proteins are chains of amino acids, each joined to it neighbor by a specific type of covalent bond. The polymerization of L-α-amino acids by peptide bonds forms the structural framework of proteins. The term protein is used for molecules composed of over 50 amino acids. The term peptide is used for molecules composed of less than 50 amino acids. The chemical and structural complexities involved demand an effective delivery system in which the physicochemical and biologic properties, including molecular size, conformational stability, solubility, sensitivity to light, moisture and heat, biological half-life, immunogenicity, dose requirements, susceptibility to break down in both physical and biological environments, requirement for specialized mechanisms for transport across biological membranes are to be considered. Peptide and Protein Structure It is essential to have an idea about structure of protein and peptide in order to deal with various problems encountered while developing drug delivery system. The proteins are relatively large molecules with complex structure. The peptide chains in peptides and proteins are seldom linear and adapt a variety of specific folded three dimensional patterns and conformations. All peptides and proteins are polymers of amino acids connected via amide linkages referred to as peptide bonds. • Primary structure: It denotes the number and specific sequence of amino acids. • Secondary structure: Arrangement of individual amino acids along the polypeptide backbone. • Tertiary structure: Three dimensional arrangement of a single protein molecule. • Quaternary structure: Proteins that contain two or more polypeptide chains associated by noncovalent forces

1. Delivery Routes of Proteins and Peptides Drug
By miyanda mungala petty
M.Pharmaceutics
CTIPS

2. Introduction
• proteins are chains of amino acids, each joined to
it neighbor by a specific type of covalent bond.
• The polymerization of L-α-amino acids by peptide
bonds forms the structural framework of
proteins. The term protein is used for molecules
composed of over 50 amino acids.
• The term peptide is used for molecules
composed of less than 50 amino acids.

3. introduction
• The route of administration has a significant impact on the therapeutic
outcome of a drug.
• Recent advances in pharmaceutical biotechnology, by virtue of the
biophysical and biochemical properties, have made specific route of
delivery as well as the design of the delivery system.
• Thus, routes which were of minor importance as parts of drug delivery in
the past have assumed added importance in protein and peptide delivery
and these include
• oral,buccal,nasal,transdermal,pulmonary ,rectal,parenteral and ocular
routes

4. General consideration in formulation design of protein
drugs
• Barriers to peptide and protein delivery
Enzymatic barriers
Intestinal barriers
Capillary barriers
Blood brain barrier

5. • Successful delivery of peptide and proteins
drugs is determined by its ability to cross
various biological barriers.
Enzymatic barriers
• Most important barrier limiting peptide
protein absorption in git due to protein
degradation mainly by hydrolytic cleavage of
peptide bonds by proteases like

6. • Insulin degrading enzyme .proteolysis is an irreversible
reaction. potentiating damage on proteins,
• For all kinds of systemic delivery except intravenous,
one of the first barriers for absorption is the
permeation across a cell layer.
• Being charged, large and hydrophilic, proteins are
notoriously poor permeators (poor bioavailability)

7. • Therefore, it is often necessary to add enhancers to the
protein formulation.
• which enhance their absorption across membranes and their
stability is very significant.
• The physical size of the protein drugs and their susceptibility
to degradation are key determinants of their delivery route

8. Oral route of delivery
• oral route is unsuitable for the systemic delivery of therapeutic
peptides and proteins because of the potential degradation by the
strongly acid environment in the stomach and by the proteolytic
enzymes in the intestinal tract, as well as presystemic elimination in
the liver.
• For such drugs to be absorbed through the gastrointestinal tract,
they must be protected from enzyme and must traverse through
the luminal barriers into the blood stream in an unchanged form.
• Currently only two peptide and protein based drugs (Interferon
alpha and human growth hormone) that can be given orally are
known to be in clinical development

9. Approaches for oral delivery of
peptide and protein
Approach Typical examples
Modifications by chemical synthesis of
prodrugs and analogues
PEG derivates ,monosacharides derivates
Use of enzyme inhibitors Bacitracin,chymostatin,aprotinin
Use of penetration enhancers EDTA ,sodium deoxy cholate ,sodium
laurly sulphate ,oleic acid
Carrier systems w/o/w emulsions ,emulsomes ,nano and
microparticles ,bioadhesive systems

10. Modifications by chemical synthesis of prodrugs and
analogues
• The strategy of altering the peptide/protein
structure by reversible (prodrug )or
irreversible (analogue )
• Aimed at transiently modifying
physicochemical properties of drug like
lipophilicity,charge,molecular size
,solubility,enzyme lability,and affinity to
carriers without compromising inherent
parent drug properties

11. • This approach manipulates pharmacokinetic
parameters ,improve therapeutic value of
parent drug, facilitates membrane
permeation, providing stability against
degradation, thus altering bioavailability

12. Use of Enzyme inhibitors
• Various enzyme inhibitors have been employed to achieve
successful delivery of peptide and protein based
• drugs.
• Examples: Metalloprotease inhibited by EDTA,
Enzymeaminopeptidases are inhibited by Bestain & Bacitracin,
Enzyme metalloendoproteases inhibited by Phosphoramidon.
• This approach has been used successfully for the delivery of insulin
and vasopressin.
• Insulin with enzyme inhibitor (Aprotinin, bacitracin, betatin) which
result in significance reduction in insulin digestion and improve in
its intestinal absorption profile

13. Use of Penetration Enhancers
• Peptide/protein drug moieties, due to their
molecular size, often require penetration
enhancers to achieve therapeutically
significant levels of luminal absorption.
• Mechanism of absorption of protein and
peptide drug is via Trans cellular & Para
cellular route.

14. Classification of intestinal permeation
enhancers
Surfactants Ionic:
Sodium lauryl sulphate
Sodium dodecylsulphate
Dioctyl sodium sulfosuccinate
Nonionic:
Polysorbitate
Nonylphenoxypolyoxetyylenes
Tween80
Bilesalts & its derivative Sodium glycholate
Sodium deoxycholate
Sodium taurocholate
Sodium dihydrofusidate
Sodium glycodihdro fusidate
Sodium glycolate

15. Fattyacids & its derivatives
Oleic acid
Caprylic acid
Lauric acids
Sodium caprate
Acyl carnites
Acyl choline
Sodium caprylat
Chelating agents EDTA
Citric acid
Salicylates
Chitosans & derivatives N-sulfanto-N,O-carboxymethylchitosan
N-trimethylated chloride(TMC)
Chitosan glutamate

16. Other enhancers
Zonula occludens toxin (Zot)
polycarbophyl-cysteine conjugate(PCP-Cys)

17. Carrier systems
• This strategy is particularly applicable in the case of
poorly absorbed peptides/proteins, which are unstable
in the Gastro intestinal (GI) lumen and their targeting
to a specific tissue or organ is to be affected.
• The proper designing of the delivery system not only
protects the drug from gastrointestinal degrading
components in the physical environment of the
formulation prior to absorption, but also localized the
drug at or near the cellular membrane to maximize the
driving force for passive permeation.

18. Various strategies employed are
• Lipid carriers and emulsions
• The most common type of lipid vesicles is liposomes.
• Since liposomes comprises of bilayers with an aqueous
core, both lipid soluble and water soluble drugs can be
• encapsulated.
• Drug denaturation during encapsulation is minimized
as they are formed under mild conditions.
• Solid lipid nanospheres and fat emulsions can also be
used.

19. • Lipid molecules can form other type of
particulates like immune stimulating complexes
(ISCOM) andcochleates.
• ISCOMs are three dimensional cages with a
diameter of 30nm to 70nm and can be formed by
mixing lipids, cholesterol and saponin (Quil A).
The potent immuno adjuvant properties of Quil A
render ISCOMs suitable for oral delivery of
antigens.

20. • Cochleates are phospholipids-calicum
precipitates
• and have a typical structure of a large continuous
solid lipid bilayer sheet rolled up into a spiral. The
• calcium ions keep the cochleates in their rolled
up forms.
• On removing calcium ions with chelating agents
the cochleates unroll and form large liposomes.

21. Emulsomes
• Emulsomes are colloidal drug carrier units. It is
typically a lipoidal drug delivery vehicle and could
be prepared using relatively higher concentration
of lecithin (5-10%).
• The interesting feature of the system is that
unlike oil phase of an emulsion (o/w), the internal
phase in the case of emulsomes remains to be in
solid or quasi-solid state at ambient
temperatures.

22. • Insulin (w/o/w oil being palmitic acid in octyl-
Decyl triglyceride) and the internal phase contain
macromolecule (protein /peptide drug) for oral
administration. This system holds promise for its
effective utilization in oral administration of
protein /peptide.
Particulate Carriers
• The particulates employed as delivery vehicles
can be replicating and non-replicating in nature.

23. • The replicating
systems comprise of attenuated or genetically modified
strains of viruses and bacteria, which continue to
propagate in vivo after administration, e.g. genetically
engineered Vaccine virus and attenuated strains of
Salmonella.
The non-replicating particulate systems are polymeric
particles and lipid containing particles. They encapsulate
the drugs within the particles and thereby lend a
protective coat .

24. Nasal routes route of delivery
• less technologically demanding than pulmonary
delivery is nasal delivery. Due to relatively rapid drug
absorption, possible bypassing of presystemic
clearance and relative ease of administration, delivery
of drug by the nasal route offers an attractive
alternative for administering systemically active drugs.
• Simple nasal drops or a nasal spray, nasal gel can be
used, and for particulate nasal delivery the particle size
is not as important. A special aspect of nasal delivery is
the possibility of achieving delivery transsynaptically
directly into the brain using nanoparticles

25. • The nasal epithelium suited for permeation
has an area of approximately 150 cm2, and
this will limit the dose range given by this
route. Higher bio-availabilities can be
obtained with more advanced delivery
systems, especially by adding enhancers that
modulate the permeability of the epithelium

26. • Pharmaceutical drugs as well as endogenous
hormones such as luteinizing-hormone-releasing
hormone LHRH, thyrotropin-releasing hormone
(TRHvasopressin,calcitonin, oxytocinACTH,
glucagon, insulin, interferons, and enkephalins
have been shown to be absorbed nasally in
animal and human.
• a number of peptide-based pharmaceuticals have
demonstrated that systemic bioavailability can be
improved by nasal route.

27. • hydrophilic peptide and protein which furthermore can
be degraded in the nasal cavity by peptidase and
absorption considerably smaller for peptide calcitonin
and insulin bioavailability.
• To overcome the barrier to nasal absorption of these
molecules, two main approaches have been utilized;
• modification of permeability of nasal membrane by
employment of absorption enhancer, such as
surfactants, bile salts, cyclodextrins, phospholipids, and
fatty acids, and use of the mucoadhesive system such
as bioadhesive, liquid formulation (e.g. chitosan)

28. • microsphere powder and liquid gelling,
formulation that decreases the mucociliary
clearance of the drug formulation and thereby
increase contact time between the drug and
site of the absorption.

29. Pulmonary route of delivery
• Pulmonary administration is an attractive route of
proteins and peptides than other alternative routes of
administration. The lungs offer a large surface area for
drug absorption, of approximately 80-140 m2.
• The alveolar epithelium is very thin (approximately
0.1–0.5 mm thick), thereby permitting rapid drug
absorption. The alveoli can be effectively targeted for
drug absorption by delivering the drug as an aerosol,
with a mass median aerodynamic diameter of less than
5 μm.

30. • Furthermore, the first-pass metabolism of the
GIT is avoided. Although metabolic enzymes
are found in the lungs, the metabolic activities
and pathways may differ from those observed
in the GIT, and this makes the pulmonary
administration of many peptides and proteins
very promising

31. • Devices such as jet or ultrasonic nebulizers, metered-dose
inhalers (MDI), and dry powder inhalers are used. MDIs are
the most frequently used aerosol delivery systems,
whereas, dry powder inhalers are designed to deliver
drug/excipient powder to the lungs.
• These inhalers are typically used to deliver bronchodilators
or corticosteroids. These are very effective for delivery of
the drugs to the upper airways by the device called as
spacers have been added, to be used with MDIs in order to
remove some of the non-respirable particles, by impaction
on their walls and valves

32. • The pulmonary route, through aerosol delivery systems is for the
administration of drugs molecules to treat pulmonary diseases,
such as asthma.
• The absorption chemical enhancers, which increase the
permeability of drugs through the epithelial membranes without
causing any tissue damage, are especially useful for the delivery of
peptide and protein drugs.
• The surfactants, bile salts and fatty acids have been evaluated as
absorption enhancers and, although most of them exhibit
permeation-enhancing effects, they also produce membrane
damage. Polyoxyethylene (PE) oleyl ether also showed good
enhancing ability for the peptide.

33. Buccal route of delivery
• The buccal mucoadhesive formulations are to be an
alternative to the conventional oral small amount of
medicaments as they can be readily attached to the
buccal cavity retained for a longer period of time and
removed at any time. The epithelium of the mouth is
accessible with small surface area approximately 100
cm2.
• Buccal adhesive drug delivery systems using matrix
tablets, films, layered systems, discs, microspheres,
ointments and hydrogel systems have been applied.

34. • A number of formulation and processing factors can
influence properties and release properties of the buccal
adhesive system.
• The formulations designed for buccal administration should
contain the following functional agents:
• Mucoadhesive agents, to maintain an intimate and
prolonged contact of the formulation with the absorption
site; penetration enhancers, to improve drug permeation
across mucosa (transmucosal delivery) or into deepest
layers of the epithelium (mucosal delivery); and

35. • enzyme inhibitors, to eventually protect the drug from the
degradation by means of mucosal enzymes.
• There are numerous important considerations that
including biocompatibility (the drug/device and
device/environment interfaces), reliability, durability;
environmental stability, accuracy, delivery scalability and
permeability which are to be considered while developing
such formulations. While biocompatibility is always an
important consideration, other considerations vary in
importance depending on the device application.

36. • Bioadhesive formulations designed for buccal
application should exhibit suitable rheological and
mechanical properties, including pseudoplastic or
plastic flow with thixotrophy, ease of application, good
spreadability, appropriate hardness, and prolonged
residence time in the oral cavity.
• These properties may affect the ultimate performance
of the preparations and their acceptance by patients

37. • The buccal mucosa represents a potentially important
site for controlled delivery of macromolecular
therapeutic agents, such as peptides and protein drugs
with some unique advantages
• such as the avoidance of hepatic first-pass metabolism,
acidity and protease activity encountered in the
gastrointestinal tract. Another interesting advantage is
its tolerance (in comparison with the nasal mucosa and
skin) to potential sensitizers

38. Transdermal drug delivery
Advantages :
• Better and improved patient compliance
• Elimination of hepatic first pass phenomenon
• Controlled administration is possible and
thereby
• avoidance of toxic effects. Also drugs with
• shorter half-life can be administered

39. • TDD represents a convenient, patient-friendly
option for drug delivery with the potential for
flexibility, easily allowing dose changes according
to patient needs and the capacity for self-
regulation of dosing by the patient.
• The noninvasive delivery of TDD makes it
accessible to a wide range of patient populations
and a highly acceptable option for drug dosing.

40. • Administration of drugs with low therapeutic
index is possible.
• Limitations of Transdermal Route for
peptide/protein Delivery are:
• A low rate of permeation for most protein drugs
• due to their large molecular weight and
• hydrophilicity and lipophilic nature of the stratum
corneum
• High intra and inter patient variability

41. Various approaches for Transdermal delivery Route of
peptidal drugs are:
• Iontophoresis
• Phonophoresis
• Penetration enhancers
• Prodrugs
• Iontophoresis
• Iontophoresis is a method that induces migration of
ions or charged molecules when an electric current is
allowed to flow through an electrolyte medium. To
undergo

42. • Iontophoresis protein/and peptide molecules
must carry
• charge. To achieve this pH and ionic strength of
solution
• are controlled. Protein/and peptide (charged
molecules)
• are repelled by the same charge on electrode and
• penetrate through the skin under the influence of
electric
• current.

43. • The two electrodes are placed on the stratum
• corneum, one of the electrode drug is loaded
(reservoir
• electrode) and current is applied which increased the
permeability of skin and drug molecule flow through
• epidermis →→→ dermis→ papillary layer →→
subdermal tissue→→blood circulation.
• Example: Insulin, TRH, Vasopressin, Leuprolide are
• successfully delivered by this technique.

44. • Phonophoresis
• In this method, ultrasound is applied via a coupling
• contact agent to the skin. The drug absorption is
• enhanced via thermal effect of ultrasonic waves and
• subsequent temporary alterations in the physical
• structure of the skin. It may be presumably due to
• fluidization of bio membrane.
• Example: Insulin and Erythropoietin.

45. Penetration enhancers
• The impervious nature of the stratum corneum is a
majorbarrier to achieve good drug flux through the skin.
• A popular solution to this problem is incorporation of
penetration enhancers into transdermal products.
Penetration enhancers have the properties of reversibly
reducing the barrier resistance of the horny layer and
thereby increasing the amount of drug reaching the living
tissue. Oleic acid, dimethylsulphoxide, surfactants and
azone have been used as Penetration enhancers.

46. • These agents fluidize the intracellular lipid lamellae of
stratum corneum and increase the pore which helps in
penetration of drug molecule.
Prodrugs
• Another strategy that ensures some promising results
especially with small peptides is based on
prodrugs/analogues. The enzymes present in the skin
selectively regenerate the active drug. Prodrug with
modeled physico-chemical characteristics permeated
well across the skin than drug.

47. Trans ferosomes
• Trans ferosomes are phosphatidylcholine based
supramolecular aggregates designed to be
sufficiently deformable so that they can cross the
intact skin barrier.
• These carriers contain at least one polar
amphiphilic component (e.g. cholate) and thus
the resultant vesicle membranes are extremely
flexible in their disposition

48. The ocular route of drug delivery
Mechanism of Drug Absorption By Ocular Route
• Barrier to ocular route is;
• Tear dilution
• Lachrymal drainage
• Protein binding
systemic delivery of peptide/protein drugs has been
attempted through the ocular route. The concept behind
ocular drug delivery to the systemic circulation exploits the
stable dynamics of the lachrymal system that exports the drug
to the nasal cavity from where considerable systemic
absorption results.

49. • Attempts have been made through this route
for administering insulin. However, palpable
movements and tears swiftly wash the insulin
solution away.
• To address this problem, viscosity of the
insulin solution has been increased by sodium
hyaluronic acid.

50. • The feasibility of ocular peptide/protein delivery
using eye drops as a delivery system is limited.
The eye drops exhibit low bioavailability, low
therapeutic efficacy and short duration of activity.
• To address these limitations eye inserts can be
employed.
• Another device based on absorbable gelatin
sponge has been successfully used to improve
upon the above mentioned limitations.

51. The treatment can be terminated simply by removal
of the device from the eye.
• Have been tried to deliver the drug in retinopathy
due to diabetic
• The first approach in ocular drug delivery system
is that to prolong the contact time by
incorporating various
• PolymersPVA (polyvinyl alcohol), PVP (polyvinyl
pyrrolidone), MC, CMC, HPMC.

52. • The device is fabricated by punching a disc of
gelatin. The drug solution is sorbed into the disc
and the wet matrices dried under vacuum. This
device has been employed for insulin delivery.
• The benefits of this device are- Relatively simple
and cheap manufacturing procedure.
• On hydration the device becomes soft and
pliable

53. RECTAL ROUTE
Advantages of Rectal route are:
• It is highly vascularized.
• It avoids to a large extent the first pass or
• presystemic metabolism.
• It is suitable for drugs that can cause
nausea/vomiting and irritate the gi mucosa on
oral administration. In case of adverse reaction or
drug overdose, the drug absorption can be
interrupted.

54. • A large dose of drug can be administered.
• Drug can be targeted to the lymphatic system.
Various factors affecting absorption from the rectal route
are:
Amount of liquid present in the rectum.
• pH and buffer capacity of the rectal fluid.
• Surface tension and viscosity of the rectal fluid.
• Luminal pressure exerted by the rectal wall which
enhances rectal absorption.
• Solubility, partition coefficient, pKa of the drug.
• Particle size and surface properties of the drug.

55. • The conventional dosage forms including gels,
solutions and suppositories have been used
for peptidal delivery.
• Among these, gels were found to offer an
optimal balance between retention at the site
of administration and rate of peptide release.
• Most of the peptide/protein drugs require
absorption enhancers to attain a reasonable
level of absorption.

56. Parenteral route
• Parenteral mode of drug delivery has been the major route
of choice for protein/peptide, owing to their poor
absorption and metabolic instability when given by other
alternative routes. Potent nature of these moieties
demands their targeting to specific receptors to improve
therapeutic index of a drug.
• If peptides are presented at high dosage levels, there
stands the possibility of generation of immune responses
and other undesirable deleterious side effects and
interactions. Targeting thus protects both the drug and
body from these contraindicative manifestations.

57. • The parenteral drug delivery system includes
Intravenous,
• intramuscular, subcutaneous, intraperitoneal,
intrathecal use.
• The drug carrier systems employed for defined
and controlled delivery of drug through this route
are particulates, soluble carriers and
miscellaneous systems as discussed below:
PARTICULATES
• Microspheres

58. • These are solid spherical particles in the particle
size range of few tenths of a micrometer to
several hundred micrometer, containing
dispersed drug in either solution or
microcrystalline from.
• Advantages:
• They can be administered subcutaneously,
intramuscularly or intraperitoneally and thus
implantation of the delivery system is not
obligatory.

59. • Using an appropriate technique and
subsequent optimization they can be
prepared economically.
• Disadvantages:
• Drug release may be poorly defined.
• They may interact or form complexes with the
blood components.

60. Nanoparticles
• They are similar to microspheres but their particle size is in
the nanometer range (10-100nm). They can be employed
for the targeted delivery of peptide/protein.
• Owing to their small size they can even pass through the
sinusoidal spaces in the bone marrow and spleen. To
enhance their specificity, moieties with targeting potential
like monoclonal antibodies can be attached to the
nanoparticles.
• The typical constitutive polymers include polybutylcyano-
acrylate, polymethacrylate, albumin, acrylic resins, chitosan
etc.

61. Liposomes
These serve as a “depot”, releasing the drug slowly following
enzymatic degradation Liposomes protect the entrapped
peptides from enzymatic degradation on intravenous
administration
• Phosphatidylcholines (lecithin) are the main components
used for the preparation of liposomes.
• Liposome membranes are semi-permeable and can thus be
used as controlled release systems .Liposomes form an
important means of targeting drugs directly to the liver

62. • Disadvantage is Phosphatidylcholines are easily
susceptible to oxidation. So, surfactant vesicles
called as “niosomes” are developed.
Advantages
• Flexibility in size, shape and structure.
• Relatively non-toxic disposition.
• Ability to encapsulate both hydrophilic and
lipophilic peptides/protein

63. Disadvantages:
• The constituent phospholipids have an inherent
tendency to interact with peptides/proteins.
• This can adversely affect their release kinetics
and shelf life of liposomal preparation.
• Poor viability to commercial scale production of
liposomes.

64. Emulsions
• Colloid sized emulsion droplets can be utilized for
parenteral delivery of peptides.
• This delivery system can be of great significance
and utility in protecting hydrophilic or lipophilic
drugs from direct contact with body fluids and
also in delivering the drug over a prolonged
period of time. Multiple emulsions can further
prolong the release of drug.
• Ex: delivery of influenza vaccine and diphtheria
toxoid in emulsion.

65. Thankyou
Any questions ?
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