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Contents: Introduction Types and Structure of Proteins Role of Proteins Delivery Challenges Routes of Administration Modification by Chemical Synthesis Enzyme Inhibitors Penetration Enhancers Ocular Route Pulmonary Route Nasal Route Transdermal Route Parenteral Route Intra muscular Route Intra venous Route Subcutaneous Route Gourav Taneja JCDMCOP, Sirsa 1 16/01/2012

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Development of Delivery Systems Stability Profile Recent Advances in Protein Delivery Polymeric Drug Delivery Microsphere Encapsulation Polymer Scaffolds Magnetic Targeted Carriers Liposomes Hydrogel Based Drug Delivery Recent Advances in Delivery of Insulin Therapeutically Useful Peptides Conclusion References Gourav Taneja JCDMCOP, Sirsa 2 16/01/2012

Biotechnology based Pharmaceuticals:

Biotechnology based Pharmaceuticals Pharmaceutical products that are basically prepared using biological molecules. This technology uses fields like molecular biology, genetics and recombinant DNA technology. NUCLEIC ACID BASED PHARMACEUTICALS PROTEIN BASED PHARMACEUTICALS BIOTECHNOLOGY BASED PHARMACEUTICALS Gourav Taneja JCDMCOP, Sirsa 3 16/01/2012


PROTEINS Proteins are probably the most important class of biochemical molecules, although of course lipids and carbohydrates are also essential for life. Proteins are the basis for the major structural components of human tissue. Proteins are natural polymer molecules consisting of amino acid units. The number of amino acids in proteins may range from two to several thousand. Gourav Taneja JCDMCOP, Sirsa 4 16/01/2012

Types of Proteins and Peptides::

Types of Proteins and Peptides: Gourav Taneja JCDMCOP, Sirsa 5 16/01/2012


STRUCTURE OF PROTEIN Array of amino acids in protein structure Spatial array of twisted polypeptide chain(hydrogen bonds) 3-D structure of functional protein Composed array of two or more polypeptide chains Gourav Taneja JCDMCOP, Sirsa 6 16/01/2012




Gourav Taneja JCDMCOP, Sirsa 8 16/01/2012

Delivery Challenges::

With the commercialization of potential protein and peptide drugs their delivery systems have also been a challenge. Various shortcomings of these systems have been observed- Low Bioavailability. Enzyme Degradation. Protein Binding. Short Half-life. Low Permeability due to high Molecular Weight. Protein Binding. Delivery Challenges : Gourav Taneja JCDMCOP, Sirsa 9 16/01/2012

Routes of Administration::

Routes of Administration: Gourav Taneja JCDMCOP, Sirsa 10 16/01/2012

Non- Parenteral route of drug administration::

Route of delivery Challenges in delivery Oral Epithelial cell barrier, Protease degradation, Extreme pH conditions, Low bioavailability Transdermal Stratum corneum barrier, Protease degradation, Macrophages, Limits in surface area and Rate of Administration Pulmonary Epithelial cell barrier, Protease degradation, Alveolar Macrophages, Limits on the amount of Protein delivered per dose, Fixed doses, Rapid Rate of Absorption, Permeability Challenges in Smokers. Ocular Tear Turnover, Protease degradation, Protein Binding, Corneal Irritation Possibility, Low Patient Acceptance. Rectal Elimination of Drug during Bowel Movements, Low Patient Compliance, Limited Surface Area. Nasal Low Bioavailability, Protease degradation, Variable Absorption, Possibility of Nasal Mucosal Irritation and Pathological Changes (on prolonged administration). Buccal Lack of Dosage form Retention, Presence of Drug Metabolizing Enzymes (Oxidases, Reductases, Cyclooxygenases, Peptidases). Non- Parenteral route of drug administration: Gourav Taneja JCDMCOP, Sirsa 11 16/01/2012

Modification by chemical synthesis::

Modification by chemical synthesis: To overcome various problems by oral route some chemical modifications are done. This aids in manipulating the pharmacokinetic parameters to improve the therapeutic values of parent drug. These modifications are basically summarized as: Gourav Taneja JCDMCOP, Sirsa 12 16/01/2012


Gourav Taneja JCDMCOP, Sirsa 13 16/01/2012

Enzyme inhibitors::

Enzyme Specificity Trypsin Arg, Lys Chymotrypsin Phe, Tyr Elastase Als, Gly, Ile, Leu, Val, Ser Carboxypeptidase Tyr, Phe, Ile, Thr, Glu, His, Ala Enzyme inhibitors: The basic problem in the delivery of protein and peptide drug delivery systems is the degradation of the proteins by the metabolic enzymes present in the body. To overcome these problems the drugs are formulated with enzyme inhibitors so as to avoid the degradation of the proteins. Here are some enzymes that have specific activity against particular amino acids. Gourav Taneja JCDMCOP, Sirsa 14 16/01/2012

Penetration Enhancers::

Facilitate the absorption of proteins and peptides across biological membranes. These are used to overcome the problem of low permeability across the cell membrane for proteins and peptides. In transdermal drug delivery, 15-70 fold increase in the flux of vasopressin in rat is observed, with a consistent antidiuretic response. Five major types of penetration enhancers are used:- Chelators (EDTA), surfactants, bile salts, fatty acids, nonsurfactants. Penetration Enhancers: Gourav Taneja JCDMCOP, Sirsa 15 16/01/2012

Mechanism of Action of Penetration Enhancers::

Mechanism of Action of Penetration Enhancers: Compromising integrity of mucosal membrane, Inhibiting proteolytic activity, Increasing the thermodynamic activity of peptide and protein drugs. Two main pathways by which enhancers act:- The transcellular pathway, The paracellular pathway. Gourav Taneja JCDMCOP, Sirsa 16 16/01/2012

Transcellular Pathway::

Transcellular Pathway: Enhancers increase transcellular permeability to protein and peptide drugs by affecting membrane lipids and proteins. Fatty acids and their derivatives act primarily on the phospholipid component of membranes thereby creating disorder and resulting in increased permeability. Various enhancers used for transcellular pathways are: Dimethyl maleate, NSAID, Monoglycerides, Na caprylate. Gourav Taneja JCDMCOP, Sirsa 17 16/01/2012

Paracellular Pathway::

Paracellular Pathway: Paracellular pathway enhances the penetration of protein and peptides by increasing in the permeability of the paracellular pathway has been depicted as increase in the “pore” size of the mucosal membrane. Various enhancers used for paracellular pathways are: EDTA Na taurocholate N- acyl amino acids N- acyl collagen derivatives Na laurate Gourav Taneja JCDMCOP, Sirsa 18 16/01/2012

Role of ca+2 in regulating paracellular permeability::

The integrity of the junctions depend upon extracellular Ca +2 . Calcium ions restore the barrier function of mucosa followed by exposure to the enhancer, this forms an inactive complex between the enhancer and Ca +2 that increases the permeability of proteins and peptides, i.e., lowering of Ca +2 enhances the permeation of proteins and peptides. Enhancement in paracellular absorption results not only from expansion of the tight junction and the intercellular space but also from increase in water influx through the space. This is characteristic of increased water flux in the paracellular pathway when compared with that in the transcellular pathway. Increase in water flux in the transcellular pathway can be induced by diethyl maleate. Role of ca +2 in regulating paracellular permeability: Gourav Taneja JCDMCOP, Sirsa 19 16/01/2012

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Increase in water influx may affect drug absorption by one of the following mechanisms: Increase in concentration gradient for penetration, Increase in solvent drag, Increase in blood flow at the absorption site. Contd… Other means to enhance paracellular permeability: Enhancing Na + transport by increasing osmolality of the dosing solution or by promoting glucose and amino acid transport. Altering actin filaments. Studies have shown that, active transport of glucose and amino acids, which is coupled to Na + transport, across the intestinal mucosa into the intercellular lateral spaces creates an osmotic force for fluid flow, resulting in increased paracellular permeability. Gourav Taneja JCDMCOP, Sirsa 20 16/01/2012

Factors influencing effectiveness of penetration enhancers::

Physiological properties of peptide and proteins. Site of administration. Nature of vehicle. Factors influencing effectiveness of penetration enhancers: Physiological properties of peptide and proteins: Solubility, dissolution, interactions, thermostability, lipophilicity and hydrophilicity of the proteins and peptides affect the permeation of the proteins and peptides through the membranes. Two model enhancers are considered: first increased only the drug diffusion co-efficient across the membranes. second decreased the effective absorption site- viable tissue partition coefficient of the drug in addition to increasing its diffusion coefficient across the membrane. Gourav Taneja JCDMCOP, Sirsa 21 16/01/2012

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Hydrophilic drugs -------- both type of enhancers. Lipophilic drugs --------- only second type of enhancer. Contd… Site of administration: The effectiveness of enhancers varies from site to site. Medium chain glycerides, dipalmitoylcarnitine and EDTA are more active in upper than the lower part of GI tract ,i.e., these enhancers are more effective when given orally then when given rectally. Other enhancers such as polyoxyethylene 9- lauryl ether , saponin and free fatty acids are more active in lower than upper GI tract. Gourav Taneja JCDMCOP, Sirsa 22 16/01/2012

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The reason for the variation in site to site administration is due to difference in the composition of phospholipids in various epithelial cell membranes. Gourav Taneja JCDMCOP, Sirsa 23 16/01/2012

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The nature of the vehicle can affect the effectiveness of enhancers because it can affect the enhancer concentration at the absorption site. As the enhancer used has the best promoting action at specific sites only, i.e., medium chain glycerides and free fatty acids are effective in upper and lower GI tract respectively. Nature of vehicle: In recent studies it is found that Tween – type surfactants are effective in promoting the rectal absorption of sulfanilic acid in the rat when administered in an oil- in – water emulsion vehicle , but ineffective when administered in aqueous solution. Gourav Taneja JCDMCOP, Sirsa 24 16/01/2012

Safety concerns::

Safety concerns: Irritation, Extent of membrane damage, Recovery rate of mucosal membrane. Alternatives to penetration enhancers: The basic alternatives to penetration enhancers are- Iontophoresis, Phonophoresis. Iontophoresis: The use of the electric ions to enhance the absorption of the drug form the site of administration. Phonophoresis: The use of the ultrasound waves to enhance the absorption of drug from the site of administration. Gourav Taneja JCDMCOP, Sirsa 25 16/01/2012

Ocular route::

Gelfoam eye device enhances the absorption of sodium insulin with an absorption enhancer Many proteins and peptides that have been investigated for ocular delivery - Enkephalins - Thyrotropin releasing hormones, - Luteinizing hormone-releasing hormone - Glucagon and Insulin All these peptides were absorbed into the blood stream to some extent Ocular route: Gourav Taneja JCDMCOP, Sirsa 26 16/01/2012


Considerations and Opportunities for Protein/Peptide Ocular Delivery: Treatment of Local Eye Disorders Systemic Absorption of Proteins and Peptides via the Ocular Route Strategies for the Ocular Delivery of Proteins and Peptides: Prodrug Approach Mucoadhesive Particulate Carriers Engineered Nanoconstructs Penetration Enhancers Gourav Taneja JCDMCOP, Sirsa 27 16/01/2012

Pulmonary drug delivery::

Formulation approaches Benefits Limitations Liposomes Improved absorption; prolonged retention time in the lungs; biocompatible. Low drug loading and encapsulation efficiency; risk of drug leakage during nebulization process; instability during storage. Solid lipid nanoparticles Biocompatible, controlled release; prolonged retention time in the lung. Low drug loading and encapsulation efficiency; risk of degradation of biopharmaceuticals during preparation; aggregation. Polymeric nanoparticles Prolonged retention time in the lung; delayed phagocytic clearance; improved epithelia penetration; controlled release. Aggregation; long-term toxicity. Large porous microparticles Delayed phagocytic clearance; improved dispersibility and flowability; dry powder form; improved storage stability; prolonged shelf-life; controlled release. Difficult to penetrate the membrane; rehydration is needed. Nanostructured microparticles Combined advantages of nanoparticles and microparticles systems. Risk of denaturation of biopharmaceuticals in the complicated preparation process; heterogeneous dispersion of nano- delivery systems in the micro particles. Pulmonary drug delivery: Gourav Taneja JCDMCOP, Sirsa 28 16/01/2012

Nasal route::

Nasal route: Nasal route is one of the most vascularised route that provide effective systemic absorption of drugs. Provide rapid absorption. Relatively high bioavailability. Opportunities : Acceptable for most of the peptides. With permeation enhancers, peptides with 20 or more amino acids provides satisfactory bioavailability. Direct nose to brain delivery of drug can be obtained for various drugs. Local sustained delivery using bioadhesive polymers. Gourav Taneja JCDMCOP, Sirsa 29 16/01/2012

Transdermal route::

Transdermal route: The drug is formulated in the form of a patch that is applied over the skin and the absorption of drug takes place through skin. Advantages: Improved patient compliance Controlled release products can be administered. Bioavailability of drugs is higher. Disadvantages: High variation depending upon patient. Low permeation rate of drugs with high molecular weight. Variation depending upon hydrophilicity and lipophilicity of stratum cranium of skin. Gourav Taneja JCDMCOP, Sirsa 30 16/01/2012

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Gourav Taneja JCDMCOP, Sirsa 31 16/01/2012

Parenteral Route::

Parenteral Route: Protein and peptides are mostly delivered by parenteral route as it provides the best choice for efficient delivery of these drugs. Gourav Taneja JCDMCOP, Sirsa 32 16/01/2012

Devices to deliver parenteral dosage forms::

Devices to deliver parenteral dosage forms: Metered dose pens Measured dose syringes Gourav Taneja JCDMCOP, Sirsa 33 16/01/2012


INTRAVASCULAR ROUTE: The drugs are administered either by intravenous or intra-arterial route but most of the times IV route is preferred as administration through intra-arterial route is difficult. Advantages: Drugs that are highly metabolized and are highly bound to tissues by IM route are basically given by this route. EX: INSULIN, INTERFERONS AND ANTIBIOTICS. Disadvantages: Causes pain, tissue necrosis and thrombopenia. Gourav Taneja JCDMCOP, Sirsa 34 16/01/2012


INTRAMUSCULAR ROUTE: The drug is injected in the muscle. Various drugs that are used for making depots are also administered by this route. Advantages: Drugs that are administered for making depots. Better bioavailability for protein bound drugs. Disadvantages: This route is not used for drugs that are metabolized at the site of administration. Some of the drugs given by this route are LHRH, GH & LONG ACTING INSULIN Gourav Taneja JCDMCOP, Sirsa 35 16/01/2012


SUBCUTANEOUS ROUTE: Drugs are administered under the fold of skin. In case of proteins and peptides, implanted polymeric devices are used. These devices are made by cross-linking the polymers and these polymers must be biocompatible and biodegradable. Examples: Poly(d,l- lactide co-glycoside), Poly lactic acid Some of the drugs used are ACTH, CALCITONIN, INSULIN, BOVINE Gourav Taneja JCDMCOP, Sirsa 36 16/01/2012

Development of delivery systems::

For the development of a delivery systems various parameters have to be considered. Formulation considerations. Pharmacokinetic considerations. Analytical considerations. Regulatory considerations. Development of delivery systems: Gourav Taneja JCDMCOP, Sirsa 37 16/01/2012

Formulation Consideration::

Formulation Consideration: The development of delivery systems for therapeutic proteins and peptides & their evaluation depend on the biophysical, chemical, physical characters of proteins & peptides include their molecular size, t 1/2 ,etc. Formulation consideration covers various aspects as: Preformulation studies. Surface absorption behavior. Stability profile. Gourav Taneja JCDMCOP, Sirsa 38 16/01/2012

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Stability Profile Gourav Taneja JCDMCOP, Sirsa 39 16/01/2012

Physical Processes:

Physical Processes Denaturation Adsorption Aggregation Precipitation Gourav Taneja JCDMCOP, Sirsa 40 16/01/2012

Pharmacokinetic consideration::

Some modifications are done in the dosage forms to overcome various limitations. Less biological half life. Instability at absorption site. Endogenous compounds interact at absorption site. Sampling error. Presence of binding proteins. Pharmacokinetic consideration: Gourav Taneja JCDMCOP, Sirsa 41 16/01/2012

Analytical considerations::

Analytical considerations: The dosage forms are analyzed so as to determine the stability and pharmacokinetics of the drugs. The techniques used are: Radioimmuno Assay. HPLC. Liquid Chromatography. Gourav Taneja JCDMCOP, Sirsa 42 16/01/2012

Regulatory considerations::

Unlike the conventional organic based pharmaceutical protein and peptide pharmaceutical have its structure have its structure, identification, quality, purity and potency. The main regulatory authorities are FDA, EPA, USDA. FDA - Food & Drug Administrations. EPA - Environmental Protection Agency. USDA - US Department of Agriculture. Regulatory considerations: Gourav Taneja JCDMCOP, Sirsa 43 16/01/2012

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Polymeric drug delivery, Microsphere Encapsulation, Polymer Scaffolds, Magnetic Targeted Carriers, Liposomes, Hydrogel based drug delivery. Recent advances in protein delivery Gourav Taneja JCDMCOP, Sirsa 44 16/01/2012

Polymeric Drug Delivery:

Frequency of doses reduced Drug utilized more effectively Drug stabilized inside the polymer matrix Reduced side effects Possibility of dose-dumping De-activation of drug inside polymer Polymeric Drug Delivery Gourav Taneja JCDMCOP, Sirsa 45 16/01/2012

Polymeric Drug Delivery:

Polymeric Drug Delivery Polymers should be: Biodegradable Bio-compatible Non-toxic Examples: Polylactides/glycolides Polyanhydrides Polyphosphoesters Gourav Taneja JCDMCOP, Sirsa 46 16/01/2012

Polymeric Drug Delivery:

Diffusion of drug out of the polymer Governing equation: Fick’s laws of diffusion Drug release is concentration dependant Less applicable for large molecules, Polymeric Drug Delivery Gourav Taneja JCDMCOP, Sirsa 47 16/01/2012

Polymeric Drug Delivery:

Drug Release by Polymer Degradation Polymer degradation by: Hydrolysis Enzymatic (Phosphotases; Proteases etc.) Polymeric Drug Delivery Gourav Taneja JCDMCOP, Sirsa 48 16/01/2012

Microsphere Encapsulation:

Microsphere Encapsulation 100 m m Gourav Taneja JCDMCOP, Sirsa 49 16/01/2012


Process involves encapsulating protein or peptide drugs in small porous particles for protection from “insults” and for sustained release Two types of microspheres nonbiodegradable biodegradable Encapsulation Gourav Taneja JCDMCOP, Sirsa 50 16/01/2012

Types of Microspheres:

Nonbiodegradable ceramic particles polyethylene co-vinyl acetate polymethacrylic acid/PEG Biodegradable (preferred) gelatin polylactic-co-glycolic acid (PLGA) Types of Microspheres Gourav Taneja JCDMCOP, Sirsa 51 16/01/2012

Microsphere Release:

Microsphere Release Hydrophilic (i.e. gelatin) best for burst release Hydrophobic (i.e. PLGA) good sustained release (esp. vaccines) tends to denature proteins Hybrid (amphiphatic) good sustained release keeps proteins native/active Gourav Taneja JCDMCOP, Sirsa 52 16/01/2012

Polymer Scaffolds:

Incorporate drug into polymeric matrix Protection of drug from enzymatic degradation – particularly Applicable to peptide and protein drugs Release drug at known rate over prolonged duration Drug dispersed or dissolved in suitable polymer Release - diffusion of drug through polymer - diffusion through pores in polymer structure - therefore different release profiles result (dissolved or dispersed) Polymer Scaffolds Gourav Taneja JCDMCOP, Sirsa 53 16/01/2012

Release Mechanisms:

Release Mechanisms Gourav Taneja JCDMCOP, Sirsa 54 16/01/2012

Magnetic Targeted Carriers (MTCs) :

Magnetic Targeted Carriers (MTCs) Microparticles, composed of elemental iron and activated carbon Drug is adsorbed into the MTCs and transported The drug attaches to the carbon component The particles serve as delivery vehicles to the area of the tumor for site-specific targeting Gourav Taneja JCDMCOP, Sirsa 55 16/01/2012

Magnetic Targeted Carriers (MTCs):

Magnetic Targeted Carriers (MTCs) Gourav Taneja JCDMCOP, Sirsa 56 16/01/2012


Liposomes Hydrophobic Hydrophilic Spherical vesicles with a phospholipid bilayer Gourav Taneja JCDMCOP, Sirsa 57 16/01/2012

Liposomes Drug Delivery :

Liposomes Drug Delivery Potential of Liposomes in drug delivery has now realized Bloemycin encapsulated in thermo sensitive Liposomes enhanced antitumor activity and reduced normal tissue toxicity S.C injection of negatively charged Liposomes produced a prolonged hypoglycemic effect in diabetic dogs Liposomes have recently been used successfully as vehicles for vaccines Gourav Taneja JCDMCOP, Sirsa 58 16/01/2012

Hydrogel Based Drug Delivery:

Hydrogel Based Drug Delivery Hydrogels are three dimensional networks of hydrophilic polymers that are insoluble Gourav Taneja JCDMCOP, Sirsa 59 16/01/2012

Hydrogel Based Drug Delivery:

Hydrogels can swell as a result of changes in pH, Temp., ionic strength, solvent composition, pressure and the application of electric fields Hydrogel Based Drug Delivery Insulin has been one drug that has been incorporated in Hydrogels and investigated by researchers extensively. Gourav Taneja JCDMCOP, Sirsa 60 16/01/2012

Recent advances in the delivery of insulin:

Buccal aerosol delivery system developed by Generex Insulin is absorbed through thin tissue layers in mouth and throat Insulin is formulated with a variety of additives and stabilizers to prevent denaturation on aerosolization and to stabilize aerosol particles Recent advances in the delivery of insulin Gourav Taneja JCDMCOP, Sirsa 61 16/01/2012

Oral Delivery by Microsphere:

Oral Delivery by Microsphere pH 2 pH 7 Gourav Taneja JCDMCOP, Sirsa 62 16/01/2012

pH Sensitive Microspheres:

Gel/Microsphere system with polymethacrylic acid + PEG In stomach (pH 2) pores in the polymer shrink and prevent protein release In neutral pH (found in small intestine) the pores swell and release protein Process of shrinking and swelling is called complexation (smart polymers) pH Sensitive Microspheres Gourav Taneja JCDMCOP, Sirsa 63 16/01/2012

Therapeutically useful Peptides:

Peptides Application ACTH Antiallergic, decongestant, anti-inflammatory Β -Endorphin Analgesic Calcitonin Paget’s disease, hypercalcemia Glucagon Hypoglycemic crisis Insulin Diabetes mellitus Leu-enkephelin Analgesic Met-enkephelin Immunostimulant Oxytocin Induce uterine contractions Somatostatin Attenuate miotic responses TRH Diagnosis of thyroid cancer Vasopressin Diabetes insipidus VIP Secretion of insulin Therapeutically useful Peptides Gourav Taneja JCDMCOP, Sirsa 64 16/01/2012


Protein and peptide pharmaceuticals are very important class of therapeutic agents. Their emergencies on the clinical & therapeutic horizon has intensified the investigation for their convenient & effective delivery through noninvasive route. The additional challenges for the pharmacist is the designing & development of viable delivery system for non parenteral administration of protein and peptide drugs. Conclusion: Gourav Taneja JCDMCOP, Sirsa 65 16/01/2012

References: :

Lee V. Changing Needs in Drug Delivery in the Era of Peptide and Protein Drugs. Florence A.T., Siepmann J. Chemical Kinetics an Drug Stability. Modern Pharmaceutics; Volume 1; Basic Principles and Systems; 5 th edition. Informa HC USA. Page no. 235-240 Mitra A.K. Peptides and Proteins as Therapeutic Agents. Ophthalmic drug Delivery Systems; 2 nd edition. Informa HC USA. Page no. 505-508. Florence A.T., Siepmann J. Biotechnology- Based Pharmaceuticals. Modern Pharmaceutics; Volume 2; Applications and Advances; 5 th edition. Informa HC USA. Page no. 259-282 Wan F. Moller E.H. Yang M. Jorgensen L. Formulation Technologies to Overcome Unfavorable Properties of Peptides and Proteins for Pulmonary Delivery. Drug Discovery Today: Technologies 2011. Lu Y. Yang J. Sega E. issues related to Targeted Delivery of proteins and Peptides. The AAPS Journal 2006; 8(3); Article 55. Page no. E466- E478 References: Gourav Taneja JCDMCOP, Sirsa 66 16/01/2012

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Gourav Taneja JCDMCOP, Sirsa 67 16/01/2012

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Gourav Taneja JCDMCOP, Sirsa 68 16/01/2012

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