Optimizing UHPLC: Chromatographic Conditions for Plant Extract Profiling

1. Significance of Plant Extract Profiling

1. Significance of Plant Extract Profiling

Plant extract profiling is a critical analytical technique that plays a pivotal role in the characterization and evaluation of natural products derived from plants. This process is essential for various applications, including quality control, pharmacological research, and the discovery of new bioactive compounds. The significance of plant extract profiling can be highlighted in the following aspects:

Quality Control and Standardization:
Plant extracts are used extensively in the pharmaceutical, food, and cosmetic industries. Profiling these extracts ensures that the final products meet the required quality standards and are consistent in their composition, which is crucial for safety and efficacy.

Pharmacological Research:
Understanding the chemical composition of plant extracts is fundamental for pharmacological studies. It helps in identifying the active constituents responsible for therapeutic effects and in elucidating the mechanisms of action.

Discovery of Bioactive Compounds:
Profiling can lead to the discovery of novel bioactive compounds with potential applications in medicine, agriculture, and other fields. It aids in the development of new drugs and the improvement of existing ones.

Biodiversity Assessment:
Plant extract profiling contributes to the assessment of biodiversity by providing insights into the chemical diversity of plant species. This information is vital for conservation efforts and the sustainable use of plant resources.

Authentication of Plant Materials:
Profiling techniques can be used to authenticate plant materials and verify their botanical identity, which is particularly important in the trade of herbal products.

Environmental Monitoring:
Plant extracts can serve as bioindicators of environmental conditions. Changes in their chemical profiles may reflect the impact of pollutants or other environmental stressors on plant health.

Nutritional Analysis:
For food products, plant extract profiling can provide information on the nutritional content, helping to ensure that consumers receive the claimed benefits.

Regulatory Compliance:
Regulatory agencies require detailed information on the composition of plant extracts to ensure compliance with safety and efficacy standards. Profiling helps manufacturers meet these regulatory requirements.

In summary, plant extract profiling is a multifaceted discipline that underpins the safe and effective use of plant-derived products. As analytical techniques, such as ultra-high-performance liquid chromatography (UHPLC), continue to advance, the depth and accuracy of profiling will further enhance our ability to harness the potential of plants for various applications.

2. Method Development for UHPLC in Plant Extract Analysis

2. Method Development for UHPLC in Plant Extract Analysis

The method development for Ultra High Pressure Liquid Chromatography (UHPLC) in plant extract analysis is a critical step to ensure accurate and reproducible results. This section will delve into the various aspects of developing a UHPLC method tailored for the profiling of crude plant extracts.

2.1 Selection of Stationary Phase
The choice of the stationary phase is crucial for the separation of complex mixtures found in plant extracts. Columns with small particle sizes and high surface area, such as those with sub-2-micron particles, are often used to achieve high resolution and speed. The chemistry of the stationary phase, such as C18, C8, or phenyl-hexyl, should be chosen based on the polarity of the compounds of interest.

2.2 Mobile Phase Optimization
The mobile phase composition is a key factor influencing the separation efficiency. A common approach is to use a gradient elution with a mixture of water and an organic solvent such as acetonitrile or methanol. The gradient profile must be optimized to achieve the desired resolution within a reasonable time frame.

2.3 Flow Rate Determination
UHPLC allows for higher flow rates compared to traditional HPLC, which can significantly reduce analysis time. However, the flow rate must be carefully determined to balance speed with separation efficiency and system back pressure.

2.4 Temperature Control
Column temperature can affect both the viscosity of the mobile phase and the solubility of the analytes. Maintaining a consistent column temperature is essential for reproducibility. The optimal temperature may vary depending on the specific plant compounds being analyzed.

2.5 Injection Volume
The injection volume in UHPLC is typically smaller than in HPLC due to the smaller column diameters. The choice of injection volume can impact both sensitivity and overloading of the column. It is important to find a balance that provides adequate sensitivity without compromising separation.

2.6 Detection Method Integration
The choice of detection method, such as UV-Vis, fluorescence, or mass spectrometry, depends on the properties of the compounds in the plant extract. The method must be sensitive and selective for the target compounds, and it should be integrated into the UHPLC system to ensure compatibility and data quality.

2.7 Method Validation
Once the initial method is developed, it must be rigorously validated to ensure it meets the required performance criteria. Validation parameters include linearity, accuracy, precision, specificity, detection and quantification limits, and robustness.

2.8 Data Processing and Analysis
The development of a robust data processing and analysis pipeline is essential for the interpretation of UHPLC data. This includes peak identification, alignment, quantification, and multivariate analysis for pattern recognition and comparison of different plant extracts.

2.9 Standardization and Quality Control
The establishment of standard operating procedures (SOPs) and the use of quality control samples are vital for ensuring the reliability and reproducibility of UHPLC methods in plant extract profiling.

2.10 Adaptation to Specific Plant Extracts
While a general method may be developed, it is often necessary to adapt the UHPLC method to the specific characteristics of different plant extracts. This may involve adjustments to the mobile phase, gradient, or other parameters to optimize the separation of the unique compounds present in each extract.

The method development process for UHPLC in plant extract analysis is iterative and requires a deep understanding of both the chromatographic system and the chemical properties of the plant compounds. With careful optimization, UHPLC can provide a powerful tool for the detailed profiling of crude plant extracts, enabling the discovery of novel bioactive compounds and the quality control of herbal products.

3. Sample Preparation Techniques for Crude Plant Extracts

3. Sample Preparation Techniques for Crude Plant Extracts

Sample preparation is a critical step in the analysis of crude plant extracts using ultra-high-performance liquid chromatography (UHPLC). The quality of the sample preparation process directly affects the accuracy and reliability of the results obtained. Here are some commonly used sample preparation techniques for crude plant extracts:

1. Solvent Extraction: This is the most common method for extracting bioactive compounds from plant materials. Various solvents such as methanol, ethanol, water, or a mixture of these can be used depending on the polarity of the compounds of interest.

2. Ultrasonic-Assisted Extraction (UAE): This technique uses ultrasonic waves to enhance the extraction process by breaking cell walls and increasing the solvent penetration into the plant material, which can lead to higher extraction efficiency.

3. Pressurized Liquid Extraction (PLE): Also known as accelerated solvent extraction, this method uses high pressure and temperature to extract compounds more rapidly and efficiently than traditional solvent extraction.

4. Supercritical Fluid Extraction (SFE): This technique employs supercritical fluids, typically carbon dioxide, to extract compounds. It offers advantages such as high extraction efficiency, low solvent residue, and the ability to selectively extract compounds based on their solubility in the supercritical fluid.

5. Solid-Phase Extraction (SPE): SPE is used to isolate specific compounds from a complex mixture. It involves passing the extract through a column packed with a solid phase that selectively retains certain compounds while allowing others to pass through.

6. Solid-Phase Microextraction (SPME): SPME is a solvent-free extraction technique where a fiber coated with a specific stationary phase is exposed to the headspace above the sample, allowing the analytes to be absorbed or adsorbed onto the fiber.

7. QuEChERS (Quick, Easy, Cheap, Effective, Rugged, and Safe): This method is used for the extraction of pesticide residues from plant materials and involves a two-step process of extraction and cleanup.

8. Dilution and Filtration: In some cases, crude extracts may be simply diluted with a suitable solvent and then filtered to remove particulate matter before injection into the UHPLC system.

9. Derivatization: Certain compounds may require chemical modification to improve their detection or chromatographic properties. Derivatization can involve the addition of functional groups to make the compounds more amenable to analysis.

10. Sample Drying: After extraction, the sample may need to be dried to remove the solvent, often using techniques such as rotary evaporation, lyophilization, or nitrogen吹流干燥.

Each of these techniques has its advantages and limitations and the choice of method depends on the nature of the plant material, the compounds of interest, and the specific requirements of the analysis. Proper sample preparation is essential to ensure that the UHPLC analysis provides meaningful and reproducible results.

4. Chromatographic Conditions and Optimization

4. Chromatographic Conditions and Optimization

In ultra high pressure liquid chromatography (UHPLC) for crude plant extract profiling, the optimization of chromatographic conditions is crucial for achieving high resolution, sensitivity, and reproducibility. This section will discuss the various factors that need to be considered and optimized to ensure accurate and reliable results.

4.1 Selection of Stationary Phase

The choice of the stationary phase, or the column, is one of the most important factors in UHPLC. Columns with different particle sizes, lengths, and chemistries are available, and the selection depends on the complexity of the plant extract and the target compounds. Smaller particle sizes (e.g., 1.7 µm) can provide higher resolution and faster analysis times, but may require higher pressures.

4.2 Mobile Phase Composition

The mobile phase, typically a mixture of water and an organic solvent such as acetonitrile or methanol, plays a critical role in the separation process. The composition of the mobile phase affects the retention time and peak shape of the compounds. Gradient elution is commonly used to separate a wide range of compounds with different polarities.

4.3 Flow Rate

The flow rate in UHPLC is significantly higher than in traditional HPLC, which contributes to the reduced analysis time. However, the flow rate must be optimized to ensure that the column efficiency is not compromised. Too high a flow rate can lead to peak broadening and reduced resolution.

4.4 Temperature Control

Temperature can significantly affect the retention times and peak shapes in chromatography. Maintaining a consistent column temperature is essential for reproducibility. Some studies have shown that higher temperatures can improve the separation of certain compounds, particularly for thermolabile compounds.

4.5 Gradient Elution Optimization

Gradient elution is a technique where the composition of the mobile phase is gradually changed during the run. The optimization of the gradient involves adjusting the start and end times, the slope, and the percentage of organic solvent. The goal is to achieve the best separation of the compounds within the shortest possible time.

4.6 Injection Volume

The injection volume can affect the sensitivity and peak shape. Smaller injection volumes can lead to better peak shapes and higher sensitivity, but may also require more concentrated samples. It is important to find a balance that provides the best analytical performance.

4.7 System Pressure and Compatibility

UHPLC systems operate at much higher pressures than traditional HPLC systems. It is essential to ensure that all components of the system, including the column, pumps, and detectors, are compatible with the high pressures. Regular maintenance and monitoring of system pressure can help prevent damage to the equipment.

4.8 Method Validation

After optimizing the chromatographic conditions, it is necessary to validate the method to ensure its reliability and reproducibility. Validation parameters may include accuracy, precision, specificity, detection and quantification limits, and robustness.

4.9 Use of Advanced Software

Modern UHPLC systems are often equipped with advanced software that can assist in method development and optimization. These tools can help in selecting the best column, predicting retention times, and optimizing gradient conditions.

By carefully considering and optimizing these chromatographic conditions, researchers can achieve high-quality profiling of crude plant extracts using UHPLC, leading to more accurate identification and quantification of bioactive compounds.

5. Detection and Quantification Methods

5. Detection and Quantification Methods

Ultra-high-performance liquid chromatography (UHPLC) is a powerful analytical technique that, when coupled with appropriate detection and quantification methods, can provide accurate and reliable results for the profiling of crude plant extracts. The choice of detection method is crucial for the identification and quantification of the various compounds present in plant extracts, which can range from simple molecules like flavonoids to complex macromolecules such as proteins and polysaccharides.

5.1 UV-Vis Detection
Ultraviolet-visible (UV-Vis) detection is one of the most common detection methods used in UHPLC due to its simplicity, sensitivity, and compatibility with a wide range of compounds. It operates by measuring the absorption of UV or visible light by the analytes in the eluent. This method is particularly useful for compounds with chromophores, such as flavonoids and anthocyanins.

5.2 Mass Spectrometry (MS)
Mass spectrometry is a highly sensitive and selective detection technique that can provide structural information about the compounds in a mixture. It is particularly useful for the identification and quantification of unknown compounds or when dealing with complex mixtures. Tandem mass spectrometry (MS/MS) can further enhance selectivity by fragmenting the analytes and analyzing the resulting fragments.

5.3 Fluorescence Detection
Fluorescence detection is another sensitive method that is particularly useful for compounds that exhibit native fluorescence, such as certain alkaloids and terpenes. It operates by measuring the light emitted by molecules after they have absorbed light at a specific wavelength.

5.4 Evaporative Light Scattering Detection (ELSD)
Evaporative light scattering detection is a universal detection method that does not rely on the optical properties of the analytes. It is particularly useful for non-UV absorbing compounds and can be used for the detection of lipids, carbohydrates, and other non-chromophoric compounds.

5.5 Refractive Index Detection (RID)
Refractive index detection is a non-destructive method that measures the change in the refractive index of the eluent caused by the presence of the analytes. It is less sensitive than other methods but can be useful for the quantification of certain compounds in complex mixtures.

5.6 Quantification Methods
Quantification in UHPLC is typically achieved using external calibration, internal standard, or standard addition methods. The choice of method depends on the complexity of the sample, the availability of pure standards, and the required accuracy and precision.

- External Calibration involves the preparation of a series of standards and the construction of a calibration curve. The concentration of the analytes in the sample is then determined by comparing their response to that of the standards.
- Internal Standard is a method where a known amount of an internal standard, which is similar in chemical properties to the analytes but not present in the sample, is added to each sample and standard. This helps to correct for variations in sample preparation and injection volume.
- Standard Addition involves the addition of known amounts of the analytes to the sample, which allows for the construction of a calibration curve within the sample matrix itself, thus accounting for matrix effects.

5.7 Data Processing and Analysis
The data obtained from the UHPLC system is processed using specialized software that allows for peak identification, integration, and quantification. Advanced software can also perform deconvolution of overlapping peaks, peak purity assessment, and multivariate data analysis to provide a comprehensive profile of the plant extract.

In conclusion, the choice of detection and quantification methods in UHPLC is critical for the accurate profiling of crude plant extracts. The combination of UHPLC with various detection techniques provides a powerful tool for the identification, quantification, and characterization of a wide range of compounds in plant extracts, contributing significantly to the fields of phytochemistry, drug discovery, and quality control of herbal products.

6. Applications of UHPLC in Plant Extract Profiling

6. Applications of UHPLC in Plant Extract Profiling

Ultra High Pressure Liquid Chromatography (UHPLC) has revolutionized the profiling of crude plant extracts due to its high resolution, sensitivity, and speed. This section will explore the various applications of UHPLC in the field of plant extract profiling, highlighting its versatility and effectiveness in different scenarios.

Phytochemical Analysis:
UHPLC is extensively used for the identification and quantification of phytochemicals in plant extracts. These compounds, which include alkaloids, flavonoids, terpenoids, and phenolic compounds, are crucial for understanding the therapeutic properties of plants.

Quality Control:
In the pharmaceutical and nutraceutical industries, UHPLC serves as a robust tool for quality control, ensuring that plant-based products meet the required standards. It helps in monitoring the consistency of active ingredients across different batches of products.

Metabolite Profiling:
UHPLC is employed in metabolomics studies to analyze the metabolic profile of plant extracts. This approach is valuable for understanding the biochemical pathways in plants and their responses to various stimuli or treatments.

Fingerprinting:
The technique is used to create chemical fingerprints of plant extracts, which are essential for the authentication and standardization of herbal products. Fingerprinting helps in distinguishing between different plant species and their parts, ensuring the correct use of plant materials.

Purity Assessment:
UHPLC is instrumental in assessing the purity of plant extracts, which is critical for safety and efficacy. It can detect the presence of contaminants or adulterants that may compromise the quality of the final product.

Stability Studies:
The stability of plant extracts can be evaluated using UHPLC to determine how they degrade over time or under different storage conditions. This information is vital for setting appropriate shelf life and storage guidelines.

Comparative Analysis:
UHPLC enables the comparison of different plant extracts or the same plant extract under various conditions. This comparative analysis is crucial for optimizing extraction methods and understanding the impact of different processing techniques on the chemical composition of plant extracts.

Environmental and Ecological Studies:
UHPLC is also applied in environmental studies to monitor the presence of plant compounds in various ecosystems. This can provide insights into the ecological roles of plants and their interactions with other organisms.

Nutritional Analysis:
In the food industry, UHPLC is used to analyze the nutritional content of plant-based foods and beverages, ensuring that they meet the nutritional claims made by manufacturers.

Drug Discovery and Development:
UHPLC plays a significant role in the discovery and development of new drugs from plant sources. It aids in the screening of plant extracts for bioactive compounds with potential therapeutic applications.

Toxicological Studies:
The technique is utilized in toxicological research to identify and quantify toxic compounds in plant extracts, which is essential for assessing their safety for human consumption.

In summary, the applications of UHPLC in plant extract profiling are vast and diverse, making it an indispensable tool in the fields of natural product research, quality control, and drug discovery. Its ability to provide detailed chemical information rapidly and accurately has significantly advanced our understanding of plant chemistry and its implications for health and the environment.

7. Case Studies: Successful Profiling of Specific Plant Extracts

7. Case Studies: Successful Profiling of Specific Plant Extracts

In this section, we delve into the practical applications of UHPLC in profiling specific plant extracts, showcasing its effectiveness in identifying and quantifying bioactive compounds in various plant matrices. These case studies highlight the versatility and efficiency of UHPLC in the field of natural product research.

7.1 Profiling of Ginkgo Biloba Extracts
Ginkgo biloba, known for its cognitive-enhancing properties, has been extensively studied for its flavonoid and terpene lactone content. UHPLC has been successfully employed to profile these bioactive compounds, providing a rapid and sensitive method for quality control and standardization of Ginkgo biloba extracts. The high resolution and speed of UHPLC have enabled the simultaneous detection of multiple biomarkers, including ginkgolides and bilobalide, within a short analysis time.

7.2 Analysis of Green Tea Polyphenols
Green tea is rich in polyphenolic compounds, such as catechins, which have been associated with numerous health benefits. UHPLC has been instrumental in the detailed characterization of these compounds, offering a comprehensive profile of green tea polyphenols. The method has been optimized to separate and quantify catechins like epigallocatechin gallate (EGCG), providing a reliable tool for assessing the quality and authenticity of green tea products.

7.3 Characterization of Curcuminoids in Turmeric Extracts
Turmeric, a popular spice with potent anti-inflammatory properties, contains Curcuminoids as its main bioactive components. UHPLC has been utilized to analyze and quantify Curcumin, demethoxyCurcumin, and bisdemethoxyCurcumin in turmeric extracts. The method has been optimized to overcome the challenges posed by the complex matrix of turmeric, ensuring accurate and reproducible results.

7.4 Profiling of Echinacea Alkaloids
Echinacea species are widely used for their immune-boosting properties, with alkaloids being one of the key bioactive groups. UHPLC has been applied to profile the alkaloid content in Echinacea Extracts, providing a sensitive and efficient method for the analysis of these compounds. The method has been tailored to the specific characteristics of Echinacea alkaloids, allowing for their accurate identification and quantification.

7.5 Analysis of Saponins in Ginseng Extracts
Ginseng is a highly valued medicinal plant known for its adaptogenic effects. Saponins, the major bioactive compounds in ginseng, have been profiled using UHPLC. The method has been optimized to separate and quantify various saponins, such as ginsenosides, providing a robust analytical tool for the quality assessment of ginseng extracts.

7.6 Conclusion
These case studies demonstrate the power of UHPLC in profiling a diverse range of plant extracts, from the cognitive-enhancing compounds in Ginkgo biloba to the immune-boosting saponins in ginseng. The method's high sensitivity, resolution, and speed make it an invaluable tool for the analysis and standardization of plant extracts, paving the way for further research and development in the field of natural products.

8. Challenges and Limitations of UHPLC in Plant Extract Analysis

8. Challenges and Limitations of UHPLC in Plant Extract Analysis

Ultra High Pressure Liquid Chromatography (UHPLC) has revolutionized the field of plant extract profiling with its high efficiency, speed, and sensitivity. However, despite its numerous advantages, there are still challenges and limitations that researchers and analysts need to be aware of when using UHPLC for crude plant extract analysis.

Complexity of Plant Matrices:
One of the primary challenges in plant extract analysis is the inherent complexity of plant matrices. Crude extracts can contain a wide range of compounds, including proteins, polysaccharides, and other biomolecules, which can interfere with the analysis and lead to peak distortion or signal suppression.

Sample Preparation:
Effective sample preparation is crucial for accurate UHPLC analysis. The extraction process must be optimized to ensure that the target compounds are efficiently extracted without degradation or loss. Additionally, the presence of highly polar or non-polar compounds can complicate the preparation process and require multiple steps or the use of specialized techniques.

Column Overloading:
Due to the high sensitivity of UHPLC, there is a risk of column overloading when analyzing crude extracts, especially if the sample concentration is too high. This can lead to poor peak shape, reduced resolution, and decreased column lifetime.

Matrix Effects:
Matrix effects can significantly impact the ionization efficiency in mass spectrometry-based detection methods used in conjunction with UHPLC. These effects can result in ion suppression or enhancement, leading to inaccurate quantification of compounds.

Method Development:
Developing a UHPLC method that can effectively separate and detect a wide range of compounds present in crude plant extracts can be challenging. The method must be robust and reproducible, yet flexible enough to accommodate the diverse chemical properties of plant constituents.

Instrumentation Limitations:
While UHPLC systems are highly advanced, they are not without limitations. For instance, the high pressures used in UHPLC can lead to increased backpressure, which may require specialized equipment and maintenance to handle.

Data Analysis:
The large amount of data generated by UHPLC can be overwhelming, and the analysis can be complex, especially when dealing with unknown compounds or when trying to identify new bioactive compounds. Sophisticated software and skilled analysts are required to interpret the data accurately.

Cost and Accessibility:
UHPLC systems can be expensive, and the cost of consumables, such as columns and solvents, can be prohibitive for some laboratories, particularly in developing countries or for small-scale research projects.

Environmental Impact:
The use of large volumes of solvents in sample preparation and chromatography can have environmental implications. There is a growing need for green chemistry approaches that minimize the environmental footprint of UHPLC analysis.

Regulatory Compliance:
Ensuring that UHPLC methods comply with regulatory guidelines for plant extract analysis can be challenging, particularly when dealing with international standards and requirements.

Despite these challenges, the benefits of UHPLC in plant extract profiling are significant, and ongoing research and technological advancements continue to address these limitations, making UHPLC an invaluable tool in the field of natural product research.

9. Future Perspectives and Technological Advancements

9. Future Perspectives and Technological Advancements

As ultra high pressure liquid chromatography (UHPLC) continues to evolve, the future of plant extract profiling is poised for significant advancements. Here are some of the key future perspectives and technological advancements that are expected to shape the field:

Enhanced Sensitivity and Resolution:
- Future developments in UHPLC are likely to focus on increasing the sensitivity and resolution of the technique, allowing for the detection of even trace compounds in crude plant extracts. This will be crucial for identifying minor bioactive constituents that may have significant pharmacological effects.

Multidimensional Chromatography:
- The integration of multidimensional UHPLC systems will enable more complex separations, providing a comprehensive profile of plant extracts. This approach will help in distinguishing between structurally similar compounds and improving the overall quality of the profiling.

Hybrid Techniques:
- The combination of UHPLC with other analytical techniques such as mass spectrometry (MS) and nuclear magnetic resonance (NMR) will offer a more detailed chemical characterization of plant extracts. These hybrid techniques will provide structural information that is essential for the identification and elucidation of novel compounds.

Artificial Intelligence and Machine Learning:
- The application of artificial intelligence (AI) and machine learning algorithms in UHPLC data analysis will streamline the profiling process, enhance pattern recognition, and improve the accuracy of compound identification and quantification. AI-driven systems will also facilitate the prediction of bioactivity based on chemical profiles.

Nanotechnology Integration:
- The use of nanotechnology in UHPLC, such as the development of nano-structured stationary phases, will lead to higher separation efficiency and faster analysis times. This could be particularly beneficial for the analysis of complex plant matrices.

Miniaturization and Portability:
- Advances in miniaturization and the development of portable UHPLC systems will enable on-site analysis of plant extracts. This will be particularly useful for field studies and in situations where samples need to be analyzed without being transported to a laboratory.

Green Chemistry Approaches:
- There will be a growing emphasis on the development of environmentally friendly UHPLC methods that minimize the use of hazardous solvents and reduce waste. This includes the use of water-based solvents and the development of recycling systems for mobile phases.

Personalized Medicine and Omics Integration:
- As personalized medicine becomes more prevalent, UHPLC will play a crucial role in profiling plant extracts tailored to individual genetic profiles. Integration with omics data (e.g., genomics, proteomics, metabolomics) will help in understanding the interaction between plant compounds and human biology at a molecular level.

Data Standardization and Sharing:
- The establishment of standardized protocols and databases for UHPLC plant extract profiling will facilitate data sharing and comparison across different studies and laboratories. This will be essential for the validation of findings and the development of a comprehensive understanding of plant chemistry.

Regulatory Compliance and Quality Control:
- With the increasing use of plant extracts in pharmaceutical and nutraceutical products, there will be a greater need for UHPLC methods that are compliant with regulatory standards for quality control. This includes the development of methods that can accurately determine the potency, purity, and safety of plant extracts.

In conclusion, the future of UHPLC in plant extract profiling is bright, with ongoing technological advancements set to enhance the efficiency, accuracy, and applicability of the technique. These developments will not only improve our understanding of plant chemistry but also contribute to the discovery of new bioactive compounds with potential therapeutic applications.