Evaluation of Protein and peptide delivery system

1. PROTEIN AND PEPTIDE
DELIVERY
UNDER THE
GUIDANCE OF
F.R. SHEEBA
PREPARED BY
MALAY PAUL
1st Sem. M.Pharma
Department of
pharmaceutics
MALLIGE COLLEGE OF PHARMACY
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2. Evaluation of protein and peptide drug
formulations
• Stability testing
• Bioassay
• UV spectroscopy
• Bradford assay
• Differential scanning calorimetry
• Chromatography
• Electrophoresis
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3. Stability testing
• The capability of a particular formulation in a specific
container/closure system to remain within its physical,
chemical, microbiological, toxicological and protective
specifications.
• Evaluates the effect of environmental factors on the
quality of the a drug substance or a formulated product
which is utilized for prediction of its shelf life and proper
storage conditions.
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4. Bioassay:-
 Due to complexity of proteins, bioassay are required
to assess potency of the formulation.
 Bioassay are of two types : in vitro and in vivo.
 In case of in vitro bioassays response of cells to
hormones and growth factors is monitored.
 In case of in vivo bioassay pharmacological response
of animals to proteins is monitored.
Eg: post injection BSL in rabbits is measured for
bioassay of insulin
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5. UV spectroscopy:-
• Proteins containing aromatic amino acid
residues such as phenyl alanine, tyrosine,
tryptophan can be detected by UV
spectroscopy. It can be used as an analytical
determination method
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6. Bradford assay:-
• This assay employs the principle that in the
presence of proteins in an acidic medium,
absorption maximum of coomassie brilliant blue
G-250 dye changes.
• If there is no protein to bind, the solution
remains brown.
• The dye forms a complex with carboxyl end of
proteins by Van der Waals forces to form a blue
colored solution.
• The intensity of the colored solution can be
measured using a spectrophotometer to
determine the concentration of the protein in the
sample.
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7. Differential scanning calorimetry:-
• Used as a tool for investigating
transitions of conformation as a function
of temperature and, more importantly,
the effect of potential stabilizing
excipients in a protein solution.
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8. • Chromatography:-
• To study stability of proteins and peptides.
Various modes used are
1. Normal Phase HPLC – polar S.P and non
polar M.p
2. Reverse Phase HPLC- non polar S.P and
polar M.P
3. Ion Exchange- separate ions and polar
molecule base on their affinity to the ion
exchanger
4. Chromatofocusing – separation of single
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9. Electrophoresis:-
• Sodium dodecyl sulphate polyacrylamide gel
electrophoresis (SDS-PAGE) used.
• Proteins are denatured by boiling in the SDS
solution.
• All charges of protein are masked by negative
charge of dodecyl sulphate.
• Thus protein moves on polyacrylamide gel strictly
on basis of size of protein molecule.
• This technique is useful for determining
molecular weight of proteins.
• For visualization of proteins on the gel, reagents
used are silver nitrate, coomassie brilliant blue
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10. REFERENCES:-
 A article of protein and peptide delivery
system by Nikitha Gangwani
 Protein and peptide analysis by
spectroscopy by John R chapman
 Drug delivery system by Praful,bhusan
 wikipedia.in
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