Methods for Laboratory Studies (Focusing Area: Natural Product Drug Discovery)

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Md. Torequl Islam, PhD – Assistant Professor of Department of Pharmacy, Bangabandhu Sheikh Mujibur Rahman Science and Technology University, Gopalganj, Bangladesh

Series: Research Advances in Communication Studies
BISAC: MED072000; SCI003000
DOI: https://doi.org/10.52305/VZOO3578

In the dynamic field of drug discovery and development, non-clinical and pre-clinical studies are the foundation of success. With global relevance, “Methods for Laboratory Studies” is your comprehensive guide, offering vital insights for researchers, educators, and students worldwide.

The book begins by highlighting the crucial role of non-clinical studies, conducted with strict adherence to Good Laboratory Practice (GLP) regulations. From experimental animals to diverse therapeutic goals, it ensures the highest safety and compliance standards.

Dive deeper into early pre-clinical studies, the compass for Go/No-Go decisions. Here, you’ll uncover the core elements of a new drug’s journey, from pharmacokinetics to safety profiles, including toxicogenetics. Emphasizing GLP-compliant studies, like repeated dosage toxicity, genotoxicity, and safety pharmacology, it guarantees safe human exposure before human trials. Before entering human trials, meticulous completion of non-clinical and pre-clinical studies establishes a robust foundation for Investigational New Drug (IND) applications.

Gain insights into the comprehensive research data package bridging the gap between animal and human trials as you turn the pages. After IND approval, additional GLP tests assess carcinogenicity, genotoxicity, chronic toxicity, and reproductive and developmental toxicity. “Methods for Laboratory Studies” isn’t just a textbook; it’s your gateway to mastering drug discovery. Whether you’re a student, educator, or seasoned researcher, this book provides lucid, essential guidelines, empowering you to shape the future of pharmaceutical science.

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Table of Contents

Preface

About the Author

Chapter 1. Natural Product Drug Discovery
1.1. Brief History
1.2. Natural Products
1.3. The Role of Traditional Practices in Drug Discovery
1.4. Traditional Practice versus Technical Research
1.5. Why Pharma Companies Have Less Interest in Natural Products?
1.6. Why Do We Prefer Natural Products for New Drug Discovery?
1.7. Traditional Bioactivity-Guided Isolation Phases Associated with Problems and Solutions
1.8. Chemical Process for Natural Product Discovery
1.9. Generic Scheme for Bioassay-Guided Fractionation
1.10. An Affinity-Based Identification System for Natural Products
1.11. Current Technologies for Natural-Product Lead Discovery
1.12. Current Applications of Natural Products in Discovery
References

Chapter 2. Introduction to Non-Clinical and Pre-Clinical Studies
2.1. Non-Clinical and Pre-Clinical Studies.
2.2. In Vitro Versus In Vivo Models
2.3. Non-Clinical Study Types
2.4. Strength of Non-Clinical Study
2.5. Special Type Non-Clinical Studies
References

Chapter 3. Collection and Extraction of Materials
3.1. Collection and Management of Plant Samples
3.2. Extraction of Plant Materials
3.3. Fractionation of Crude Extract
3.3.1. Precipitation
3.3.2. Solvent-Solvent Extraction
3.3.3. Distillation.
3.3.4. Dialysis
3.3.5. Electrophoresis
3.3.6. Cautions
3.4. Chemical Investigations in Plants
3.5. Qualitative Identification of Secondary Metabolites
3.6. Example of Isolation and Identification of Compounds in a Crude Extract
3.7. Drugs from Animal Origins
3.8. Drugs from Marine Sources
3.9. Extractions from Microorganisms
3.9.1. Bacterial Material Extraction: The Isolation and Identification of Antibiotic Drug Candidates
3.9.2. Fungal Material Extraction: The Isolation and Identification of Drug Candidates from Fungi
3.10. Advances in Compound Identification and Structural Characterization
References

Chapter 4. Isolation of Lead Compounds
4.1. Herbal Remedies
4.2. Plant-Origin Bioactive Ingredients
4.2.1. Atropine
4.2.2. Andrographolide
4.2.3. Bacosides
4.2.4. Caffeine
4.2.5. Camphor
4.2.6. Capsaicin
4.2.7. Colchicine
4.2.8. Curcumin
4.2.9. DIGOXIN
4.2.10. Diosgenin
4.2.11. Emetine
4.2.12. Ergometrine
4.2.13. Eugenol
4.2.14. Gingerols and Shogaols
4.2.15. Glycyrrhetinic Acid
4.2.16. Guggulsterone
4.2.17. Hesperidin
4.2.18. Levodopa
4.2.19. Menthol
4.2.20. Nicotine
4.2.21. Opium Alkaloids
4.2.22. Piperine.
4.2.23. Podophyllotoxi
4.2.24. Quinine and Quinidine
4.2.25. Reserpine
4.2.26. Sennosides
4.2.27. Solasodine
4.2.28. Strychnine and Brucine
4.2.29. Vasicine
4.2.30. Vinca Alkaloids
References

Chapter 5. Management of Experimental Animals
5.1. Animal Experimentation
5.2. Selection of Laboratory Animals
5.3. Most Frequently Used Laboratory Animals
5.3.1. Rodents
5.4. Caring of Laboratory Animals
5.4.1. Housing
5.4.2. Bedding
5.4.3. Feeding
5.4.4. Water
5.4.5. Sanitation
5.4.6. Vermin Control
References

Chapter 6. Acute Toxicity Study and Dose Determination
6.1. Acute Toxicity Study
6.2. Hippocratic Analysis
6.3. Lethal Dose Fifty (LD50) Calculation
6.4. Selection of Test Dose/Concentration
6.5. Conversion of Human Dose to Experimental Animal Dose
6.5.1. Human Equivalent Dose (HED) Calculation Based on Body Surface Area
6.6. Drug, Dose and Route of Administration for Chicken/Poultry
6.7. Test Dose/Concentration Related Ques
References

Chapter 7. Administration Route Selection
7.1. Route of Administration and Its Impacts on Drug Screening
7.2. Different Routes of Drug Administration
7.2.1. Oral Route
7.2.2. Intraperitoneal and Intravenous Routes
7.2.3. Topical Route
7.2.4. Subcutaneous Route
7.2.5. Intramuscular Route
7.2.6. Intrathecal Route
7.2.6. Intranasal and Intratracheal Routes
References

Chapter 8. Nanotechnology: The Interaction of Metal Ions with Extracts
8.1 Nanosynthesis from Medicinal Plants
8.2. Why Do You Use Plant Extracts to Synthesize NPs?
8.3. Examples of Some Nanoparticle Synthesis Methods
8.3.1. AgNPs
8.3.2. AuNPs
8.3.3. CuNPs
8.3.4. PtNPs
8.3.5. PdNPs
8.3.6. NiNPs
8.3.7. FeNPs
8.3.8. Ag-PdNPs
8.3.9. Fe-PdNPs
8.4. Limitations and Challenges of Medicinal Plant-Based Nanoparticle Synthesis
References

Chapter 9. Biological Activity Studies
9.1. Biological Activity Study
9.2. The Importance of Biological Activity Screening

Part I: Antioxidant Tests
In Vitro Models
Ex Vivo Assay(s)
In Vivo Assays

Part II: Anti-Inflammatory Tests
In Vitro Assays
In Vivo Assays

Part III: Immunomodulatory Activity Tests
Human Cell Model
Monkey Cell Model

Part IV: Anti-Arthritic Activity Tests
In Vitro Assays
In Vivo Assays

Part V: Hypoglycemic/Anti-Diabetic Test
In Vitro Assay
In Vivo Assays

Part VI: Anti-Obesity Test
In Vitro Models
In Vivo Model

Part VII: Anti-Hemolytic Tests
Preparation of Rat Erythrocytes
CuOOH-Induced Hemolysis Assay
H2O2-Induced Hemolysis Assay (Model 1)
H2O2-Induced Hemolysis Assay (Model 2)

Part VIII: Thrombolytic Activity Analysis
Clot Lysis Model

Part IX: Anti-Emetic Activity Study
Chick (In Vivo) Model

Part X: Anti-Pyretic Activity Study
Brewer’s Yeast Induced Pyrexia (Mouse/Rat Model)
Yeast Induced Pyrexia (Chick Model)

Part XI: Anti-Microbial Assays
Factors Affecting Anti-Microbial Activity of Test Sample
Agar Disk-Diffusion (Simple) Method
Latin Square (agar well diffusion) Method
Turbidimetric or Serial Dilution (broth dilution) Method
Challenge Test
Some Other Popularly Used Anti-Microbial Assays
Microbiological Assay of Vitamin
Anti-Viral Test

Part XII: Anthelmintic Tests
In Vitro Assay
In Vivo Assay

Part XIII: Anti-Diarrheal Activity Tests

Part XIV: Anti-Cancer Activity Test
Cell Viability Model.
DMBA-Induced Breast Cancer Test
Part XV: Spasmolytic Activity Tests
Isolated Tissue Model
Ach-Induced Contraction Model
Kcl-Induced Contraction Model
Nitric Oxide Pathway Model
Rat Ilium Model
Part XVI: An Evaluation of Biological Effects on Specific Organs
Neuroprotective Activity Tests
Anti-Nociceptive Activity Study
Memory-Enhancing Capacity Test
Anti-Convulsant Activity Tests
Anti-Epileptic Activity Tests
Depression Behavior Models
Anti-Compulsive Model
Motor Coordination Model
Other Popularly Used Neruopharmacological Test Models
Amphetamine-Induced Stereotyped Behavior Study
Cardio-Protective and Cardiological Activity Studies
Anti-Hypertensive Test
Hepatoprotective Activity Tests
Renal (Kidney) Protective Activity Tests
Part XVII: Miscellaneous Activity Studies
Hair Growth Promoting Activity Test
Reproductive Capacity Test
Insect Repellent Test
Mosquito Repellent Tests
Adulticidal Activity Test
Anti-Vitiligo Activity Test
Diuretic/Anti-Diuretic Activity Test
References

Chapter 10. Biopharmaceutical Studies
10.1. Absorption Assays
10.1.1. In Vitro Permeation Test (Franz Diffusion Model)
10.1.2. Percutaneous Absorption Test (Multiple Membrane Model)
10.2. Protein Binding Assays
10.2.1. Filter Paper Assay
10.3. Drug Metabolism Assays
10.3.1. In Vitro Metabolism Test
10.3.2. Food Interference Test
10.4. Drug Excretion Assays
10.4.1. Urinary Excretion Test (Rat Model)
10.5. Assay of Pharmacokinetic Parameters
10.6. Bioavailability Assay
10.6.1. Plasma Sampling Model (Mouse Model)
References

Chapter 11. Toxicity Analysis
11.1. Toxicity Analysis
11.1.1. Acute and Sub-Acute Toxicity Study (Animal Model)
11.2. Cytotoxicity Analysis
11.2.1. Allium cepa Test
11.2.2. Hemolysis Test (rat erythrocyte model)
11.2.3. MTT [3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] Assay
11.3. Genotoxicity Analysis
11.3.1. Plant Model
11.3.2. Animal Models
11.4. Mutagenicity Analysis
References

Chapter 12. Computer-Aided Drug Design (In Silico Study)
12.1. Computational Drug Analysis
12.2. Structure-Based Drug Design
12.3. Ligand-Based Drug Design
12.4. The Total Process of Structure-Based Drug Design
12.4.1. Target Identification
12.4.2. Protein (Target) Structure Modeling, Energy Minimization and Validation
12.4.3. Binding Site Prediction and Analysis
12.4.4. Ligand Selection
12.4.5. Molecular Docking, Virtual Screening and Visualization
12.4.6. Molecular Dynamics (MD) Simulation
12.4.7. Molecular Mechanics/Poisson-Boltzmann Surface Area (MM-PBSA) Calculations
12.4.8. ADMET Analysis
12.4.9. Network Pharmacology
References

Chapter 13. Reagent Preparation
13.1. Reagents
13.2. The Importance of Reagents in Analysis
13.3. Chemicals
13.4. The Importance of Chemicals in Analysis
13.5. The Difference between Chemicals and Reagents
13.6. Reagent Preparation
References

Index

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