Ekstraksi, Karakterisasi dan Uji Aktivitas Antioksidan Astaxanthin dari Produk Fermentasi Udang (Cincalok)

Shrimp is one of the aquatic organisms that contain several active compounds, including astaxanthin. Cincalok is one of the fermented shrimp products containing astaxanthin. This study aims to determine the characteristics of astaxanthin extract from cincalok and its antioxidant activity. Extraction of astaxanthin from cincalok was carried out using the reflux method with acetone : cyclohexane (20:80 v/v) as a solvent. The identification and characterization of astaxanthin was carried out using thin-layer chromatography (TLC), UV-Vis spectrophotometry, and High-Pressure Liquid Chromatography (HPLC). Meanwhile, the antioxidant activity test was carried out using the 1,1-diphenyl-2-picrylhydrazyl (DPPH) method in one serial concentration (5; 15; 25 ppm). The results of TLC analysis showed that astaxanthin in cincalok extract has Rf value (0.32). The analysis using a UV-Vis spectrophotometer produced a spectrum with a maximum wavelength of 477 nm, which corresponds to the maximum wavelength of standard astaxanthin. The yield of astaxanthin extract from cincalok in this study was 1.47 mg/100 g wet weight. The chromatogram from the results of UHPLC analysis showed that the retention time of cincalok astaxanthin extract was 6.27 minutes with a purity of 18.03%. The antioxidant activity of cincalok astaxanthin extract was 568.32 ppm. Udang merupakan salah satu organisme air yang mengandung banyak senyawa aktif, termasuk astaxanthin. Cincalok merupakan salah satu produk hasil fermentasi udang yang mengandung astaxanthin. Penelitian ini bertujuan untuk mengetahui karakteristik ekstrak astaxanthin dari cincalok dan aktivitas antioksidannya. Ekstraksi astaxanthin dari cincalok menggunakan metode refluks dengan pelarut aseton:sikloheksan (20:80 v/v). Identifikasi dan karakterisasi astaxanthin dilakukan dengan menggunakan kromatografi lapis tipis (KLT), spektrofotometri UV-Vis, dan High Pressure Liquid Chromatography (HPLC). Sedangkan uji aktivitas antioksidan dilakukan menggunakan metode 1,1-difenil-2-pikrilhidrazil (DPPH) dengan memvariasikan konsentrasi larutan uji, yaitu 5; 15; 25 ppm. Hasil dari penelitian ini melaporkan astaxanthin pada ekstrak cincalok menunjukkan nilai Rf 0,32 pada kromatografi lapis tipis (KLT). Hasil analisis menggunakan spektrofotometer UV-Vis menghasilkan spektra dengan panjang gelombang maksimum 477 nm, yang sesuai dengan panjang gelombang maksimum astaxanthin standar. Randemen ekstrak astaxanthin dari cincalok pada penelitian ini adalah 1,47 mg/100 g berat basah. Kromatogram dari hasil analisis UHPLC menunjukkan waktu retensi ekstrak astaxanthin cincalok yaitu selama 6,27 menit dengan kemurnian sebesar 18,03%. Aktivitas antioksidan dari ekstrak astaxanthin cincalok diperoleh sebesar 568,32 ppm.

Phase II Metabolism of Asarone Isomers In Vitro and in Humans Using HPLC-MS/MS and HPLC-qToF/MS

(1) Background: Metabolism data of asarone isomers, in particular phase II, in vitro and in humans is limited so far. For the first time, phase II metabolites of asarone isomers were characterized and human kinetic as well as excretion data after oral intake of asarone-containing tea infusion was determined. (2) Methods: A high pressure liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (HPLC-qTOF-MS) approach was used to identify phase II metabolites using liver microsomes of different species and in human urine samples. For quantitation of the respective glucuronides, a beta-glucuronidase treatment was performed prior to analysis via high pressure liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS). (3) Results: Ingested beta-asarone and erythro and threo-asarone diols were excreted as diols and respective diol glucuronide conjugates within 24 h. An excretion rate about 42% was estimated. O-Demethylation of beta-asarone was also indicated as a human metabolic pathway because a corresponding glucuronic acid conjugate was suggested. (4) Conclusions: Already reported O-demethylation and epoxide-derived diols formation in phase I metabolism of beta-asarone in vitro was verified in humans and glucuronidation was characterized as main conjugation reaction. The excretion rate of 42% as erythro and threo-asarone diols and respective asarone diol glucuronides suggests that epoxide formation is a key step in beta-asarone metabolism, but further, as yet unknown metabolites should also be taken into consideration.

Supramolecular Chromatographic Separation of C60 and C70 Fullerenes: Flash Column Chromatography vs. High Pressure Liquid Chromatography

A silica-bound C-butylpyrogallol[4]arene chromatographic stationary phase was prepared and characterised by thermogravimetric analysis, scanning electron microscopy, NMR and mass spectrometry. The chromatographic performance was investigated by using C60 and C70 fullerenes in reverse phase mode via flash column and high-pressure liquid chromatography (HPLC). The resulting new stationary phase was observed to demonstrate size-selective molecular recognition as postulated from our in-silico studies. The silica-bound C-butylpyrogallol[4]arene flash and HPLC stationary phases were able to separate a C60- and C70-fullerene mixture more effectively than an RP-C18 stationary phase. The presence of toluene in the mobile phase plays a significant role in achieving symmetrical peaks in flash column chromatography.

Enhanced pigment content estimation using the Gauss-peak spectra method with thin-layer chromatography for a novel source of natural colorants

Alternative pigment sources that are harmless to human health and can be produced in an eco-responsible way are of great research interest. The experiments undertaken in this study were conducted using autumn leaves of Aesculus hippocastanum as potential novel colorant sources. This study focused on improving the Gauss-peak spectra method (a less expensive alternative to high-pressure liquid chromatography) in combination with thin-layer chromatography, leading to the development of a new methodology. The collected leaves were stored at two different temperatures: 20°C and −20°C. The data obtained by spectrophotometric scanning of the samples were analyzed using the Gauss-peak spectra method in the R program with three wavelength ranges: 350–750 nm, 390–710 nm, and 400–700 nm. The results were then assessed for statistically significant differences in the estimated concentrations for the different wavelength ranges regarding (1) total pigment, carotenoid, and chlorophyll concentration (two-sample t-test) and (2) concentration of each indicated pigment (two-way analysis of variance). The results were also tested for differences between the estimated concentrations of samples stored under the different conditions. The Gauss-peak spectra results with and without thin-layer chromatography were statistically compared using a paired t-test. The results showed that thin-layer chromatography greatly enhanced the efficiency of the Gauss-peak spectra method for estimating the major and minor pigment composition without generating high additional costs. A wavelength range of 400–700 nm was optimal for all Gauss-peak spectra methods. In conclusion, the proposed method is a more successful, inexpensive alternative to high-pressure liquid chromatography.