case study Dr. ZIMMER Aline Cell Culture World Congress Munich, 29/02/2012
  • Agenda 1 DMEM-F12 media characterization • Physico-chemical characterization • Cellular performance testing (CHO-S cells) • Fingerprinting • Performance prediction 2 3 2 Quality improvement - case studies Perspectives in future media preparation Cell Culture World congress, 29/02/2012
  • Impact of raw materials on dry powder media quality Physicochemical properties Processing Packaging concept Purity Raw material quality 3 Cell Culture World congress, 29/02/2012
  • Impact of production processes on dry powder media quality Blending process Milling process Quality of raw materials 4 Cell Culture World congress, 29/02/2012 Dry powder media quality Packaging concept
  • Benchmarking of DMEM-F12 • Recipe containing 50 raw materials (inorganic salts, amino acids, vitamins, glucose, lipids...) INORGANIC SALTS VITAMINS OTHERS Chloride salts Calcium / Magnesium Potassium / Sodium Unpolar / hydrophobic Ala / Met / Leu / Ile / Pro / Phe / Trp Biotin Calcium pantothenate Dextrose Putrescine Sodium pyruvate Phosphate salts Sodium dibasic / monobasic Neutral Tyr / Thr / Gln / Gly / Ser / Cys / Cystine / Asn Choline chloride Cyanocobalamin Folic acid Linoleic / lipoic acid Sulfate salts Cupric / Ferrous Magnesium / Zinc Basic Lys / Arg / His i-inositol Niacinamide Pyridoxal HCl Thymidine Hypoxanthine Nitrate salts Ferric nitrate 5 AMINO ACIDS Acid Glu / Asp Pyridoxine HCI Riboflavin Thiamine HCl Phenol red Sodium bicarbonate Cell Culture World congress, 29/02/2012
  • Benchmarking of DMEM-F12 • Recipe containing 50 raw materials (inorganic salts, amino acids, vitamins, glucose, lipids...) • Comparison of dry powder media from several suppliers regarding: • Appearance • pH • Osmolality • Humidity • Particle Size • Cellular Performance 6 Cell Culture World congress, 29/02/2012
  • Raw materials: the Fundamentals of Media Visual differences of DMEM-F12 dry powder media from different suppliers • Supplier A and B: white appearance • Merck Millipore media: pinkish color (resulting from Vitamin B12) gentle production process 7 Cell Culture World congress, 29/02/2012
  • Quality of powdered Media … … starts with a reliable production process Supplier A lot Merck Millipore lots UPLC Analysis Homogeneity Commercial DMEM-F12 powder showing non homogeneous blending Supplier A Merck Millipore • Pinkish color: marker for mild raw material treatment • Brownish / yellow color: degradation of a raw material during the production process detected by UPLC Gentle production process preserves critical raw materials from degradation 8 Cell Culture World congress, 29/02/2012
  • pH value observed after solubilisation of DMEM-F12 from different suppliers Reconstituted media from supplier S exhibits a lower pH value 9 Cell Culture World congress, 29/02/2012
  • Osmolality observed after solubilisation of DMEM-F12 from different suppliers Reconstituted media from supplier S exhibits a lower osmolality 10 Cell Culture World congress, 29/02/2012
  • Turbidity analysis of DMEM-F12 from different suppliers All tested DMEM-F12 are soluble without precipitation 11 Cell Culture World congress, 29/02/2012
  • Humidity in DMEM-F12 dry powder media 12 Cell Culture World congress, 29/02/2012
  • DMEM-F12 particle size analysis by air jet sieving Two types of particle size reduction effects are observed based on the processing technology 13 Cell Culture World congress, 29/02/2012
  • DMEM-F12: CHO-S performance testing 50% difference in viable cell density between the lowest and the highest performer medium 14 Cell Culture World congress, 29/02/2012
  • Serial amplification assay of commercial DMEM-F12 dry powder media Constant performance of Merck Millipore media 15 Cell Culture World congress, 29/02/2012
  • Agenda 1 DMEM-F12 media characterization • Physico-chemical characterization • Cellular performance testing (CHO-S cells) • Fingerprinting • Performance prediction 2 3 16 Quality improvement - case studies Perspectives in future media preparation Cell Culture World congress, 29/02/2012
  • Near Infrared spectroscopy Fast, simple and non destructive method to measure physicochemical properties of powders via light absorption Applications: - Quality assessment - Identification of raw materials - Process control (homogeneity) - Chemical quantitative analysis of complex mixtures • One NIR spectrum is a fingerprint from a raw material or a mixture • Combination with multivariate data analysis analysis of large and complex datasets 17 Cell Culture World congress, 29/02/2012
  • Principle component analysis (PCA) of dry powder media: Clustering commercial media by NIR PCA of DMEM (blue), DMEM-F12 (red) and RPMI 1640 (green) Analysis by NIR-Spectroscopy (NIR-Spectrometer Bruker, Vektor MPA) 18 Cell Culture World congress, 29/02/2012
  • Principle component analysis (PCA) Correlation of cellular performance and NIR analysis Supplier C-1 Supplier S Supplier C-2 Supplier B Supplier C-3 Merck Millipore Supplier A PCA of DMEM-F12. Analysis by NIR-Spectroscopy (Bruker, Vektor MPA) 19 Cell Culture World congress, 29/02/2012
  • Agenda 1 DMEM-F12 media characterization • Physico-chemical characterization • Cellular performance testing (CHO-S cells) • Fingerprinting • Performance prediction 2 3 20 Quality improvement - case studies Perspectives in future media preparation Cell Culture World congress, 29/02/2012
  • Solubility: L-Cystine dihydrochloride: 50 mg/mL in 2M HCL (CAS: 30925-07-6) L-Tyrosine disodium salt: 50 mg/mL H2O (CAS: 69847-45-6) 21 Cell Culture World congress, 29/02/2012
  • L-Cystine dihydrochloride Milled raw material Raw material 22 Cell Culture World congress, 29/02/2012
  • Solubility of L-Cystine dihydrochloride milled in PBS @ 0.12 g/L 23 Cell Culture World congress, 29/02/2012 not milled
  • Complete Solubilisation of Media … L-Cystine 2HCl cumulative particle distribution Frequency (% particles by count) … is part of raw material and processing evaluation L-Cystine 2HCl is partially insoluble under defined media preparation conditions Milled L-Cystine 2HCl Particle size (µm) 24 Cell Culture World congress, 29/02/2012 cumulative particle distribution Frequency (% particles by count) Particle size (µm) Milled L-Cystine 2HCl is completely soluble within minutes
  • L-Tyrosine Disodium Salt Milled raw material Raw material 25 Cell Culture World congress, 29/02/2012
  • Particle size distribution L-Cystine HCl Accumulated particles (%) L-Cystine HCl L-Cystine HCl (milled) particle size reduction by factor 5.5 26 particle diameter (µm)
  • Particle size distribution L-Tyrosine 2Na salt – Supplier 1 accumulated % particles L-Tyrosine 2Na salt L-Tyrosine 2Na salt (milled) particle size reduction by factor 2.7 27 particle diameter (µm)
  • Particle size distribution of raw materials Size reduction 5.5 x 2.7 x The effect of milling is dependent from : - The type of raw material - The starting size of particles 28
  • Effect of the raw material source on the particle size distribution The effect of milling is dependent from : - The starting size of particles - The raw material source (supplier) 29
  • L-Tyrosine 2Na salt limit of solubility (supplier 2) 1 g/L (milled) 4 g/L (milled) 4 g/L (not milled) ~ 20 fold the concentration in DMEM-F12 Increased solubility of milled L-Tyrosine 2Na salt. Possible addition of at least 1 g/L L-Tyrosine 2Na salt in a chemically defined cell culture media with an amino acid load of > 9 g/L. 30
  • Perspectives in future media preparation • The raw material quality is key for optimal media performance on cellular level • Raw material selection • Raw material pre-processing • Supply chain • The production process is directly linked with batch to batch consistency and cellular performance Raw material quality Production process • Milling and blending process • Packaging • The combination of raw materials and the production process is mandatory for excellent dry powder media performance 31 Cell Culture World congress, 29/02/2012 Cellular performance
  • Acknowledgments Research and development • Dr. Joerg von Hagen • Maria Wehsling • Ronja Müller • Marc Vieten Production & Operations • Tanino Scaramuzza • Frank De Groot • Jochen Bausch Quality control & Analytics • Joachim Eichner • Dr. Robert Schiwon • Christoph Hoffman 32 Cell Culture World congress, 29/02/2012
  • Leading Pharma Raw Material Production Lifted synergies for production of mammalian cell culture media High Quality Raw Materials Industrial Golden Standard in QC / QA / Reg Years of Expertise in Media Development Expertise in Solid Powder Production 33