Physical Pharmacy 2 1

Particle Size Sizing Technique 2:

hydrodynamic chromatography sieving

microscopy

Kausar Ahmad

Kulliyyah of Pharmacy, IIUM

http://staff.iiu.edu.my/akausar

Physical Pharmacy 2 2

Hydrodynamic Chromatographya technique for separating particles based on their size, eluting in the order: largest to smallest. – compare with SEC

Measures complex particle size distributions in the range 5nm to 2µm

Makes no assumptions regarding the shape of the particle size distribution

Results are independent of particle density

Analysis time less than 10 minutes

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HDC Operation

A water-based eluent

is continuously

pumped through the system at

constant flow rate.

sample & a small

molecule marker

solution, are introduced

into the system.

The unit contains a separating 'cartridge'.

dynamic operating

range from e.g. 20nm to

1.2µm.

UV detector response is

used to calculate

concentration of particles of different size

present in sample.

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Results generated by HDC

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LiposomePL-PSDA (Polymer Lab: HDC mechanism)

What is the volume average diameter?

What is the polydispersity index?

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Sieving POWDER can be separated

into various size fractions by vibrating sieve loaded with sample to enable the particles of size less than that of the mesh openings to pass through and the over size to remain in the sieve.

Dry sieving is adopted for free flowing powder samples of size range varying from e.g. 4mm down to 25μm.

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Sieves A standard test sieve is generally made of a

woven wire mesh cloth specified wire thickness with square openings

Standard sieves are made according to recommended norms to maintain opening size interval between successive sieves.

The ratio between successive sieves is kept as a constant such as 1.414 and hence the sieve size varies in geometric progression.

fixed to a rectangular or circular frame

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Sieving procedure

Sieves stacked

pana

b

c

d

e

f

g

Wire mesh

Minimum size

Maximum size

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Table of Sieve ResultsSieve no./size (um)

Weight of sieve (g)

Wt of sieve & sample (g)

Wt of sample (g)

% Wt of sample

% cumulative weight

(<25) na na a ax100/M ax100/M

25 x1 y1 b (a+b)x100/M

41 x2 y2 c (a+b+c)x100/M

60 x3 y3 d

116 x4 y4 e

450 x5 y5 f

2000 x6 y6 g 100

Total M

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Interpretation of Sieve Results

From the % cumulative weight, the mean diameter can be obtained i.e.

d(0.5) – the diameter at which 50% of the samples, based on weight falls below it.

Wei

ght

(%)

Cumulative weight (%)

Microscopyhttp://www.mos.org/sln/SEM/index.html

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To fly, moths must be light. A close-up of the Cecropia Moth scale shows that it is mostly air, adding very little weight to the moth

The deer tick, is about the size of a freckle.

The shells of radiolarians, single-celled animals.

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Principles of Microscopyhttp://micro.magnet.fsu.edu/primer/lightandcolor/polarizedlighthome.html

The microscope is an instrument designed to make fine details visible.

The Concept of Magnification

• The image of an object can be magnified when viewed through a simple lens.

• By combining a number of lenses in the correct manner, a microscope can be produced that will yield very high magnification values.

Lenses and Optics

• The action of a simple lens, is governed by the principles of refraction and reflection .

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Using light microscopy equipped with graticule to determine size

Comparison of microscopesFeature Light TEM SEM

Use morphology Small particles, 40-150 nm

morphology

Source of illumination

Visible light High speed electrons

High speed electrons

Best resolution 200 nm 0.2 nm 3-6 nm

Magnification 10-1000X 500-500,000X 20-150,000X

Depth of field 0.002-0.05 nm 0.004-0.006 0.003-1 mm

Lens type Glass Electromagnetic Electromagnet

Image ray-formation spot

On eye by lenses

On phosphorescent plate by lenses

On cathode tube by scanning device

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Scanning Electron Microscope

Conventional light microscopes use a series of glass lenses to bend light waves & create a magnified image.

The SEM creates the magnified images by using electrons instead of light waves.

The SEM shows detailed 3D images at much higher magnifications than is possible with a light microscope.

The images created without light waves are rendered black and white.

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SEM Technique

Samples have to be prepared

to withstand vacuum inside

SEM.

Specimens are dried to

prevent from shriveling.

Because SEM use electrons, they must be

made to conduct

electricity. Samples are coated with a very thin layer

of gold by sputter coater.

Now the prepared

specimen is ready.

The sample is placed inside

the microscope's

vacuum column

through an air-tight door.

Inside the SEM

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After air is pumped out of

column, an electron gun emits

a beam of high energy electrons. This beam travels downward through

a series of magnetic lenses

to focus electrons to a fine spot.

Near the bottom, a set of scanning coils moves the focused beam back and forth

across the specimen, row by

row.

As the electron beam hits each

spot on the sample,

secondary electrons are

knocked loose from its surface. A

detector counts these electrons and sends the signals to an

amplifier.

The final image is built up from the

number of electrons emitted from each spot on

the sample.

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Information given by SEM

pore diameter

particle size

shape

surface condition.

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References

SL Flegler, JW Heckman, KL Klomparens, Scanning and

transmission electron microscopy, Oxford, New York

(1993)

http

://micro.magnet.fsu.edu/primer/lightandcolor/polarizedlig

hthome.html