Teknik Aplikasi Sampel Pada Pengujian Kuantitatif Kromatografi Lapis Tipis: Tinjauan Terhadap Area Dan Faktor Retensi

Idha Kusumawati; Riesta Primaharinastiti; Hanif Rifqi Prasetyawan

doi:10.32382/mf.v20i2.655

Penulis

Idha Kusumawati Universitas Airlangga, Surabaya, Indonesia https://orcid.org/0000-0002-9451-3098
Riesta Universitas Airlangga, Surabaya, Indonesia https://orcid.org/0000-0003-1052-7765
Hanif Universitas Airlangga, Surabaya, Indonesia https://orcid.org/0009-0002-8857-6188

DOI:

https://doi.org/10.32382/mf.v20i2.655

Kata Kunci:

Densitometry, Peak Area, Retention Factor, Thin Layer Chromatography

Abstrak

Sample Application Techniques In Quantitative Determination of Thin Layer Chromatography: Review on Area and Retention Factor

Thin Layer Chromatography (TLC) is one of the most commonly used chromatographic techniques to identify compounds in mixtures and determine the purity of a compound. However, the crucial factor that is difficult to control, which can affect the accuracy of the results, is the stage of sample application. This study focused on investigating the effect of different techniques of sample application (manual and semi-automatic) on the area and retention factor (Rf) of the compound spot. Curcuminoid was used as a sample in this study. The manual sample application techniques used a 2μL capillary tube. The semi-automatic sample application techniques used a Camag Linomat-5 instrument with 0,7, and 8 mm band lengths. After the development of the TLC plate using toluene: acetic acid (4:1, v/v)., the curcuminoid was separated into three spots: curcumin, demethoxycurcumin, and bisdemethoxycurcumin. A one-way ANOVA was used to analyze the spot area and Rf of the three compounds, followed by a Duncan test. The difference in sample application techniques causes differences in the size and shape of the initial sample spot, which affects the compound spot produced after developing of TLC plate. This study showed that the differences in application techniques cause significant differences in the Rf of the compound spot, but do not show significant differences in the compound spot area. Based on this research, optimizing the application techniques of samples is a critical stage and it is essential to optimize how to apply the sample on TLC plate, including the concentration, volume, shape, and size of the spot on TLC.

Kromatografi Lapis Tipis (KLT) adalah salah satu teknik kromatografi yang paling umum digunakan untuk mengidentifikasi senyawa dalam campuran dan menentukan kemurnian suatu senyawa. Namun, faktor krusial yang sulit dikontrol dan dapat mempengaruhi keakuratan hasil adalah tahap aplikasi sampel. Penelitian ini difokuskan untuk menyelidiki pengaruh teknik aplikasi sampel yang berbeda (manual dan semi-otomatis) terhadap luas area dan faktor retensi (Rf) spot senyawa. Kurkuminoid digunakan sebagai sampel dalam penelitian ini. Teknik aplikasi sampel secara manual menggunakan tabung kapiler 2μL. Teknik aplikasi sampel semi-otomatis menggunakan instrumen Camag Linomat-5 dengan panjang pita 0, 7, dan 8 mm. Setelah pengembangan plat KLT menggunakan toluena: asam asetat (4: 1, v/v), kurkuminoid dipisahkan menjadi tiga bercak: kurkumin, demetoksikurkumin, dan bisdemetoksikurkumin. ANOVA satu arah digunakan untuk menganalisis area bercak dan Rf dari ketiga senyawa tersebut, diikuti dengan uji Duncan. Perbedaan teknik aplikasi sampel menyebabkan perbedaan ukuran dan bentuk bercak sampel awal yang mempengaruhi bercak senyawa yang dihasilkan setelah pengembangan plat KLT. Berdasarkan penelitian ini dapat diketahui bahwa proses pengapikasian atau penotolan sampel menjadi tahapan penting untuk dilakukan optimasi cara pengaplikasian sampel meliputi konsentrasi atau volume penotolan serta bentuk dan ukuran spot noda penotolan sampel, saat analisis menggunakan metode KLT.

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