Isolation and Characterisation of Protoberberines with Antioxidant, Antiproliferative and Protease Inhibitory Activity from Berberis Aristata

Contents

Chapter 1 Introduction and review of literature

1.1 Introduction

1.2 Classification of antioxidants

1.3 Applications of antioxidants

1.4 Source of natural antioxidants

1.4.1 Berberis aristata

1.4.2 Carum carvi

1.4.3 Foeniculum vulgare

1.4.4 Glycyrrhiza glabra

1.4.5 Madhuca indica

1.4.6 Myristica fragrans

1.4.7 Nardostacy’s jatamansi

1.4.8 Swertia chirayita

1.4.9 Trachyspermum ammi

1.4.10 Zingiber officinale

1.5 Protoberberine

Chapter 2 Screening of medicinal plant extracts for antioxidant activity and phytochemicals

2.1 Introduction

2.2 Aim and Objectives

2.3 Materials and Methods

2.3.1 Preparation of plant extracts

2.3.2 DPPH radical scavenging assay

2.3.3 Superoxide radical scavenging assay

2.3.4 Hydroxyl radical scavenging assay

2.3.5 In-vitro inhibition of lipid peroxidation

2.3.6 Total antioxidant assay by FRAP method

2.3.7 Screening of medicinal plant extracts for phytochemicals

2.3.7.1 Tests for alkaloids

2.3.7.2 Tests for flavonoids and polyphenolics

2.3.7.3 Test for saponins

2.3.7.4 Test for tannins

2.4 Statistical analysis

2.5 Results and Discussion

2.6 Conclusion

Chapter 3 Activity guided isolation of antioxidants from B.aristata

3.1 Introduction

3.2 Aim and Objectives

3.3 Materials and Methods

3.3.1 Fractionation of ethanolic extract of B.aristata

3.3.2 Antioxidant activity

3.3.3 Phytochemical analysis

3.3.3.1 UV-Visible absorption spectral analysis

3.3.3.2 Qualitative analysis

3.3.3.3 Quantification of total alkaloids

3.3.3.4 Quantification of total phenolics

3.3.3.5 Quantification of total flavonoids

3.4 Statistical analysis

3.5 Results and Discussion

3.6 Conclusion

Chapter 4 Antioxidant potential of ethyl acetate fraction against oxidative stress in E.coli model, on erythrocytes and DNA

4.1 Introduction

4.2 Aim and Objectives

4.3 Materials and Methods

4.3.1 Effect of H2 O2 induced oxidative stress in E.coli model

4.3.2 Effect of AAPH radical induced oxidative stress on RBC

4.3.3 DNA fragmentation assay

4.4 Statistical analysis

4.5 Results and Discussion

4.6 Conclusion

Chapter 5 Purification and characterisation of antioxidant alkaloids from B.aristata

5.1 Introduction

5.2 Aim and Objectives

5.3 Materials and Methods

5.3.1 Purification using silica gel column chromatography

5.3.2 High performance liquid chromatography

5.3.3 Structure prediction

5.3.3.1 UV-Visible absorption spectral analysis

5.3.3.2 IR spectroscopy

5.3.3.3 Nuclear magnetic resonance spectroscopy

5.4 Results and Discussion

5.5 Conclusion

Chapter 6 In silico docking studies of berberrubine, jatrorrhizine and thalifendine with caspase 3, cathepsin B, MMP-9 and telomeric DNA

6.1 Introduction

6.2 Aim and Objectives

6.3 Materials and Methods

6.3.1 Retrieval of receptors from Protein DataBank (PDB)

6.3.2 Designing of ligands

6.3.3 Docking with hex v6.3

6.3.4 Docking with iGEMDOCK v2.1

6.4 Results and Discussion

6.5 Conclusion

Chapter 7 Antiproliferative and protease inhibitory activities of protoberberines

7.1 Introduction

7.2 Aim and Objectives

7.3 Materials and Methods

7.3.1 Antiradical activity of berberrubine, jatrorrhizine and thalifendine

7.3.2 Cell proliferation assay

7.3.3 In vitro inhibition of cathepsin B enzyme activity

7.3.4 In vitro inhibition of MMP-9 enzyme activity

7.4 Statistical analysis

7.5 Results and Discussion

7.6 Conclusion

Bibliography

Research Objectives and Focus

The research primarily aims to identify and isolate potent antioxidant compounds from the medicinal plant Berberis aristata, evaluate their biological activity, and investigate their therapeutic potential against oxidative stress and cancer through in vitro and in silico methods.

• Screening of various medicinal plants for antioxidant properties using diverse free radical scavenging assays.
• Activity-guided isolation and structural characterization of active alkaloids (protoberberines) from B. aristata.
• Assessment of the protective role of isolated compounds against oxidative stress in cellular and DNA models.
• In silico molecular docking studies to predict the interaction of isolated protoberberines with cancer-regulating target proteins.
• Investigation of the antiproliferative and protease inhibitory activities of the purified compounds against cancer cell lines.

Excerpt from the Book

Summary of Chapters

Chapter 1: Provides an introduction and literature review regarding antioxidants, their classification, and the therapeutic applications of various medicinal plants.

Chapter 2: Details the screening of various solvent extracts from medicinal plants for their antioxidant potential against different radicals and qualitative phytochemical analysis.

Chapter 3: Describes the activity-guided isolation process of antioxidants from the ethanolic extract of B. aristata using liquid-liquid fractionation.

Chapter 4: Investigates the antioxidant potential of the ethyl acetate fraction of B. aristata against oxidative stress induced in E. coli, red blood cells, and DNA.

Chapter 5: Covers the purification and structural characterization of antioxidant alkaloids (protoberberines) from the ethyl acetate fraction of B. aristata.

Chapter 6: Presents in silico docking studies to analyze the binding affinity of isolated protoberberines with cancer-relevant targets like caspases, MMP-9, and telomeric DNA.

Chapter 7: Analyzes the antiproliferative and protease inhibitory activities of the purified protoberberines against specific human cancer cell lines.

Keywords

Berberis aristata, antioxidants, protoberberines, oxidative stress, DPPH, lipid peroxidation, DNA fragmentation, docking, caspase 3, MMP-9, cathepsin B, antiproliferative, cancer, alkaloids, medicinal plants.

Frequently Asked Questions

What is the core focus of this research?

The research is dedicated to the study of antioxidant, antiproliferative, and protease inhibitory effects of protoberberine alkaloids derived from the plant Berberis aristata.

What are the primary themes explored?

Key themes include plant-based antioxidant screening, isolation and purification of bioactive compounds, structural characterization via spectroscopy, and the investigation of anti-cancer properties through molecular docking and cell culture assays.

What is the ultimate objective of this thesis?

The primary goal is to validate the traditional medicinal use of B. aristata by identifying its active components and establishing their efficacy as therapeutic agents against oxidative stress and cancer.

Which scientific methods are employed?

The study uses liquid-liquid fractionation, silica gel column chromatography, HPLC, UV-Visible, IR and NMR spectroscopy, in vitro radical scavenging assays, MTT proliferation assays, and in silico molecular docking software like Hex v6.3 and iGEMDOCK.

What is covered in the main section of the work?

The main sections cover the screening of various medicinal plants, activity-guided isolation, the protective effect of B. aristata extracts against oxidative stress, chemical purification, computational docking studies, and final efficacy testing in cancer cell lines.

Which keywords best describe this study?

Key terms include Berberis aristata, antioxidants, protoberberines, oxidative stress, docking, protease inhibition, and cancer treatment.

How does the ethyl acetate fraction contribute to the study's findings?

The ethyl acetate fraction was found to be a rich source of antioxidant alkaloids, exhibiting the highest radical scavenging and lipid peroxidation inhibition activity compared to other solvents.

What conclusions were drawn regarding the protoberberines?

The study concluded that berberrubine, jatrorrhizine, and thalifendine exhibit significant antioxidant and anti-cancer properties, with binding potential comparable to existing drugs like doxorubicin.