Protein and peptide are biopolymers which yield more than two amino acids on hydrolysis. Although the terms ‘proteins’ and ‘peptides’ are used freely, peptides are those with molecular weight below 10,000 and proteins are molecules with higher molecular weight. Most therapeutic proteins and peptide-based drugs are administered by parenteral route and are incorporated in liposomes to prolong their action or fused with Immunoglobulins or Albumin to improve their half-life. PEGylation is a proven technique for improving the potentials of Proteins/peptide delivery systems.

1. Protein and Peptide
Drug Delivery System
Presented by-
PRODIPTA CHAKRABORTY
Dept. of Pharmaceutics
Himalayan Pharmacy Institute

2. INTRODUCTION
 Protein and peptide are biopolymers which yield more than two amino acids on
hydrolysis.
 Although the terms ‘proteins’ and ‘peptides’ are used freely, peptides are those
with molecular weight below 10,000 and proteins are molecules with higher
molecular weight.
 Most therapeutic proteins and peptide-based drugs are administered by
parenteral route and are incorporated in liposomes to prolong their action or
fused with Immunoglobulins or Albumin to improve their half-life.
 PEGylation is a proven technique for improving the potentials of
Proteins/peptide delivery systems.

3. Barrier to protein and peptide delivery
1) Enzymatic barrier to protein and peptide absorption.
2) Blood-brain barrier and Blood-cerebrospinal fluid barriers.
3) Gastro- intestinal barriers.
4) Mucosal barriers.

4. 1) Enzymatic barrier-
 Degradation of proteins and peptides by enzymes occurs not only in the liver
but also at the site of administration, in the blood, in the kidney etc. Therefore,
protection of proteins and peptides against such degradations in more than
one anatomical site is necessary to maximize bioavailability. However,
protection of these substances from degradation does not necessarily mean an
increase in its stability or increased therapeutic effect.
 The half-life of peptides and proteins is only about 15 minutes due to their
rigid clearance by the liver and kidney.
 Renal clearance can be avoided by binding the administered protein/peptide
to albumin or immunoglobulins.

5.  There is preliminary evidence that the protein and peptides
degradation in the liver can be altered by changing the chemical
composition of these substances.
 For instance, Strunz et al. (1978) modified the molecular size of
gastrin which significantly altered its stability in the liver.

6. 2) Blood-Brain Barrier and Blood-CSF Barrier-
 Entry of proteins and peptides to the brain is restricted by two barriers that exist between the
CNS and the blood- BBB and Blood-CSF Barrier. Surface area of BBB is 5000 times greater than
Blood-CSF barrier so BBB is considered the primary barrier.
 The Blood-Brain Barrier is composed of highly specialized cells that are difficult to permeate
through for proteins and peptides. This barrier is a tight intercellular junction of endothelial
cells.
 Presence of cells such as pericytes and astrocytes forms a solid envelope around the brain
capillaries.
 Therefore, a drug may enter the brain through transcellular route not paracellular route.

7.  Blood-cerebrospinal fluid is formed by the choroid plexus of the
lateral, third and fourth ventricles and is similar in composition to
the ECF of brain. The capillary endothelium that lines the choroid
plexus have a open junction and the drugs can flow freely into the
extracellular space between the capillary wall and the choroidal
cells. The choroidal cells are joined to each other by tight junctions
forming the blood-CSF barrier.
 A drugs that enters the CSF slowly cannot achieve a high
concentration because of bulk flow of CSF continuously removes
the drugs.

8. 3) Gastro- intestinal Barrier-
 The gastrointestinal(GI) tract contains many endo and exopeptidases, enzymes that hydrolyze
peptide bond and act synergistically to degrade protein and peptides. For these it is important
to have both qualitative and quantitative data for oral delivery.
 In the gastro-intestinal tract, enzymatic degradation can occur at the lumen, brush border,
mucosal mesh etc. Due to metabolism of proteineous drugs reduces their bioavailability.
 The small intestine is the main site for absorption of drugs, but the efficiency and mechanism
of drugs absorption changes between the different segments of the small intestine. Towards
the distal sections, the villi becomes smaller and fewer thereby reducing the surface area of
absorption.
 For instance, it is reported that in the upper jejunum the mucosal area per cm of serosal
length is, 98cm2, whereas it is only 20cm2 at the ileal side.

10. Table 1: Summary of the gastro-intestinal barriers to peptides and proteins
absorption.
Stomach Enzymatic (enzymes) Intestine absorption.
Acidic environment
Non-specific proteolytic
enzymes like pepsin
Trypsin
Chymotrypsin
Elastase
Pepsin
Carboxypeptidase A
Carboxypeptidase B
Aminopeptidase
A limiting step is the very
poor absorption of
macromolecules either
through the enterocytes
tight junction.

11. 4) Mucosal Barrier-
 Mucus play an important role in determining the absorption and bioavailability of orally
administered drugs, the major component is the secreted glycoprotein mucin.
 It is reported that the mucin layer is thickest in the stomach and colon, whereas in he small
intestine the thickness varies depending on the extent of digestive activity.
 The digestive enzyme activity of trypsin and chymotrypsin decreases to almost one-half in the
jejunum and to one-third in the ileum.
 Aoki et al. investigated the enzymatic barrier to the absorption of insulin by invitro studies.
He reported that the apparent permeability coefficient (Papp) of insulin for the hyaluronidase-
treated segment was significantly higher than that of the control group in all small intestinal
regions.
 The Papp of insulin in both groups increased in the order duodenum<jejunum<ileum.

12.  Their study also suggested the possibility of mucous/ glycocalyx layer acting as
an enzymatic but not a diffusional barrier, irrespective of the intestinal region.
 Morishita et al. he observed the same report by using an insitu absorption
study with different intestinal segment loops. And he found that there is
increase in insulin absorption from ileum, then the distal part of the small
intestine.

13. Formulation approaches:
 Polymer which is stable in GIT but decomposes at the ileocaecal junction have
ben used for insulin delivery and found to be very effective for oral insulin
delivery.
 Chitosan-EDTA-protease inhibitor conjugates have been used for many peptide
delivery.
 Molecules-N acylated non-α ,aromatic amino acid compound was found to
increase the absorption of human growth hormone by altering the
conformation of molecule reversibly and facilitate transport across intestinal
mucosa.
 hGH was lyophilized to get stable form with reduction in aqueous solubility. It
decreases the potential for degradation in protein molecule mobility and
thereby ensured the stability and improved the bioavailability of orally
administered peptide hormones.

14. Table 2: Approaches of formulation of oral protein drug delivery system.
Protein’s nature Formulation approach
1. Unstable in solution. Lyophilization using cryoprotectants and incorporating drug into delivery
matrix as a solid powder.
2. Adsorb to delivery matrix (PLG) a) Incorporation of hydrophilic surfactants (Polysorbate 20/80,Ppluronic
F.68)
b) addition of another protein as a competitor for adsorption surface.
3) High protein concentration required
in delivery system-prone to
aggregation.
a) Addition of surfactant to reduce self association.
b) use of less soluble prodrug. eg; complexation with metal(zinc-insulin)
4) Poor stability at low pH. a) Lyophilization.
b) Formulation in high pH buffer.
c) Addition of soluble basic salt in delivery matrix to neutralize acid
degradation products of delivery matrix
d) Formulation of microporous delivery matrix rather than monolithic
device.
5) Heat sensitivity a) Using low temperature homogenization encapsulation process.

15.  Other formulation tested are emulsion, liposomes, nanoparticles, soft
gelatines coated capsules.
 Methods for production of protein formulations are emulsification,
coacervation, extrusion, spray drying and polymerization. Special procedure
like double emulsion method and prolease microsphere technology may also
be adopted.
 As proteins are more stable in solid state than in liquid, its incorporation in
solid form in delivery matrix is advantageous.

17. Reference
1) Gupta H., Sharma A., 2009, Recent trends in protein and peptide drug delivery system, Asian
Journal of Pharmaceutics, Page no: 69-70.
2) Vincent H.L.Lee., 1986, Enzymatic barriers to peptide and protein absorption and the use of
permeation enhancers to modify absorption.
3) Chris Vander Walle, 2011, Protein and Peptide delivery, Page no: 109
4) Ratnaparkhi M.P., Chaudhari S.P., Pandya V.A., 2011, Peptides and Proteins in
Pharmaceuticals, International Journal of current Pharmaceutical Research, Volume 3, Issue
2.
5) Singh. R., Saraf Shubhini A., Semalty A., 2007, Properties and Formulations of Oral Drug
Delivery System of Proteins and Peptides, Indian Journal of Pharmaceutical Science,
Volume-69, Issue-6, Page no:744-745.

18. Thank you