Sophesticated hyphenated analytical techniques - CE MS

CE MS

1. SOPHESTICATED HYPHENATED ANALYTICAL TECHNIQUES –CE-MS NAME-SHRUTI PANDIT. CLASS-FINAL YEAR B PHARMACY (SEM-VII ) under the Guidance of MISS. PRAJAKTA THETE . (M pharmacy) College name- K C T S R G Sapkal college of pharmacy anjaneri nashik.

2. Sr no. TITLE PAGE NO, 1 AIM AND OBJECTIVE 03 2 LITERATURE REVIEW 04 3 INTRODUCTION 07 4 PRINCIPLE 10 5 MASS SPECTROMETRY 17 6 COUPLING OF CE MS 21 7 FUTURE SCOPE 23 7 APPLICATIONS 24 8 CONCLUSION 25 9 REFERENCES 26

3. AIM – A REVIEW: TO STUDY THE SOPHESTICATED HYPHENATED ANALYTICAL TECHNIQUES ON CE MS. OBJECTIVES – I . To Understanding the principle and mechanism of capillary electrophoresis and mass spectroscopy . II . To study the advantages, disadvantages, principle, and procedure of the CEMS . III . Provide an overview of capillary electrophoresis (CE) and mass spectrometry (MS) highlighting their individual capabilities and limitations. IV . Explore the principle and benefits of hyphenated technique combining CE and MS. V . Highlight the future prospects and potential areas of improvement for CE MS hyphenation .

4. 3. LITERAT URE RE VIE W Dr. K . Bhavyasri et al*- (2020) Dr. k. Bhavyasri work in the international journal of pharmaceutical sciences focused on the application of CE MS (capillary electrophoresis and mass spectrometry,) CE MS is a powerful analytical technique that combines the separation capabilities of capillary electrophoresis with the detection and identification capabilities of mass spectrometry. By utilizing CE MS Dr Bhavyasri aimed to enhance the efficiency and accuracy of pharmaceutical analysis. Their work shed light on the utility of CE MS as a valuable tool. Dr Bhavyasri studied on instrumentation in detail, technical aspects of CE MS interface, application and the advancement made in this year is discussed. S. NAGAJYOTHI et al* – (2017) S. Nagajyothi has made significant contributions to the field of hyphenated techniques through her work published in the international journal of advanced research and development. Her paper titled ‘hyphenated techniques: a comprahensive review’. She studied on detail overview and analysis of various hyphenated techniques. The concept, principles, and applications, of hyphenated techniques. This technique involve the combination of two or more analytical methods. She studied the strength and limitation of each method like CE MS.

5. N RAMARAO,et al (2017) N Ramarao s work published in the international journal of research and development on the topic of A comprehensive review on sophesticated techniques. N Ramarao explores the different hyphenated techniques that have been developed and used in wide range these techniques involve two or more analytical technique such as chromatography, and spectroscopy to obtain a reliable information. Also studied on the principle methodologies, advantages, limitation of hyphenated techniques. JAMES M MILLER,et al (2005) He is the author of the book of chromatography concept and contrast He studied on the detail study about the capillary electrophoresis it involves the principle, advancement techniques, advantages, application, limitation, instrumentation of the capillary electrophoresis. It includes the study of separation of ions based on their electrophoretic mobility. H KAUR ,et al H Kaur is the author of the book of spectroscopy by pragati edition.In that he studied on the detail information about the mass spectrometry and other spectroscopies.it contains the principle, types, application, limitation, solution of the limitations etc he studied ‘the mass spectrometry is not a true spectroscopic technique because absorption of electromagnetic energy is not involve in any way’.

6. CAPILLARY ELECTROPHORESIS IN MASS SPECTROMETRY

7. INTRODUCTION HYPHENATED ANALYTICAL TECHNIQUES -; It is an separation technique.it referred as the combination of two or more techniques to separate chemicals from solutions and detect them.

8. CAPILLARY ELECTROPHORESIS (high separation efficiency in liquid phase ) MASS SPECTROMETRY (high separation efficiency in gas phase ) CE - MS INTRODUCTION

9. DEFINATION -: ; Capillary electrophoresis is an analytical technique that separates ions based on their electrophoretic mobility with the use of an applied voltage 1000volts/cm. A capillary is present by anode and cathode together.

10. Principle Capillary electrophoresis is a widely used analytical technique in various fields, such as chemistry ,biochemistry ,and pharmaceuticals. It is based on the principle of separating charged molecules in a capillary filled with an electrolyte solution under the influence of an electric field. One of the fundamental principle of capillary electrophoresis is electrophoretic mobility.

11. ELECTROPHORETIC MOBILITY Electrophoretic mobility refers to the movement of charged particles in an electric field. The principle of electrophoretic mobility allows for the separation and analysis of various analytes within a mixture by controlling the experimental conditions ,such as the PH of the electrolyte solution and the strength of the electric field . It is possible to optimize the separation of compounds with different electrophoretic mobilities.

12. The movement of analytes is affected by several factors including the analytes charge ,size ,shape, and interactions with the surrounding electrolyte and capillary wall . Under the effect of an electric field , Positively charged analyte move towards the negatively charged electrode (cathode). Negatively charged analyte move towards the positively charged electrode (anode).

13. INSTRUMENTATION

14. CAPILLARY – The capillary is a fused silica tube with a small inner diameter ( 10 – 100 um )that acts as the separation channel. Fused silica is preffered due to its excellent optical transmission properties.. The length of the capillary depends on the desired separation efficacy .

15. BUFFER RESERVIORS - This method requires two buffer reservoirs, an anode and a cathode reservoir, connected to the capillary. The buffer solutions in these reservoirs facilitate the movement of analytes by providing necessary conductivity and PH conditions. The anode reservoir contains a positively charged electrode while the cathode reservoir contains a negatively charged electrode. Common buffer systems include phosphate, borate, and tris-based buffers.

16. POWER SUPPLY - The power supply creates an electric field between the anode and cathode reservoirs, which drives the analytes through the capillary. Constant voltage or constant current modes can be employed, depending on the specific separation requirements. DETECTORS - UV- Visible absorbance detectors are commonly used, as they provide good sensitivity and allow for the detection of the wide range of analytes. Fluroscence detectors can enhance sensitivity for fluroscently labeled compound.

17. MASS SPECTROMETRY This technique works by exposing a vaporized sample to a high voltage electric current .the sample loses electrons and forms positively charged ions that are pushed along a circular path using magnetic and electrical field . The size of the path depends on the mass to charge ratio of the ions. (m/e). When the sample is exposed to an electron beam, it causes an electron to be knocked out from the sample molecule, resulting in the creation of a parent ion (M +).

18. SAMPLE INLET SYSTEM An interface between an ion source and a sample is called an inlet system. The inlet system's goal is to introduce the sample into the ion source with as little vacuum loss as possible. A port through which the sample is injected or placed into a chamber at high vacuum and heated to achieve vaporisation can serve as the basic inlet system. A GC inlet system will evaporate the sample and separate the mixture into its constituent parts if it is a mixture of compounds. As the components enter the mass spectrometer, their respective mass spectra are recorded one after the other. Therefore, it is possible to obtain the mass spectra of individual components of a complex mixture without first separating them. ION SOURCE As the mass analyzer only uses gaseous ions, ionisation becomes necessary. The molecules are typically ionised by adding a proton or removing an electron when they enter the mass spectrometer in the gas phase. b y subjecting the molecules to a high energy electron beam—typically consisting of 70 eV electrons—an electron can be removed from them. The energy of this electron beam is far greater than the energy at which the molecules are ionised during the bombardment. Because it has one unpaired electron, the resultant ion is a radical ion .

19. ELECTROSTATIC ACCELERATING SYSTEM Due to the potential difference between the first accelerator plate and the second repeller plate, a strong electric field exists between them, which accelerates the production of positively charged ions in the ionisation chamber. After going through the second accelerator plate, these ions reach their ultimate speeds. The first and second accelerating plates maintain a potential difference of roughly 400–4000V, which accelerates the ions of masses m1 , m2, m3, etc., to their final velocities. The ions that emerge from the slit are a collimated beam of high-kinetic energy ions travelling at high speeds . MAGNETIC FIELD The ions travel in a curved path as they enter the magnetic field after being accelerated by the electric field. The ion mass (m), accelerating voltage (V), electron charge (e), and magnetic field strength (H) all affect the radius of curvature (r). The mass-to-charge ratio (m/e) and curvature radius (r) are the foundations of mass spectrometry, which in turnhave a mutual reliance. Any modification to the magnetic field's (H) strength or accelerating voltage in alters the values of m/e and r.

20. ION SEPERATOR Another name for an ion separator is a mass analyzer. The core of a mass spectrometer is where ionised masses are taken and divided according to charge to mass ratios. There are numerous commercial types of mass analyzers available depending on the sorting technique. ION COLLECTOR Ions travel from the analyzer to the detector, where they produce a signal. The m/z value is obtained by further amplifying the signal. Typically, a direct writing recording oscilloscope with three to five galvanometers is used as the readout system. VACUUM SYSTEM Vacuum is necessary to permit the ions to reach the detector without colliding with any extraneous materials including atmospheric gases which themselves undergo ionization and fragmentation giving their spectra.

21. COUPLING OF CE MS Capillary Electrophoresis (CE) coupled with Mass Spectrometry (MS) combining CE’s high efficiency and high speed with the high sensitivity and high selectivity offered by MS detection is very attractive. There are several factors that must be considered when coupling the CE instrument to an MS detector.

22. 5 FUTURE SCOPE I Proteomics: CE-MS allows for the separation, identification, and characterization of complex protein mixtures . II Metabolomics: CE-MS allows for the comprehensive analysis of small molecules, metabolites, and their modifications within biological samples. III . Pharmaceutical analysis: CE-MS has the potential to become a key tool in drug development and quality control processes. IV . Environmental analysis: CE-MS proves useful in environmental monitoring, facilitating the detection and quantification of organic pollutants, heavy metals, and other contaminants in air, water, soil, and biological samples . V Forensic analysis: CE-MS plays a crucial role in forensic analysis by providing highly sensitive and selective detection of drugs, metabolites, and other toxic compounds in various forensic samples. VI. Food and beverage analysis: CE-MS finds application in food quality control, analyzing contaminants, additives, and adulterants. VII. Clinical diagnostics: CE-MS has potential in clinical diagnostics for the analysis of biological samples such as urine, blood, and cerebrospinal fluid

23. APPLICATIONS High efficiency of separation . Ability to separate both charged and non charged molecules. Offers new selectivity, an alternative to HPLC. Easy and predictable selectivity. Small size of sample (1 – 10 ul ) Faster separation (1-45 min ) Can be automated. Easily coupled to MS . It is use in the pharmaceutical analysis for purity and quality control. It is use in the bioanalysis for for analysis of protiens, peptides, nucleic acid, carbohydrate and metabolites. It is also used in environmental analysis for determining the presence and conc. Of various pollutants. (heavy metals, pesticides etc.) It is used in forensic analysis for identification and quantification of drugs, toxins, metabolites. It is widely used in food analysis for separation of food components, additives, contaminants, and allergens.

24. CONCLUSION Capillary electrophoresis is electrophoresis performed in a capillary tube. The hyphenated techniques are far better and useful than normal single techniques. Hyphenation includes both separation and identification which makes the analysis of samples easy. Nowadays the hyphenated techniques are more used than normal spectroscopic and chromatographic techniques.

25. REFERENCES 1) Ruchira chin chole et. Al.recennt applicatios of hyphenated liquid chromatography techniques international journal of pharmaceutical science review and research; 2012, 14 (1)57-63. 2) Kemp G; capillary electrophoresis; a versatile family of analytical techniqus. Biotechnology and applied biochemistry 1998; 27(1);9-17. 3) Skoog DA Holler FJ and Crouch SR; principles of instrumental analysis. Thomson Brooks/ Cole publishing sixth edition 2007. 4) S. Pleasance, P.THIBAULT and J. KELLY, J Chromatogr.,591 (199) 325-339. 5) J Henion, A Mordhai and J. Cai, Anal. Chem., 66 (1994) 2103-2109. 6) Janini GM and Issaq HJ ; THE BUFFER IN CAPILLARY ZONE electrophoresis. Chromatographic science 1993; 64; 119-160. 7) Cifuentes A and Poppe H; Rectangular capillary electrophoresis ; some heretical considerations. Chromatographia 1994; 39(7-8); 391-404. 8) Geiser L, Cherkaoui S and Veuthey JL: Simultaneous analysis of some amphetamine derivatives in urine by nonaqueous capillary electrophoresis coupled to electrospray ionization mass spectrometry. J Chromatography A 2000; 895(1-2): 111-21 9) Wiley HE: mass spectroscopy principles and applications.Wiley-Interscience, Third edition 2007 10) Ross GA: Capillary electrophoresis-mass spectrometry:practical implementations and applications. Agilent Technologies 2001

26. 20) JAMES . M . MILLER chromatography concept and contrast edition second 2005 page no. 365-383. 21) H KAUR spectroscopy by pragati edition page no. 443-509. 22) WWW.WIKIPEDIA.ECYCLOPEDIA . 23) https://www.slideshare.net . 24) www.ezyaccess.in . 25) http://www.delnet.in . 26) https://www.sciencedirect.com 27) https://www.chromatographytoday.com

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