X ray crystallography

complete introduction, types of crystal, x-ray production, x-ray diffraction, braggs law, interference, x-ray methods, applications.

1. X-ray Crystallography Guided By: Dr. Chandana Majee (Assistant Professor) Presented By: Himanshu Singh M.Pharma (Pharmaceutical Chemistry)

2. Content Introduction Principle Types Of Crystal X-ray Diffraction Bragg’s Law Interference Thomson scattering Instrumentation & limitation Different X-ray Methods Application Of X-ray Crystallography References

3. Introduction X-ray crystallography is a technique used for determining the atomic and molecular structure of a crystal, In which the crystalline atoms cause a beam of incident x-rays to diffract into many specific directions. X-ray crystallography is a method of determining the arrangement of atoms within a crystal .

4. Principle : Beam of x-ray strikes a crystal The beam of light to spread into specific directions From the angles and intensities of these diffracted beams Crystallographer can produce a three dimensional picture of the density of electrons within the crystals . From this electron density : The mean positions of the atoms in the crystal can be determined, as well as, Their chemical bonds and Their disorder.

5. Why X-rays ? An electromagnetic wave of high energy and very short wavelength, which is able to pass through many materials opaque to light . (wavelength is 0.1 to 100 angstrom) The wavelength of X-ray photos is on the order of the distance between atomic nuclei in solids ( bonds are roughly 1.5-2.5 angstrom ). You can think of it like the waves fit nice between the atoms and fill the crystal .

6. Why Crystal ? Researchers crystallize an atom or molecule, because the precise position of each atom in a molecule can only be determined if the molecules is crystallized. If the molecule or atom not in a crystallized form, the X-rays will diffract unpredictably and the data retrieved will be too difficult if not impossible to understand .

7. Crystal system Conditions Triclinic None Monoclinic α = γ = 90° Orthorhombic α = β = γ = 90° Tetragonal a = b; α = β = γ = 90° Trigonal a = b; α = β = 90°; γ = 120° Hexagonal a = b; α = β = 90°; γ = 120° Cubic a = b = c; α = β = γ = 90° The seven crystal systems:

8. X-ray Diffraction Diffraction is the slight bending of light as it passes around the edge of an object. X-ray crystallography uses the uniformity of light diffraction of crystals to determine the structure of molecule or atom. Then X-ray beam is used to hit the crystallized molecule. The electron surrounding the molecule diffract as the X-rays hit them. This form a pattern that pattern is known as x-ray diffraction.

9. X-ray Diffraction pattern of single alum crystal

10. Bragg’s Law There is a definite relationship between the angle at which a beam of x-rays must fall on the parallel planes of atoms in a crystal in order that there be strong reflection, the wavelength of the X-rays and the distance between the crystal planes. Bragg Equation : 2d sin θ = n λ   Here, d – spacing between diffracting planes Θ - the incident angle n – any integer λ – wavelength Bragg’s law identifies the angles of the incident radiation relative to the lattice planes for which diffraction peaks occurs. Bragg derived the condition for constructive interference of the X-rays scattered from a set of parallel lattice planes.

11. Bragg considered crystals to made up of parallel planes of atoms. Incident waves are reflected secularly from parallel planes of atoms in the crystal, with each plane is reflecting only a very small fraction of the radiation, like a lightly silvered mirror. When X-ray strikes a layer of a crystal, some of them will be reflected. we are interested in X-rays that are in-phase with one another. X-rays that add together constructively in x-ray diffraction analysis in-phase before they are reflected and after they reflected.

12. These two X-ray beams travel slightly different distances. The difference in the distances traveled is related to the distance between the adjacent layers . Connecting the two beams with perpendicular lines shows difference between the top and the bottom beams. The length DE is the same as EF, so the total distance traveled by the bottom wave is expressed by:

13. Constructive interference of the radiation from successive planes occurs when the path difference is an integral number of wavelength. This is the Bragg Law. When X-rays are scattered from a crystal lattice, peak of scattered intensity are observed which correspond to the following conditions: 1. the angle of incidence = angle of scattering 2. the path length difference is equal to an integer number of wavelengths. Bragg reflection can only occur for wavelength: n λ <= 2d This is why we cannot use visible light. The diffracted beams from any set of lattice planes can only occur at particular angles predicted by the Bragg Law.

14. Interferences When an incident x-ray beam hits a scatter x-rays are emitted in all directions. Most of the scattering wave fronts are out of phase interfere destructively. Some sets of wave fronts are in phase and interfere constructively. A crystal is composed of many repeating unit cells in 3-dimensions, and therefore, acts like a 3-dimensional diffracting grating. The constructive interference from a diffracting crystal is observed as a pattern of points on the detector. The relative positions of these points are related mathematically to the crystal’s unit cell dimensions.

15. Thomson Scattering The x-ray scattering is determined by the density of electrons within the crystal. Since the energy of an x-ray is much greater than that of a valence electron , the scattering may be modeled as Thomson scattering, the interaction of an electromagnetic ray with a free electron. The intensity of Thomson scattering for one particle with mass m and charge q is:

16. Instrumentation Production of X-rays Collimator Monochromator Liquid nitrogen steam to keep crystal cold Movable mount to rotate crystal Detectors

17. Production of X-ray Discovered by: ROENTGEN in 1895 The x-rays are produced when fast moving electrons or cathode rays hit a heavy metal . Most x-rays have a wavelength ranging from 0.01 to 10 nanometers . Common device used to produce x-rays is called Coolidge tube (made up of glass) Coolidge tube is connected with two electrodes. Cathode(tungsten)-filament Anode- target metal Pressure maintained 10-6 mmHg Cathode connected to low tension battery (heating cathode).

18. Procedure For X-ray Production: Two electromagnetic field plates E1 and E2 are arranged on either side of the cathode, which controls the acceleration of cathode rays (electrons) emitted by filament. When high voltage (20kv) is produced across the electrode, the cathode rays are emitted and they hit the target which is made up of molybdenum etc. Invisible X-rays are produced at 45 degree in path rays. The distance between filament and the x-ray tube target is 1 cm . Velocity of electron is raised from zero to half the speed of light .

19. Electron travelling from cathode to anode: Projectile electron interacts with the orbital electron of the target atom. This interaction results in the conversion of electron kinetic energy into thermal energy and electromagnetic energy (in the form of X-rays and infrared radiation).

20. Collimator: In order to get a narrow beam of x-rays, the x-rays generated by the target material are allowed to pass through a collimator which consists of two sets of closely packed metal plates separated by a small gap. The collimator absorbs all the x-rays except narrow beam that passes between the gap. Monochromator: Monochromator is an optical device that transmit a mechanically selectable narrow band of wavelength of light or other radiation chosen from a wider range of wavelength available at input.

21. Types of Monochromators : 1. Filter 2. Crystal Monochromator A. Flat Crystal Monochromator B. Curved Crystal Monochromator Filter: X-ray beam may be partly monochromatized by insertion of a suitable filter. A filter is a window of material that absorbs undesirable radiation but allows the radiation of required wavelength to pass. Crystal Monochromator: It is made up of suitable crystalline material positioned in the x-ray beam so that the angle of reflecting planes satisfied the Bragg's equation for the required wavelength the beam is split up into component wavelengths. Material used- Nacl, Lithium fluoride, Quartz, etc.

22. Detectors : The x-ray intensities can be measured and recorded either by; Photographic method Counter methods a. Geiger-Muller tube counter b. Proportional counter c. Scintillation detector d. solid state semi-conductor detector e. Semiconductor detectors Both these of methods depends upon ability of x-rays to ionize matter and differ only in the subsequently fate of electrons produced by the ionizing process.

23. Photographic Method: To record the position and intensity of x-ray beam a plane or cylindrical film is used. The film after exposing to x-ray is developed The blackening of the developed film is expressed in terms of density units D given by: D=log Io/I Io – incident intensities I- transmitted intensities D- total energy that causes blackening of the film D is measured by densitometer. The photographic method is mainly used in diffraction studies since it reveals the entire diffraction pattern on a single film. Disadvantage : time consuming and uses exposure of several hours.

24. Counter Methods: Geiger-Muller tube counter : Geiger tube is filled with inert gas like Argon . Central wire anode is maintained at a positive potential of 800 to 2500 V. collision with filling gas production of anion pair X-RAY Electron central anode Positive ion-moves to outer electrode The electron is accelerated by the potential gradient and causes the ionization of large number of argon atoms, resulting in the production of avalanche of electron that are travelling towards central anode.

25. Proportional counter: Construction is similar to Geiger tube counter. Proportional counter is filled with heavier gas like xenon and krypton . Heavier gas is preferred because it is easily ionized Operated at a voltage below the Geiger plateau. The dead time is very short (~ 0.2 micros), it can be used to count high rates without significant error.

26. c . Scintillation Detector: In a scintillation detector there is large sodium iodide crystal activated with a small amount of thallium . When x-ray is incident upon crystal, the pulses of visible light are emitted which can be detected by a photo multiplier tube. Useful for measuring x-ray of short wavelength Crystal used in scintillation detectors include sodium iodide, anthracene, naphthalene and p-terphenol.

27. d. Solid State Semi-conductor Detector: In this type of detector, the electrons produced by x-ray beam are promoted into conduction bands and the current which flows is directly proportional to incident x-ray energy. Disadvantage: Semi-conductor device should be maintained at low temperatures to minimize noise and prevent deterioration. e. Semi-conductor Detectors: When x-rays falls on silicon lithium drifted detector an electron and a hole (+e). Pure silicon made up of with thin film of lithium metal plated on to one end. Under the influence of voltage electrons moves positive charge and holes towards negative.

28. Voltage generated is measure of the x-ray intensity falling on crystal. Upon arriving at lithium pulse is generated. Voltage of pulse = q/c Where , q- total charge collected on electrode c- detector capacity.

29. X-ray Diffraction Methods

30. Laue method The Laue method is mainly used to determine the orientation of large single crystal while radiation is reflected from or transmitted through a fixed crystal. The diffracted beams form arrays of spots, that lie on curves on the film. The Bragg angle is fixed for every set of planes in the crystal. Each set of planes picks out and diffracts the particular wavelength from the white radiation that satisfies the Bragg law for the values of d and θ involved. Determine the orientation of single crystals by means of illuminating the crystal with continous spectrum of x-rays.

31. Transmission Laue Method In this method, the film is placed behind the crystal to record beams which are transmitted through the crystal. One side of the cone of Laue reflections is defined by the transmitted beam. The film intersects the cone, with the diffraction spots generally lying on an ellipse. Back Reflection Laue Method In this method, the film is placed between the x-ray source and the crystal. The beams which are diffracted in a backward direction are recorded. One side of the cone of Laue reflections is defined by the transmitted beam. The film intersects the cone, with the diffraction spots generally lying on hyperbola.

32. Rotating crystal method In the rotating crystal method, a single crystal is mounted with an axis normal to a monochromatic x-ray beam . A cylindrical film is placed around it and the crystal is rotated about the chosen axis . As the crystal rotates, set of lattice planes will at some point make the correct Bragg angle for the monochromatic incident beam, and at that point a diffracted beam will be formed.

33. Lattice constant of the crystal can be determined by means of this method; for a given wavelength if the angle θ at which a reflection occurs is known, can be determined. The reflected beams are located on the surface of imaginary cones. By recording the diffraction patterns (both angles and intensities0 for various crystal orientations, one can determine the shape and size of unit cell as well as arrangement of atoms inside the cell.

34. Photographs can be taken by: Complete rotation method: In this method series of complete revolutions occur. Each set of a plane in a crystal diffracts four times during rotation. Four diffracted beams are distributed into a rectangular pattern in the central point of photograph. 2. Oscillation method: The crystal is oscillated at an angle of 15 or 20 degree. The photographic plate is also moved back and forth with the crystal. The position of the spot on the plate indicates the orientation of the crystal at which the spot was formed.

35. Powder method (debye-scherrer) X-ray powder diffraction is a rapid analytical technique primarily used for phase identification of a crystalline material and can provide information on unit cell dimensions (lattice parameter) . Types of material to be determined: finely ground, homogenized and average bulk composition.

36. Procedure of Powder method: A very small amount of powdered materials is sealed into a fine capillary tube made from glass that does not diffract x-rays(fine powder is struck on a hair with a gum). The specimen is placed in the Debye-Scherrer camera and is accurately aligned to be in the center of the camera. X-rays enter the camera through a collimator The powder diffracts the x-rays in accordance with Braggs law to produce cones of diffracted beams. These cones intersect a strip of photographic film located in the cylinder camera to produce a characteristics set of arcs on the film. When the film is removed from the camera, flattened and processed, it shows the diffraction lines and the holes for the incident and transmitted beams.

37. If the angle of incidence = θ then, the angle of reflection = 2 θ If the radius = r , the circumference = 2 π   r correspond to a scattering angle of 360 degree. θ = 360 * 1/ π  r From the above equation the value of θ can be calculated and substituted in Bragg’s equation to get value of d. There is no need to rotate the specimen. Applications: Useful for determining the complex structures of metals and alloys. Characterization of crystalline materials. Identification of fine grained minerals such as clays and mixed layer clays that are difficult to determine optically. Determination of unit cell dimensions. Measurement of sample purity.

38. Application f XrD XRD is a nondestructive technique. Some of the uses of x-ray diffraction are: Differentiation between crystalline and amorphous materials Determination of the structure of crystalline materials Determination of electron distribution within the atoms, and throughout the unit cell. Determination of the orientation of single crystals Determination of the texture of polygrained materials Measurement of strain and small grain size, etc. Miscellaneous application Soil classification based on crystallinity Analysis of industrial dusts Assessment of weathering & degradation of minerals & polymers Study of corrosion products Examination of tooth enamel & dentine Examination of bone state & tissue state Structure of DNA & RNA

39. Advantages of x-ray: X-ray is the cheapest, the most convenient and widely used method. X-rays are not absorbed very much by air , so the specimen need not be in an evacuated chamber. Disadvantage of x-ray: They do not interact very strongly with lighter elements Limitation: Cannot identify amorphous materials. No depth profile information. Minimum spot size of 50 micrometer.

40. Applications of XRC HIV: scientists also determined the x-ray crystallographic structure of HIV protease , a viral enzyme critical in HIV’s infection. Pharmaceutical scientists hoped that by blocking this enzyme, they could prevent the virus from spreading in the body. Arthritis: to create an effective painkiller in case of arthritis that doesn’t cause ulcers, scientists realized they needed to develop new medicines that shut down COX-2 but not COX-1. In Dairy Science: this technique has been a widely used tool for elucidation of compounds present in milk and other types of information obtained through structure function relationship . The mineral constituents and lactose are the only true crystalline constituents in dairy products that can be analyzed. Analysis Of Milkstones: this technique also applied for analyzing the chemical composition of milk stones . Since each chemical compound gives a definite pattern on a photographic film according to atomic arrangement, x-rays can be used for qualitative chemical analysis as well as structural analysis .

41. 5 . Differentiation Of Sugar: since each crystalline compound gives a definite pattern according to the atomic arrangement, the identification of the common sugars ( sucrose, dextrose and lactose )is made simple by x-rays. 6 . In Case Of New Material: x-ray crystallography is till the chief method for characterizing the atomic structure of new materials and in discerning materials that appear similar by other experiments. 7. X-rays Analysis Of Milk Powder: this technique has also been used in study of milk powder. Most work has been confined to determine the effect of different milk powdering processes upon structural group spacing's within the milk proteins.

42. References: https://www.slideshare.net/AkashArora45/xray-crystallography-its-applications-in-proteomics https://www.slideshare.net/bharathpharmacist/81347482-xraydiffractiontechnique-39635806 https://www.slideshare.net/HasanulKarim2/x-ray-crystallograpy https://www.slideshare.net/MartinJacob13/x-ray-crystallography-for-mpharm