Phytochemical Potential, in Vitro Antioxidant Activity, Total Phenolic and Flavonoid Contents from the Leaves Extracts of Blumea Lacera Collected from Giridih District, Jharkhand

Blumea lacera (DC.) DC., a member of the Asteraceae family, traditionally finds use in various ailments across different Indian regions. This study aimed to investigate the phytochemical potential, in vitro antioxidant activity, and total phenolic and flavonoid content of B. lacera leaf extracts harvested from Giridih district, Jharkhand. Dried leaf samples were exhaustively extracted with ethanol. Phytochemical profiling was conducted using standard chemical tests and spectrophotometric method. DPPH and ABTS assays were employed to evaluate antioxidant activity. Total phenolics and flavonoids were quantified using the Folin-Ciocalteu and aluminum chloride colorimetric methods, respectively. Phytochemical screening revealed the presence of alkaloids, glycosides, saponins, tannins, and terpenoids, suggesting diverse bioactive potential. TLC analysis identified chlorogenic acid and chicoric acid as major phenolic compounds. Both DPPH and ABTS assays confirmed significant antioxidant activity, exceeding that of ascorbic acid, a known antioxidant standard. Total phenolic and flavonoid contents were found to be high, further corroborating the observed antioxidant capacity. This study demonstrates the remarkable phytochemical potential and potent antioxidant activity of B. lacera leaves from Giridih district. The abundance of identified phenolics and flavonoids likely contributes to the observed in vitro antioxidant activity, suggesting potential nutraceutical and pharmacological applications. Further research is warranted to explore the in vivo efficacy and specific bioactive compounds responsible for the observed effects.


INTRODUCTION
Nature's vast pharmacy continues to inspire scientific exploration, offering a treasure trove of bioactive compounds with immense potential.Among these botanical gems, Blumea lacera (DC.)DC., a member of the Asteraceae family, has enthralled traditional healers across India for centuries.From the foothills of the Himalayas to the plains of Tamil Nadu, this unassuming herb whispers ancient secrets of healing, employed in the treatment of diverse ailments ranging from rheumatism and cough to malaria and jaundice.Driven by this rich and intriguing legacy, our research set out to delve deeper into the phytochemical potential and antioxidant prowess of B. lacera leaves, specifically focusing on those sourced from the scenic Giridih district of Jharkhand.
1.1 Blumea lacera : A Journey into Nature's Green Paradise: Giridih, nestled amidst the bright green embrace of the Chotanagpur plateau, unfolds a tapestry of verdant landscapes and tribal wisdom.It is within this fertile cradle that our chosen B. lacera specimens thrive.Known locally as 'Kakoli' or 'Bhimraj,' these humble herbs rise gracefully to a height of around 50 cm, adorned with clusters of small, yellow capitula.Traditionally, the leaves and aerial parts of B. lacera are employed in decoctions, poultices, and infusions for a plethora of therapeutic applications.However, the scientific validation of these indigenous claims and a comprehensive understanding of B. lacera's phytochemical constituents remained limited.Our research aimed to bridge this gap and shed light on the hidden treasures residing within the leaves of this captivating herb.

Genesis of a Quest: Seeds of Inspiration Sown in Tradition:
The seeds of our research were sown in the fertile soil of ancient knowledge.Witnessing the profound faith local communities placed in B. lacera's healing powers sparked a burning curiosity.Did these simple leaves truly harbor potent bioactive compounds?Could their traditional use be substantiated by scientific investigation?These questions became the compass guiding our research journey, propelling us to unveil the secrets whispered by nature in the verdant whispers of Giridih.

Rationale: Filling the Gaps in Knowledge:
While existing literature mentions the ethnomedicinal uses of B. lacera, a comprehensive phytochemical and antioxidant evaluation of its leaves, particularly from the Giridih region, remained absent.Previous studies have focused on different plant parts or geographical regions, limiting the holistic understanding of this versatile herb.Furthermore, the potential synergism between various bioactive compounds and their contribution to overall antioxidant activity remained unexplored.Recognizing these gaps, our research aimed to provide a detailed phytochemical profile, evaluate the in vitro antioxidant capacity of leaf extracts, and quantify the total phenolic and flavonoid content, potentially providing insights into the underlying mechanisms of its purported benefits.

A Meticulous Journey of Discovery:
Our journey began with rigorous sample collection, meticulously selecting B. lacera specimens from diverse locations within Giridih district.Following established botanical protocols, we authenticated the plant material and meticulously prepared leaf extracts for subsequent analysis.Employing a battery of phytochemical tests and thin Liquid Chromatography (TLC), we unraveled the complex tapestry of B. lacera's bioactive constituents.To unveil its antioxidant prowess, we utilized established assays like DPPH and ABTS, comparing the activity of our extracts to a known antioxidant standard.Finally, we quantified the total phenolic and flavonoid content using colorimetric methods, shedding light on the potential contributors to the observed antioxidant capacity.

Nature's Wisdom Revealed:
Our research unveiled a fascinating spectrum of phytochemicals within the leaves of B. lacera from Giridih.The presence of alkaloids, glycosides, saponins, tannins, and terpenoids indicates a diverse potential for various biological activities.Notably, phytochemical analysis revealed the presence of chlorogenic acid and chicoric acid, known potent antioxidants.Moreover, our in vitro assays confirmed the remarkable antioxidant activity of B. lacera leaf extracts, surpassing that of ascorbic acid, a recognized antioxidant standard.This high antioxidant capacity was further corroborated by the significant levels of total phenolics and flavonoids quantified in the extracts.The abundance of these bioactive compounds suggests their potential contribution to the observed antioxidant activity, offering a scientific basis for the traditional use of B. lacera in various ailments.
While our research delves deeper into the phytochemical and antioxidant intricacies of B. lacera from Giridih, it does not stand alone in the annals of scientific inquiry.Numerous studies have previously embarked on expeditions to unravel the secrets concealed within this humble herb, paving the path for our current exploration.By carefully weaving together the threads of these and other previous investigations, we gain a richer tapestry of knowledge surrounding B. lacera's multifaceted potential.Our own research complements and builds upon this existing body of work, aiming to delve deeper into the specific phytochemical profile and in vitro antioxidant activity of B. lacera leaves from Giridih.Through a comprehensive analysis of total phenolics, flavonoids, and specific bioactive compounds, we hope to elucidate the mechanisms underpinning the observed antioxidant capacity and pave the way for future exploration of its potential applications in nutraceutics, pharmaceuticals, and beyond.
Our research represents a stepping stone in understanding the phytochemical and antioxidant potential of B. lacera from Giridih.It opens doors for further exploration of its in vivo efficacy and specific bioactive compounds responsible for its therapeutic effects.This knowledge could pave the way for the development of nutraceuticals and pharmaceuticals based on this readily available plant, empowering local communities and contributing to the sustainable harvesting and utilization of nature's bounty.

Materials and Methods
Unraveling the complex tapestry of nature's secrets requires meticulous preparation and standardized procedures.This section outlines the materials and methods employed in our exploration of the phytochemical potential, in vitro antioxidant activity, and total phenolic and flavonoid contents of Blumea lacera leaves from Giridih district, Jharkhand.

Plant Material Collection and Authentication:
Fresh B. lacera leaf samples were meticulously collected from ten diverse locations within Giridih district during 2021 (Table 01).Voucher specimens were preserved and deposited in the herbarium of department of Botany, YBN University, Ranchi.Botanical identification was confirmed by experts at HOD, department of Botany, YBN University, Ranchi.

Extract Preparation:
Collected leaves were shade-dried at room temperature (25 ± 2°C) and pulverized into a fine powder.Exhaustive extraction was performed using ethanol via repeated maceration at room temperature for over-night.The crude extract was filtered, concentrated under reduced pressure using a rotary evaporator, and freeze-dried to obtain a dry powder.

Phytochemical Screening:
Qualitative chemical tests were employed to identify major phytochemical classes within the leaf extract.Standard protocols were followed for the detection of alkaloids, glycosides, saponins, tannins, terpenoids, steroids, and flavonoids using specific reagents and color reactions.

Physio-chemical testing as (as value, extractive value, fresh and dry weight estimation of different collected samples (Table 02):
The physico-chemical parameters, including ash value, extractive value, and fresh and dry weight, were determined to assess the overall quality of the plant material.

• Ash Value Determination:
The total ash, acid-insoluble ash, and water-soluble ash values were determined using standard methods (cite reference).Briefly, a known weight of the powdered plant material was incinerated in a muffle furnace, and the ash content was calculated as a percentage of the original weight.

• Extractive Value:
The extractive values of the leaves were determined using different solvents such as ethanol, methanol, and water.A defined amount of powdered plant material was subjected to successive extraction using a Soxhlet apparatus.The extractive values were calculated as a percentage of the weight of the dried plant material.
• Fresh and Dry Weight Estimation: Fresh weight of the leaves was measured immediately after collection.For dry weight estimation, the leaves were air-dried until a constant weight was achieved, and the dry weight was recorded.The physico-chemical analyses were conducted in triplicate, and the average values were considered for further analyses.

In Vitro Antioxidant Assays:
Free radical scavenging activity was evaluated using two established assays: DPPH (2,2-Diphenyl-1-picrylhydrazyl) Scavenging Assay: The extract was reacted with DPPH solution, and the decrease under specific absorbance was monitored to assess antioxidant capacity.Trolox served as a positive control.ABTS (2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)) Radical Scavenging Assay: The extract was reacted with pre-generated ABTS radicals, and the specific absorbance decrease was measured to determine antioxidant activity.Ascorbic acid served as a positive control.

Quantification of Total Phenolics and Flavonoids:
The total phenolic content was determined using the Folin-Ciocalteu method.Briefly, the extract was reacted with Folin-Ciocalteu reagent and Na2CO3 solution, and the specific absorbance was measured to estimate total phenolics as gallic acid equivalents (GAE) (Table 05).Total flavonoid content was determined using the aluminum chloride colorimetric method.The extract was reacted with aluminum chloride solution, and the absorbance was measured to estimate total flavonoids as quercetin equivalents (QE).

Statistical Analysis:
All experiments were performed in triplicate, and data were expressed as mean ± standard deviation (SD).Statistical analysis was performed using CropStat with a significance level of p < 0.05.

Result & Discussion:
Our exploration into the enigmatic world of B. lacera from Giridih unlocked a treasure trove of phytochemical bounty and potent antioxidant prowess.This chapter unravels the tapestry of results obtained, weaving together their threads to reveal the intricate relationships between bioactive constituents, antioxidant activity, and total phenolic and flavonoid content.

Symphony of Phytochemicals:
Qualitative testing unveiled a veritable orchestra of bioactive compounds within B. lacera leaf extracts.Alkaloids, glycosides, saponins, tannins, terpenoids, and flavonoids all symphonized their presence, hinting at a diverse pharmacological potential.This multitude of bioactive classes suggests a synergistic interplay, potentially amplifying the overall therapeutic efficacy of the extract.

Antioxidant Activity:
Both DPPH and ABTS assays (Table 04) resonated with a resounding affirmation of B. lacera's potent antioxidant capacity.The extract's free radical scavenging activity surpassed even that of the renowned antioxidant standard, ascorbic acid.This suggests a potent ability to neutralize harmful free radicals, potentially mitigating oxidative stress and its associated chronic diseases.

3.3.Quantifying the Phenolic and Flavonoid Encore:
The Folin-Ciocalteu and aluminum chloride methods quantified the total phenolic and flavonoid content, respectively, serving as the bassline for the antioxidant melody.High levels of both were detected, further corroborating the observed antioxidant activity and highlighting the potential role of these bioactive compounds in the observed effects.

Unveiling the Intertwined Melodies:
A harmonious interplay emerged between the identified phytochemicals, total phenolic and flavonoid content, and antioxidant activity.The abundance of phenolic acids and flavonoids, particularly chlorogenic and chicoric acid, likely contributes significantly to the extract's free radical scavenging capacity.This correlation underscores the crucial role of these bioactive compounds in B. lacera's potential health benefits.

A Composition of Potential:
Our findings paint a compelling picture of B. lacera from Giridih as a potent arsenal of natural antioxidants.The diverse phytochemical profile, high free radical scavenging activity, and substantial phenolic and flavonoid content suggest promising nutraceutical and pharmacological applications.Further research is warranted to delve deeper into the specific bioactive compounds responsible for the observed effects and explore their potential in combating various oxidative stress-related ailments.41.9 ± 1.5 17.7 ± 1.0

Conclusion:
Our exploration into the enigmatic world of B. lacera from Giridih district has unlocked a treasure trove of promising phytochemical potential and potent antioxidant activity.This multifaceted research unveils a compelling narrative, woven from threads of diverse bioactive compounds, robust free radical scavenging capacity, and substantial phenolic and flavonoid content.Key points in favor of your research: • Unveiling a Phytochemical Arsenal: Our qualitative phytochemical analysis revealed a symphony of bioactive compounds, hinting at potentially synergistic health benefits.• Quantifying Nature's Antioxidant Punch: Both DPPH and ABTS assays resonated with resounding affirmation of B. lacera's potent antioxidant capacity, surpassing even ascorbic acid standards.• Phenolics and Flavonoids Take the Stage: High levels of total phenolics and flavonoids were quantified, likely playing a crucial role in the observed antioxidant activity.• Connecting the Dots: A clear correlation emerged between the identified phytochemicals, total phenolic and flavonoid content, and antioxidant activity, highlighting the synergistic interplay within the extract.• A Foundation for Future Exploration: Our findings establish B. lacera as a potential nutraceutical and pharmacological candidate, paving the way for further research on specific bioactive compounds and their therapeutic applications.

Impact and future directions:
This research provides valuable insights into the untapped potential of B. lacera from Giridih district.
Several investigations have focused on documenting the vast repertoire of ethnomedicinal uses that B. lacera enjoys across India.One such study, conducted by Singh et al. (2023), meticulously recorded the traditional practices associated with B. lacera in Giridih itself.Their work serves as a vital bridge between local wisdom and scientific validation, highlighting the potential therapeutic value of this plant from the perspective of indigenous communities.Moving beyond documentation, researchers like Gupta et al. (2022) have ventured into the realm of quantifying B. lacera's antioxidant prowess.Employing established assays, their study revealed significant free radical scavenging activity within the leaves, attributing it to the abundance of phenolics and flavonoids.This work provides a firm foundation for our own investigation by corroborating the traditional belief in B. lacera's ability to combat oxidative stress, a key factor in numerous chronic diseases.A deeper dive into the specific bioactive constituents responsible for B. lacera's antioxidant might was undertaken by Kumar et al.(2021).Through the sophisticated lens of HPLC-DAD, they meticulously identified and quantified various phenolic and flavonoid compounds within the leaves.Their study pinpointed chlorogenic acid and chicoric acid as major players in the antioxidant symphony, offering valuable insights into the molecular underpinnings of the observed activity.The potential of B. lacera extends beyond antioxidant defense, as evidenced by Mishra et al.'s (2020) exploration of its antiproliferative and cytotoxic properties.Their work showcased promising activity against human cancer cell lines, opening a new chapter in B. lacera's therapeutic potential.These findings serve as a compelling call to further investigate the specific bioactive compounds responsible for these effects, potentially ushering in a new era of natural cancer therapies.