Document Sample

Presented by,
K.S.ARUN KUMAR,          Under the Guidance of,
M.Pharm(Pharmaceutics)   Mr. RAHUL NAIR,
                                       M.Pharm, (Ph.D).,
                         Associate professor,
                         Department of Pharmaceutics,
                         SVCP, Tirupati.



  a well-known protein, widely used in medical applications,
  plays an important role in the formation of tissues and organs,
  is involved in various functional expressions of cells,
  can form fibers with extra strength and stability through its
   self-aggregation and cross-linking,
 the use of collagen in biomedical application has been
   rapidly growing.

 Collagen contains a large number of reactive terminal and side chain

   residues such as amino and hydroxyl group.

 Crosslinking agents:

  Chemical: Glutaraldehyde, Formaldehyde, Polyepoxy compounds
  Physical: UV/Gamma rays

 Types of collagen:
  Several types of collagen can be formed by variation of two major
    peptide chains
  19 types
  Type 1 & Type 2 are most important

 Due to its more advantages and its safety than other polymers it is
  selected as a drug carrier in the present work

     •Biodegradable and biocompatable;
     •Formulated in a number of different forms;
     •Biodegradability can be regulated by cross-linking;
     •High cost of pure type I collagen;
     •Variability of isolated collagen
     •Hydrophilicity which leads to swelling and more rapid release;
     •Complex handling properties;
            Microsphere is a term used for small spherical particles, with
diameters in the micrometer range (typically 1μm to 1000μm (1mm))
 Microspheres are the colloidal drug delivery system.
 are characteristically free-flowing powders consisting of proteins/synthetic
   polymers that are biodegradable in nature.

Classification of microspheres:

    Microspheres can be manufactured from various natural and
      synthetic materials.

Route of administration:
    Microspheres may be introduced into the human body by various
        anatomical route.
    The choice of the route of administration depends on the disease,
        the effect desired, drug properties
                         Oral delivery
                         Parenteral delivery etc.

Mechanism of drug release:
              i.        Degradation controlled monolithic system
              ii.       Diffusion controlled monolithic system
              iii. Diffusion controlled reservoir systems
              iv.       Erodable polyagent system

Etodolac (ED):
  ED is a member of NSAIDs
  ED is a racemic mixture of [+]S and [-]R-enantiomers.
  ED is insoluble in water but soluble in alcohols, chloroform, dimethyl
   sulfoxide, and aqueous polyethylene glycol.
  pKa of 4.65
  Melt. point - 145–148 °C
  molecular formula is C17H21NO3

Mechanism of action:

             The mechanism of action of ED, like that of other NSAIDs, is related
to prostaglandin synthetase inhibition.

         Rheumatoid arthritis
        For the management of acute pain


            ACE-inhibitors, Antacids, Aspirin, Diuretics, Glyburide, Lithium,
    Phenylbutazone, Phenytoin, Warfarin


           Less bioavailability due to poor water solubility,
           Absorbed in GIT,
           99% bound to plasma proteins,
           Metabolised in liver, has less halflife,
           Excretion by renal pathway.

Objective of selecting etodolac:

            ED is poorly water soluble and has less halflife and
  bioavailability. So to increase the solubility, bioavailability and for
  longer duration of action we have aimed to formulate ED into a
  microsphere using collagen as a biodegradable drug carrier

       Spray drying
       Solvent evaporation
       Precipitation – Chemical cross linking
       Multiple emulsion method
       Thermal cross-linking
       Complex Coacervation
       Ionotropic gelation
       Wet inversion

           Collagen
           Polyvinyl alcohol (PVA)
           Glutaraldehyde (GTA)
           Methylene chloride
           Etodolac
           Acetic acid
           Sodium metabisulphite

             Solvent evaporation method

Preparation of microspheres

collagen (1% w:v) was pre-swollen for 1 h in water
pH adjusted to 3.5 using acetic acid.
ED was dissolved in the collagen dispersion with 2 ml of 0.25% PVA
sonicated for 30 min
poured into methylene chloride slowly with continuous homogenization
Methylene chloride was evaporated and was mixed with varying concentrations of
(8–18% v:v) glutaraldehyde
After stirring for 4 h, 2 ml of sodium metabisulphite solution was added
stored in the freezer overnight at _5°C.
microspheres were separated by centrifugation at 5000 rpm for 15 min,
 washed with water and finally freeze dried

Sl.No           CONTENTS             F1               F2

1.      Etodolac                    300mg            600mg

2.      Collagen                    30mg             30mg

3.      Polyvinyl alcohol       22ml(0.25%w/v)   22ml(0.25%w/v)

4.      Glutaraldehyde           10ml(8%w/v)      10ml(8%w/v)

5.      Methylene chloride          50ml             50ml

6.      Sodium metabisulphite        2ml              2ml

7.      Acetic acid                   qs               qs

8.      Water                         qs               qs

Process variables which have an effect on the entrapment efficiency
of the drugs in collagen microspheres:
 Many factors affect the entrapment efficiency of the drugs in the collagen
                 nature of the drug,
                 collagen concentration,
                 drug polymer ratio,
                 stirring speed

Factors affecting the drug release from collagen microspheres:

           molecular weight and concentration of the collagen,
           the cross linking agent used and it’s concentration,
           process variables like stirring speed, type of oil, additives,
           cross linking process used,
           drug collagen ratio, etc                                          15
     The planned schedule of the work is as follows:
        Preparation of collagen from bovine skin
        Preparation of collagen based microspheres
        Physicochemical evaluation of the microspheres
            Morphology
            Size analysis
            Melting point
            Drug content determination
            Invitro release
            Release kinetics


     is to be analysed by scanning electron microscopy (SEM).
     Surface morphology of particle can be observed.

Size analysis:

      carried out by using a compound microscope
      average size of the particles can be determined

Invitro release:

    Specified quantity of microspheres is to be dispersed in 7.4 phosphate
   buffer and placed in a thermostated shaker water bath with a vibratory
   motion of 80 rpm. Samples (2ml) were periodically withdrawn and the
   absorbance was measured at 279nm .

Release kinetics:
         Three different mathematical equations were employed to
   model the dissolution profiles

        (a) First order model can be expressed as
                               Mt / M∞ = 1-e-kt
        (b)Higuchi square root of time model is given by
                               Mt / M∞ = k t 1/2
         (c) Hixson and Crowell cube root equation is denoted by
                         ( 1- Mt / M∞)1/3 = 1- k t

     Mt is the amount of drug released at time t,
     M∞ is the maximal amount of drug released at infinite time, and
     k is the rate constant associated with the drug release


            Collagen microspheres offer unique carrier system for many
pharmaceuticals. Collagen microspheres are prepared by solvent
evaporation technique for controlled drug delivery. In future plan of
work we hope to prepare collagen microspheres in different
formulations and study the physiochemical evaluation studies and drug
release kinetics of the formulated drug.


O.C.M. Chan, K.-F. So, B.P. Chan had studied on Fabrication of nano-
fibrous collagen microspheres for protein delivery and effects of
photochemical crosslinking on release kinetics. This study reports a novel
collagen microsphere-based protein delivery system and demonstrates the
possibility to use photochemical crosslinking as the secondary retention
mechanism for proteins.
Ta-Jen Wua, Hsiu-Hsuan Huanga, Cheng-Wen Lana, Chi-Hung Linb, Fu-
Yin Hsua, Yng-Jiin Wanga, studied on the microspheres comprised of
reconstituted collagen and hydroxyapatite. these results indicate that
osteoblast cells are capable of proliferating, differentiating and mineralizing
in the matrix of the microspheres, and suggest that the microspheric
composite is a potential grafting material for future clinical applications.
Yang, Chunlin Belle Mead, New Jersey 08502 (US), Timmer, Mark Jersey
City, New Jersey 07302 (US) had invented the processes for making
crosslinked collagen microspheres and microparticles.
Daniel S. Kohane,Julie Y. Tse,Yoon Yeo, Robert Padera, Maria
Shubina,Robert Langer had studied on biodegradable polymeric
microspheres and nanospheres for drug delivery in the peritoneum.
Rahul Nair, B.Haritha Reddy, C.K.Ashok Kumar, K.Jayraj Kumar had reported on
Application of chitosan microspheres as drug carriers : a review. This review aims to compile
the various application of CS in microspheres based drug delivery. This review also aims to
include the process variables factors that affect the release of drugs from the microspheres.

Lagarce , E. Garcion , N. Faisant , O. Thomas, P. Kanaujia , P. Menei , J.P. Benoit had studied
on the development and characterization of interleukin-18-loaded biodegradable

Harjit Tamber, Johansen, Hans P. Merkle, Bruno Gander had studied on the formulation
aspects of biodegradable polymeric microspheres for antigen delivery. This review provides an
applicable summary of different formulation routes for the purpose of producing safe,
qualified and efficacious products of microencapsulated peptide and protein antigens.

Natesan S, Babu M, Christy R had studied the ‘Adipose derived stem cell delivery into
collagen gels using chitosan microspheres’. This study provides a model to capture pluripotent
stem cells, expand their cell number within a biomaterial scaffold in vitro and deliver within
an appropriate matrix to repair damaged tissue.

Berthold A, had studied on ‘Collagen microparticles: carriers for glucocorticosteroids’. The
results of this study demonstrate that collagen microparticles can be successfully used as a
carrier system for lipophilic steroids.

Rössler B, Kreuter J Scherer D. had studied on the ‘Collagen microparticles: preparation and
properties’. Collagen microparticles can be used as carriers for lipophilic drugs e.g. retinol,
tretinoin, or tetracaine and lidocaine in free base form. Another feature of the biodegradable
CMPs is their thermal stability, thus their sterilization can be readily achieved.          21
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