This is our 7th SEM Practice School work related to Chemistry. This ppt contains various docking, characterization of the anti-bacterial drug Sulfacetamide, one of the Sulfonamides.

1. Presented By,
ANAS. H*
170090658
AMEENA KADAR K. A*
170090656
[VIIth SEMESTER B. PHARM]
GRACE COLLEGE OF PHARMACY
SYNTHESIS, CHARACTERIZATION AND
MOLECULAR DOCKING OF
SULPHACETAMIDE
Under the Guidance of,
Dr. Baskar L
M. Pharm, PhD
Head of Pharmaceutical
Chemistry
Grace College of Pharmacy
1

2. ACKNOWLEDGEMENT
It is a great pleasure to express our gratitude to our guide Dr. Baskar L
M. Pharm, PhD and Head of Pharmaceutical Chemistry and sincere thanks
to the whole CHEMISTRY department.
We honestly express warm regards to our principal Dr. Y Haribabu M.
Pharm, PhD and vice principal Dr. C I Sajeeth M. Pharm, PhD.
We also extend our sincere thanks to all the supportive mentors of our
college.
2

3. CONTENTS
 INTRODUCTION
 AIM & OBJECTIVES
 PLAN OF WORK
 EXPERIMENTAL SECTION
 RESULT ANALYSIS
 CONCLUSION
 BIBILOGRAPHY
3

4. INTRODUCTION
 A Drug, or a pharmaceutical, is a substance used to prevent or cure a
disease or ailment or to alleviate its symptoms.
 Drugs are very essential for the welfare of human beings.
 To meet the requirements, several multidisciplinary approaches are
required for the process of drug discovery and development.
 Drug discovery is a multifaceted process, which involves the
identification of a drug, chemical therapeutically useful in treating and
management of a disease condition
 The process of drug discovery includes the identification of drug
candidates, synthesis, characterization, screening, and assays for
therapeutic efficacy.
 It takes about 12 - 15 years from discovery to the approved medicine.
.
4

5. DRUG DISCOVERY
Approval
New Drug Application
Clinical trials
Investigational New Drug
Preclinical research
Formulation and development
Product characterization
Lead optimization
Lead identification
Target validation
Target identification
5

6. DRUG DESIGN
 It is the process of finding new medications based on the knowledge of a
biological target.
 In the most basic sense, drug design involves the design of molecules that
are complementary in shape and charge to the biomolecular target with
which they interact and therefore will bind to it.
 Various approaches;
 Rational drug design
 Insilico/ Computer Aided Drug Design (CADD)
 De Novo Drug Design
6

7. COMPUTER AIDED DRUG DESIGN (CADD)
 CADD also known as In silico screening has become powerful technique
because of its utility in various phases of drug discovery and
development.
 It can help in identifying drug targets via bioinformatics tools.
 They can also be used to analyse the target structures for possible
binding active sites, generate candidate molecules, check for their drug
likeness, dock these molecule with the target, rank them according to
their affinities, further optimize the molecules to improve binding
characteristics.
7

8. 8

9.  LBDD methods focus on known ligands for a target to establish a
relationship between their physico-chemical properties and activities
referred to as structure-activity relationship (SAR), that can be used for
optimization of known drugs or guide the design of new drugs with
improved activity.
 SBDD (structure of 3D target protein is known, hence test ligands
calculated based on affinity after docking).
9

10.  Molecular Docking
 It denotes ligand binding to its receptor or target protein.
 It is used to generate multiple ligand conformation and orientation and the
most appropriate ones are selected.
 Docking is a molecular modeling technique that is used to predict how a
protein (enzyme) interacts with small molecules (ligands).
Molecular
Docking
Rigid
Docking
Flexible
Docking
10

11.  Basic steps involved in Molecular Docking:
Visualization
Docking
Receptor Grid Formation
Ligand Preparation
Protein Preparation
11

12. Overview of Molecular Docking
12

13. LIGAND
 Sulfacetamide is a sulfonamide, that is sulfanilamide acylated on the
sulfonamide nitrogen.
 It inhibits bacterial folic acid synthesis by competing with para amino benzoic
acid.
 With a broad spectrum of action, it is used as an anti-infective topical agent to
treat skin infections and as an oral agent for urinary tract infections.
S N H 2
O
O
H N
C
O
C H 3
13

14. Target – DIHYDROPTEROATE SYNTHASE
 PDB ID – 1AJ0
 Dihydropteroate synthase (DHPS) is essential for the de novo synthesis of
folate in prokaryotes, in lower eukaryotes such as protozoa and yeast, and in
plants.
 DHPS is absent in mammals.
 This makes it an ideal drug target.
DIHYDROPTEROATE
SYNTHASE
DIHYDROPTEROIC ACID
PTERIDINE + PABA
14

15. SOFTWARES USED IN DOCKING
USE SOFTWARE FEATURES
1. Drawing tools Chem Draw • Helpful for the analysis of NMR
and Mass spectroscopic studies.
• Useful for MOT, VBT and also
for Hybridization.
Chem Sketch • Gives the physical properties of
molecules.
• Covert structure to SMILE form.
2. Data Banks RCS PDB • Information about the 3D shapes
of proteins, nucleic acids, and
complex assemblies.
TTD • Information about the known and
explored therapeutic protein
and nucleic acid targets, the
targeted disease, pathway
information and the
corresponding drugs directed
at each of these targets.
15

16. 16

17. 17

18. 18

19. Pubchem • Freely accessible chemical
information
Drug Bank • Informations of drug
interactions, pharmacology,
chemical structures, targets,
metabolism
ChEMBL • ChEMBL is a database of
bioactive drug-like small
molecules, it contains 2-D
structures, calculated properties
(e.g. logP, Molecular Weight,
Lipinski Parameters, etc.) and
abstracted bioactivities (e.g.
binding constants,
pharmacology and ADMET
data)
3. Energy
Minimization
MOE • An integrated computer-aided
molecular design platform.
• Mainly used for the
macromolecules.
19

20. 20

21. 21

22. Chem 3D Pro • To generate 3D models
• To minimize the energy of
ligand.
4. Prediction Pass Prediction • A prediction of activity spectra
for substances.
Swiss Target
Prediction
• To predict the most probable
protein targets of small
molecules.
5. Protein
Validation
Ramachandhran Plot
Server & PROCHECK
• Main-Chain Conformational
Tendencies of Amino Acids.
22

23. 7. Pharmacokinetic
Parameters &
toxicity
Swiss ADME • To compute physicochemical
descriptors as well as to predict
ADME parameters,
pharmacokinetic properties,
drug like nature and medicinal
chemistry friendliness of one or
multiple small molecules to
support drug discovery.
ADMET SAR • Free tool for the prediction of
chemical ADMET properties
based on QSAR.
8. Chemical Tool
Box
Open Babel • Chemical toolbox designed to
speak the many languages of
chemical data.
6.
Chemoinformatics
Molinspiration • Molecule manipulation and
processing, including SMILES
and SD file conversion,
generation of tautomers,
calculation of various molecular
properties needed in QSAR.
23

24. AutoDock Vina • Significantly improves the
average accuracy of the binding
mode predictions compared to
AutoDock 4.2.
PatchDock • An object recognition and image
segmentation techniques used in
computer vision.
• Online docking program.
SwissDock • A web server program dedicated
to the docking of small
molecules on target proteins
9. Docking Tools AutoDock 4.2 • Suite of automated docking tools.
• It is designed to predict how
small molecules, such as
substrates or drug candidates,
bind to a receptor of known 3D
structure.
24

25. 10. Visualization
Tools
Discovery Studio • An interactive package with a
broad variety of features for
modelling and simulation.
PyMol • An open source, user-
sponsored, molecular
visualization system.
Chimera • Program for the interactive
visualization and analysis of
molecular structures and
related data.
25

26. SYNTHESIS OF SULFACETAMIDE
 The discovery of sulfonamides is a significant milestone event in the human
chemotherapeutic history.
 Sulfonamides are synthetic compounds that have activity against both gram-
positive and gram- negative bacteria.
 Originally they were synthesized in Germany as azodye known as prontosil.
 Prontosil breaks down in the human body to produce Sulfanilamide which is
the active agent that kills streptococcus bacteria.
 Sulfacetamide found highly successful use in fighting urinary tract infections
starting in 1941.
26

27. RECRYSTALLIZATION
 Separation of a mixture of solids or purification of solids are most readily
achieved by recrystallization.
 The basic principles employed in recrystallization is the dissolution of the
crude material in a suitable solvent and filtration is the necessary step, to
remove any suitable impurities in the solution.
FILTRATION TECHNIQUES
 Technique used to separate solids from liquids, is the act of pouring a
mixture onto a membrane (filter paper) that allows the passage of liquid (the
filtrate) and results in the collection of the solid.
 Two filtration techniques are generally used in chemical separations in
general chemistry lab: “gravity filtration” and “vacuum/suction”
filtration.
27

28.  Suction filtration is a technique which allows for a greater rate of filtration.
 Whereas in normal filtration gravity provides the force which draws the
liquid through the filter paper, in suction filtration a pressure gradient
performs this function.
 It is used in recrystallization experiments.
28

29. DRYING
 Drying has been defined as the process whereby moisture is vaporized from
a material and is swept away from the surface, sometimes under vacuum, but
normally by means of carrier gas which passes through or over the material.
THIN LAYER CHROMATOGRAPHY
 TLC is simple, convenient and useful technique because it is quick, cheap
and accurate and easy to use.
 Useful to separate the required product from the mixture.
 This technique is helpful to monitor organic reactions, similarly in quality
control studies on stability testing, shelf life determination or degradation.
29

30. Overview of TLC Method
30

31. MELTING POINT DETERMINATION
 The temperature at which a solid melts and becomes a liquid is the Melting
Point.
 Hence, different compounds tend to have different melting points.
 A pure, nonionic, crystalline organic compound usually has a sharp and
characteristic melting point.
 A mixture of very small amounts of miscible impurities will produce a
depression of the melting point and an increase in the melting point range.
 Consequently, the melting point of a compound is a criterion for purity as
well as for identification.
31

32. SPECTRAL CHARACTERIZATION TECHNIQUES
 The most widely used methods of organic compound identification is that to
ensure the interactions of compounds with electromagnetic radiations of
different wavelength.
 There are a number of analytical technique which facilitates to identify and
characterize the compound, some of them are:
i. Ultra Violet -Visible Spectroscopy
ii. Infrared Spectroscopy
 UV-Visible Spectrophotometer
 Fourier Transform Infra-Red Spectrophotometer
32

33. AIM AND OBJECTIVES
 AIM
The purpose of the study is to carry out Synthesis, Characterization And
Molecular Docking Of Sulphacetamide.
 OBJECTIVES
 To design and characterization of crude drug of Sulphacetamide.
 Calculation of lipinskis rule and drug likeness score using
Molinspiration software.
 Determination of pharmacokinetic properties and toxicity using
Swiss ADME and ADMET SAR.
 Determination of pharmacological effects by Pass Prediction.
 Molecular docking of lead molecule towards Dihydropteroate
synthase using Autodock 4.2, Autodock Vina, PyRx, Patch dock
and Swiss dock.
 To carry out the spectroscopic (FTIR, UV), TLC and Melting Point of
Sulphacetamide.
33

34. PLAN OF WORK
REVIEW OF LITERATURE
Insilico screening
Synthesis & Characterization
Elemental Analysis
UV FTIR
TLC
MP
Autodock 4.2
Autodock Vina
Swiss Dock
Patch Dock
Autodock with PyRx
34

35. EXPERIMENTAL SECTION
i. Docking of sulphacetamide by different software
1. AutoDock 4.2
 It is a suite of automated docking tools.
 It is designed to predict how small molecules, such as substrates or
drug candidates, bind to a receptor of known 3D structure.
 It actually consists of two main programs: autodock performs the
docking of the ligand to a set of grids describing the target
protein; autogrid pre-calculates these grids.
 In addition to using them for docking, the atomic affinity grids can be
visualised.
35

36.  Protein Preparation
 Available in RCS PDB, TTD, download it in the PDB format.
 Refine it by using MOE.
 Protein ID – 1AJ0
 Open Autodock window File Read Molecule Open Protein
 Give Charges: > Edit Charges Add kollman charges
 Saving file: > Save as PDBQT , where Q Charges & T Type
of Atom
36

37.  Ligand Preparation
 The structures were built by ChemDraw Ultra 8.0 version, ACD Chem
sketch and saved in PDB format.
 Energy Minimization - Chem 3D Pro
 Open Autodock window Ligand Open Choose File
 Ligand Torsion tree Choose Root Detect Root Choose
torsions Set number of torsions.
 Ligand Output Save as PDBQT.
 Grid Formation
 Grid Macromolecule Choose
 Grid Set map types Choose ligand grid box confine the
receptor to the grid box file close saving current
 Grid Output Save as GPF (Grid Parameter File).
37

38.  Docking
 Docking Macromolecule Set rigid filename Receptor
 Docking Ligand select default parameters Accept Search
parameters Genetic algorithm Accept
 Docking Lamarckian GA (4.2) Save as Ligand.dpf (Docking
Parameter File)
 Run Autogrid Program path name Select autogrid4.exe.file
Launch
 Run Autodock Select autodock4.exe and select ligand.dpf
Launch.
 Visualization of Docking Result
 Analyse Docking Open Choose ligand.dlg (docking log file)
 Analyse Macromolecule
 Analyse Conformations Load
 Analyse Docking Show interactions
 Analyse Clusterings Show
38

39. 2. AutoDock Vina
 It significantly improves the average accuracy of the binding mode
predictions compared to AutoDock 4.
 Pre-calculating grid maps (Vina does that internally), and some other
implementation tricks, such as pre-calculating the interaction between
every atom type air at every distance.
 It also uses the same type of structure format (PDBQT) for maximum
compatibility with auxiliary software.
3. SwissDock
 It is a docking web server, the structure of the target protein, as well as that
of the ligand, can be automatically prepared for docking.
 The interpretation of docking results and their integration into existing
research pipelines is greatly facilitated by the seamless visualization of
docking predictions in the UCSF Chimera molecular viewer.
39

40. 4. PatchDock
 It is a geometry-based molecular docking algorithm.
 It is aimed at finding docking transformations that yield good molecular
shape complementarity.
 SYNTHESIS OF SULFACETAMIDE
NH2
SO2NH2
(CH3CO)2O
Sulphanilamide
-H20
NHCOCH3
SO2NHCOCH3
Partial deacetylation
Acetylated Sulphanilamide
NH2
SO2NHCOCH3
Sulphacetamide
40

41. RECRYSTALLIZATION
 Dried Drug + few ml of Ethanol/Hot water.
FILTRATION
 Done by Suction/ Vacuum filtration.
 CHARACTERIZATION OF SULPHACETAMIDE
 Melting Point Determination
 Thin Layer Chromatography (TLC)
 Spectral Studies
i. UV – Visible Spectroscopy
ii. Fourier Transform Infra- Red Spectroscopy (FTIR)
41

42. THIN LAYER CHROMATOGRAPHY
 TLC Plate – Coating the plate with Silica gel
 Mobile Phase – A mixture of 50 volume of 1-Butanol + 25 volume of ethanol +
25 volume of water + 10 volume of strong ammonia solution.
 Test Solution – Dilute a suitable volume with water to produce a solution
containing 4%w/v of Sulphacetamide.
 Reference Solution – 0.2%w/v solution of Sulphanilamide in Water.
 Detecting Method – Iodine Chamber.
UV- VISIBLE SPECTROSCOPY
 Calculate the λmax with the help of “UV Probe” software.
FOURIER TRANSFORM INFRA-RED SPECTROSCOPY
“IR Solution” software helps to determine the FTIR peak, which determines the
Structural Conformations of the particular drug.
42

43. RESULTS AND DISCUSSION
1. CHEM DRAW
NH2
H3COCHNO2S
N-(4-aminophenylsulfonyl)acetamide
Sulphacetamide
C8H10N2O3S
Exact Mass: 214.04
Mol. Wt.: 214.24
C, 44.85; H, 4.70; N, 13.08; O, 22.40; S, 14.97
Boiling Point: 799.04 [K]
Log P: -0.31
MR: 52.03 [cm3/mol]
Henry's Law: 4.9
CLogP: -0.976
CMR: 5.2618
43

44. 2. CHEM 3D PRO
44

45. 3. CHEM SKETCH
NH2
S
O O
NH
O
N-(4-aminobenzene-1-sulfonyl)acetamide
Nc1ccc(cc1)S(=O)(=O)NC(C)=O
Sulphacetamide
Molecular Formula: C8H10N2O3S
Formula Weight: 214.2416
Composition: C(44.85%) H(4.70%) N(13.08%) O(22.40%) S(14.97%)
Molar Refractivity: 52.21 ± 0.4 cm
3
Molar Volume: 155.9 ± 3.0 cm
3
Parachor: 424.7 ± 6.0 cm
3
Index of Refraction: 1.584 ± 0.02
Surface Tension: 55.0 ± 3.0 dyne/cm
Density: 1.373 ± 0.06 g/cm
3
Dielectric Constant: Not available
Average Mass: 214.2416 Da
45

46. Wireframe Sticks Ball & Stick
• Different 3D visualization by using Chemsketch 3D Viewer
46

47. 4. PASS PREDICTION
47

48. 5. Swiss Target Prediction
48

49. 6. Molinspiration
miLogP TPSA MW No.of H-
bond
acceptors
No.of H-
bond
donors
No. of
violations
No. of rotatable
bonds
Volume
-0.56 89.26 214.25 5 3 0 2 174.71
•Properties
GPCR ligand Ion channel
modulator
Kinase inhibitor Nuclear
receptor ligand
Protease
inhibitor
Enzyme inhibitor
-0.46 -0.48 -0.70 -1.28 -0.36 -0.12
•Bioactivity
49

50. 7. Lipinski’s Rule of Five
50

51. Formula Mol.Wt No. of
Heavy
atoms
No.of arom.
heavy atom
No.of
Rotatble
bonds
No.of H-
bond
acceptors
No.of H-
bond
donors
Molar
Refractivity
TPSA
C8H10N2O3S 214.24
g/mol
14 6 3 3 2 51.75 97.64
A2
Water Solubility GI Absorption BBB Permeant Bioavailability Score Log Kp
Very soluble High No 0.55 -8.29 cm/s
8. SWISS ADME
•Physicochemical Properties
9. ADMET SAR
Human Intestinal
Absorption
BBB Human Oral
Bioavailability
Carcinogenicity (binary) Eye corrosion
+ 0.9939 + 0.9764 + 0.8286 - 0.6429 - 0.9706
51

52. Eye Irritation Hepatotoxicity Acute Oral
Toxicity
Water Solubility Plasma Protein
Binding
Biodegradation
+ 0.6471 + 0.8750 2.261 -1.292 0.533 -0. 8250
10. RAMACHANDRAN PLOT SERVER
52

53. No. of residues in favored regions No. of residues in allowed regions No. of residues in outliner region
240 (97.166%) 5 (2.024%) 2 (0.810%)
11. AUTODOCK
Result Analysis
Software
Protein Ligand Docking Score Amino acid residue
AutoDock 4.2 1AJ0 Sulphacetamide -5.87 GLY217, ILE20.
ARG265, ASN 115,
ILE 117, MET139,
PHE190, LYS 221,
SER 219
AutoDock Vina 1AJ0 Sulphacetamide -5.7 SER 219, LYS 221,
ARG 255, GLY 217,
LEU 215, PHE188,
ASP 185, ASN 115,
ILE 20, PHE 190
53

54. 54

55. 55

56. 12. AUTODOCK VINA
56

57. 13. SWISSDOCK
S. No. Compound Full fitness (Kcal/mol) Estimated Δ G (Kcal/mol)
1. Sulphacetamide -1089.31 - 6.93
57

58. 14. Patch Dock
ACE
(Kcal mol-1)
Area Amino acid Residue
-239.73 330.20 PRO 145, PRO 190, GLN 149, LYS
192, GLY 65, ARG 63, THR 62, PHE
190, GLY 191, GLY 189, ALA 151,
ASN 197, MET 148
58

59. CHARACTERIZATION OF SULPHACETAMIDE
 Characterization testing is used to gain an understanding of the physical and
chemical properties of pharmaceutical materials.
a. Physical Properties
b. Chemical Properties
PHYSICAL PROPERTIES
PHYSICAL PROPERTIES
PROPERTY VALUE
Physical state Solid
Colour White Powder
Molecular Formula C8H10N2O3S
Molecular Weight 214.243
PH 7.4
Melting Point 1830C
Rf Value 0.93
59

60. SPECTRAL STUDIES
 UV-VISIBLE SPECTROSCOPY
λmax = 263nm
60

61.  FTIR
15
%T
7.5
-0
4000 3500 3000 2500 2000 1500 1000 500
1
3005.10
2833.43
2673.34
2636.69
2601.97
2098.55
1932.67
1915.31
1897.95
1784.15
.55
1438.90
188.15
1002.98
968.27
6
0
87
8
61

62. CONCLUSION
 Sulphacetamide drug had synthesized and characterized which has proven
that and leads to the path of its attractive feature of the antimicrobial activity.
 Drug likeness of the compound was confirmed by the software viz.,
Molinspiration, Lipinski’s rule of five, and pass prediction.
 Protein validation was done by the Ramachandhran plot.
 Molecular docking was performed by using AutoDock 4.2, AutoDock Vina,
SwissDock, and PatchDock and found that the drug majorly binds to amino
acids like Asparagine, Glycine, and Serine which has been matched and
supported with the reference literature.
 Pharmacokinetic parameters were evaluated by Swiss ADME, ADMET SAR
and ADRs are studied by Pass Prediction.
62

63.  Sulphacetamide was synthesized from Sulphanilamide and the product
was obtained with a good yield.
 Physicochemical properties like Melting Point, Retention factor were
performed and reported.
 The product was confirmed by FTIR (Functional groups); UV- visible
spectroscopy was done to get λmax.
 In near future, we aim to elaborate on the chemistry and
pharmacological background of the present moiety that we had taken
it.
63

64. BIBILOGRAPHY
1. Hira Saleem, Arooma Maryam, Saleem Ahmed Bokhari, Ayesha
Ashiq, Sadaf Abdul Rauf, Rana Rehan Khalid, Fahim Ashraf
Qureshi, and Abdul Rauf Siddiqi : Design, synthesis, characterization and
computational docking studies of novel sulfonamide derivatives. EXCLI
journal, Published online 2018 Feb 1, Pages: 169-179.
2. https://www.technologynetworks.com/drug-discovery/articles/target-
identification-validation-in-drug-discovery-312290
3. https://slidetodoc.com/aim-preparation-and-evaluation-of-sulphacetamide-
sodium-eye/
4. https://www.molinspiration.com/cgi-bin/properties
5. https://www.uniprot.org/uniprot/P0AC13
64

65. 6. Amol B. Deore*, Jayprabha R. Dhumane, Hrushikesh V Wagh, Rushikesh
B. Sonawane: The Stages of Drug Discovery and Development Process. Asian
Journal of Pharmaceutical Research and Development, pages: 62-66.
7. Neema Bisht and B. K. Singh: ROLE OF COMPUTER AIDED DRUG
DESIGN IN DRUG DEVELOPMENT AND DRUG DISCOVERY. IJPSR,
2018; Vol. 9(4): 1405-1415.
8. http://dx.doi.org/10.22192/ijarbs.2017.04.02.009
9. Supplementary Table S1: A list of 211 pieces of software for CADD.
Number of citations was obtained using Google Scholar, last accessed on 14th
April 2021.
10. http://dx.doi.org/10.22270/ajprd.v7i6.616
11. http://dx.doi.org/10.13040/IJPSR.0975-8232.9(4).1405-15
12. https://www.elsevier.com/locate/bfopcu
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