THIN LAYER CHROMATOGRAPH Y Pn. Suryati Bt. Syafri 2009 Learning Objective 1. 2. 3. 4. 5.

State the definition of TLC Explain the phases used in TLC List the materials & methods used in TLC List the application of TLC List the advanteges & disadavantages of TLC Thin Layer Chromatography What is KLN?... One of analysis method that is used to identify the unkonwn compounds and

to determine the purity of mixture. This method is simple, rapid and cheap Widely used in pharmaceutical & food stuff industry. -A plate of TLC can be made from aluminium or glass which is coated by a solid matter as a stationary phase. - The coated material has 0.1-0.3mm in

thickness -some of them has been added by fluorescent indicator that will make it florescence during the UV light exposure. STATIONARY PHASE Silica is commonly used as stationary

phase The separation of sample mixture will be depent on the polarity of sample. Some modified silica is also used in certain purposes. Stationery phase Description Application Silica gel G

Silica gel with average particle size 15m containing ca 13% calcium sulfate binding agent Used in wide range pharmacopoeial test Silica gel G254 Silica gel G with

fluorescence added Same application with Silica gel G where visualization is to be carried out under UV light. Cellulose Cellulose powder of less than 30m particle size.

Identification of tetracyclines MOBILE PHASE The ability of mobile phase to move up is depent on the polarity itself Volatile organic solvents is preferably used as as mobile phase. MOBILE PHASE

SOLVENT POLARITY INDEX Heksana 0 Butanol 3.9 Chloroform

4.1 Methanol 5.1 Ethanol 5.1 Acetonitrile 5.8

Air 9.0 MATERIALS TLC plate

Developing container - chamber/ jar/ glass beaker Pencil Ruler Capillary pipe Solvents / mobile phase - organic solvents UV lamp METHOD 1.Developing Container Preparation Solvent is transferred

into the container with 0.5-1cm in dept from the bottom 2. TLC Plate Preparation Commercialy obtained with 5cm x 20cm in size Prepare your size when neccesary Line 1 cm from the bottom

with a pencil as a part should be spotted. 3.Spotting TLC plates Make sure that your sample is liquified already. stick it using capillary pipe & spott onto the line you have made

4.Develop the plate after spotting, put the plate inside the chamber in the ascendant position Make sure that the dept of solvent doesntb touch the spots Let it develop up to the 1cm from

the top of plate After that, pull out the plate from the cahmber and let the solvent be vaporized 5. Detection of spots - The color samples are easy to be seen and no need to use UV lamp to detect them 6. DETECTION OF SPOT 1)

2) 3) 4) Iodination-put the plate in which the spots face to the iodine crystall and see what is the spot color changing Ninhydrin: -spesific identification of amino acid compounds. - Ninhydrin solution will show a purple spot when it is sprayed to the amino acid spot.

KMnO4 used to identify a reducing agent such as glucose, fructose, vitamin C and others. Alkaline tetrazolium blue specificaly used for corticosteroid identification The use of Rf as separation parameter - The distance taken through by the solvent to move up will be assigned as solfent front - The distance taken trrough by the sample to move up will be assign as sample front - Rf value is obtained by dividing the sample front toward solvent front

Rf = sample front solvent front - Thin-Layer Chromatography: Determination of Rf Values solvent front Rf of component A = dA component B

dS Less polar! dS Rf of component B = dB dB dS The Rf value is a decimal fraction, generally only

reported to two decimal places component A dA More polar! origin 7. Quantitative determination of known sample - Done by scratching the spot using spatula, and extract the compound using the suitable solvent

- The liquid extract can be determined its content using other method such as spectroscopy. Prob;ems commonly occur in TLC and how to solve a. The spot shape is too broad - Diameter is supposed to be < 1-2mm b. The movement of solvent - should be straight up - unproportionality in stationary phase surface will inhibit the movement of solvent

c. streaking formation - caused by too concentrated sample TLC Compared to Paper Chromatography 1. Precise and effective 2. More stable toward various organic solvents Advantages

Cheap Simple The developing can be monitored visually Able to use various chemical as a detector Identification unknown drugs using standard Reference Rujukan 1. 2.

3. R.A.Day, Jr. A.L.Undewood(1987). Analisis Kualititatif. Edisi ke Empat, ms: 474-529. David G. Watson(2005). Pharmaceutical analysis. Edisi ke-2, ms 315-331 http// hndbooksupport/TLC/ TLCprocedure.html Terima kasih

Recently Viewed Presentations

  • Behavioral Research Chapter 10 - Texas Christian University

    Behavioral Research Chapter 10 - Texas Christian University

    Factorial design. Designs with more than one indep var or factor . all levels of each indep var are combined with all levels of the other indep var. The simplest type of factorial design is a 2 X 2—has two...
  • Jacob S. Kounin

    Jacob S. Kounin

    Kounin is a classroom behaviourist theorist Best known for his two studies done in 1970 He wrote the book, "Discipline and Group Management in Classrooms" Kounin worked to combine both discipline and learning in the classroom Kounin believed that organization...
  • CNIT131 Internet Basics & Beginning HTML Week 12

    CNIT131 Internet Basics & Beginning HTML Week 12

    Coaxial cable was the transmission medium of choice for many early networks and still is found in cable television connections. Coaxial cable (Figure 6-9), pronounced coax (KO-ax), consists of a single copper wire surrounded by insulating material, woven or braided...
  • Unit 1 Lesson 5 - Welcome to Biology and Botany/Ecology Class!

    Unit 1 Lesson 5 - Welcome to Biology and Botany/Ecology Class!

    Darwin realized the struggle for survival due to the difficulties (or selection pressure) caused by nature must be selecting for heritable trait variations that will allow one organism within a population to out-reproduce the other members of the population. Darwin...
  • Multimodal Approaches to QTL Analysis.

    Multimodal Approaches to QTL Analysis.

    Multimodal Complex Trait Analysis: Combined Use of F2, HS, RI, SNPs, SSLP Haplotypes and Arrays to Fine-Map Behavioral Traits" Robert Hitzemann, Ph.D.


    FOUR WAYS OF SETTING ASIDE GOD'S LAW SETTING ASIDE GOD'S LAW The Bible contains God's law. As long as man lives in harmony with that law, he pleases God. When he violates that law, he sins. Sin is the transgression...
  • Detective Constable Mark Aldridge FELO 1 Detective Constable

    Detective Constable Mark Aldridge FELO 1 Detective Constable

    UK's national fraud and internet crime reporting centre. Set up in 2009 by National Fraud Authority (originally part of the Home Office) Report and record fraud and information about fraud on behalf of the Police. All reports are sent to...
  • PowerPoint - Famous Chefs and Entrepreneurs in the Food ...

    PowerPoint - Famous Chefs and Entrepreneurs in the Food ...

    The contemporary kitchen, from institutional to fine-dining restaurant, runs according to a strict hierarchy in which the chef plays the lead role. Assignments differ based on the precise needs of a given kitchen, but in most upscale American and European...