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STABILISING PROTEINS BY SPRAY DRYING WITH ADJUVENTS Geoffrey Lee Friedrich Alexander University, Erlangen. Why spray dry a protein ? The spray drying process: machines & process conditions Two examples of spray-dried pharmaceutical proteins Sources of damage to proteins during spray drying
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STABILISING PROTEINS BY SPRAYDRYING WITH ADJUVENTSGeoffrey LeeFriedrich Alexander University, Erlangen • Why spray dry a protein ? • The spray drying process: machines & process conditions • Two examples of spray-dried pharmaceutical proteins • Sources of damage to proteins during spray drying • Formulation measures to stabilize proteins carbohydrates, surfactants • Single droplet drying via levitation • Is spray drying a potentially useful process for my product ?
Why Use Spray Drying for Proteins ? • Spray drying (SD) of protein-containing systems is not new ! • Advantages: robust, standard equipment, process development straightforward, relatively inexpensive, scale up; Disadvantages: needs exact in-process control, yield optimization required, minimization of deposit formation, 'continuous' process. • Applications of SD proteins in pharmacy: - inhaleable powders; - injectable powders; - stable, flowable storage-form for bulk protein.
Process Variables Control Product Quality •Independent process variables: - Drying Air Inlet Temperature, Tinlet - Drying Air Relative Humidity, % RH - Drying Air Flow Rate, vda - Liquid Feed Flow Rate, Vlf- Atomising Air Flow rate, vaa • Dependent variable: - Outlet Air Temperature, Toutlet
Laboratory Scale Spray Dryer: Niro Mobile Minor Drying Capacity up to 7 kg/hr; Maximum Tinlet 350°C; 3' x 6' x 6' high
Pilot Scale Spray Dryer: Niro P6.3 Drying capacity up to 60 kg/hr; Chamber 1.6 m x 0.8 m x 60°; Size 11.5' x 9' x 15'
Micro Spray Dryer: Büchi B-290 Drying capacity up to 1.5 kg/h; size 500 x 600 x 1000 mm
Feasibility of spray drying a protein • Product quality (peptide/protein) investigated by: - activity loss (enzymes) - change in aggregation status (HPLC, SEC) - gel electrophoresis: eg, isoelectric focussing - alteration in FT-IR amide bands • Example: model protein trypsinogen (Tzannis & Prestrelski, 1999) - ca 15 % activity loss on SD at Tin/Tout = 110oC/70oC - ca 20% loss of monomer (SEC) • 2 further examples of pharmaceutical proteins illustrate use of analytical techniques…
Example I: A low molecular weight peptide • Substance: Peptide with 20 amino acids = ca. 2.5 kDa • SD conditions: - Büchi 191 Micro Spray Dryer • Liquid Feed: - 2 mg/mL peptide (very low !)
Example I: SEM Appearance Residual Moisture = 2.85 % w/w
Example II: A high molecular weight protein • Substance: IgG (AMG 162) with MWt ca. 150 kDa • SD conditions: - Büchi 191 Micro Spray Dryer • Liquid Feed: - 115 mg/mL IgG Sorbitol
Example II: SEM appearance Residual moisture = 4.4/5.0 % w/w
Potential sources of protein damage Drying air 2. Shearing forces Nozzle Atomizing air 1. Adsorption Liquid feed 3. Liquid/air interface expansion 4. Thermal stress Drying tower
The 2 periods of droplet drying Various morpholgies Critical point Constant-rate phaseT = approx. Twetbulb Falling-rate phase T Toutlet Residence time: 1s – 25s eg, Tinlet/Toutlet = 130oC/90oC
Primary formulation measure to reduce protein damage • Glass-forming carbohydrates or amino acids can reduce level of protein damage - prevent unfolding & aggregation - which carbohydrate/protein mass ratio ? during SD: water replacement mechanism after SD: glassy immobilisation ? • Low residual moisture content ensures high glass transition temperature, Tg important for protein storage stability • Sufficient storage stability of carrier ? amorphous systems are hygroscopic must prevent moisture uptake & crystallisation also deterioration in powder properties
Secondary formulation measures • Addition of surfactant to liquid feed can reduce protein surface excess at water/air interface of atomised liquid feed • Use of non-aqueous solvents for peptides with low aqueous solubility higher w/v improves particle formation • Polymers to taylor particle morphology eg, dextrans or hydroxy ethyl starches eg, surfactants
Thermal inactivation of lactate dehydrogenase (LDH) in trehalose
Improvement of process stability of LDH in trehalose + 0.1 % g/g Polysorbat 80
Development of a spray drying process for a protein • Laboratory scale SD of protein solution: - sufficient water solubility = alternative solvent - does damage to protein occur ? (aggregation) - residual moisture content OK ? - which process conditions give best result ? • Which formulation measures are necessary ? - do I need a carbohydrate ? Probably yes. - which protein/carbohydrate weight ratio ? (maximize) - adjustment of required particle size = useage ? • 2. Move to pilot scale machine: - depends on required process throughput (kg of powder per h) - upscale increases residence time in chamber - can I use the same nozzle setup ?
Single-droplet drying kinetics of carbohydate solution I II III IV
Final Particles Removed from Levitator catalase/trehalose (6:4) catalase
Summary & Conclusions • Spray drying is one of a number of processes that canbe used for the production of fine particles. • It is an established technique where much expertise andexperience is available. • Development can be performed under GMP conditions. • The selection of a suitable machine & process conditionshas a (fairly) sound scientific basis. • The product capacity can be adjusted within wide boundaries. • The powder properties can also be taylored by processor formulation. • Potential problems: some questions need to be addressed: - how do I obtain a high product yield ? - how do I minimize protein damage ? - how much stabilizing adjuvent do I need, and which one is the best for my protein ? - what is the patent situation ?