**4.3 Supercritical fluid extraction (SFE)**

*Mycotoxins and Food Safety*

usually is present in food [20].

infertility in swine and poultry [21].

**4.1 Solid-phase extraction (SPE)**

combination with mass spectroscopy.

**4.2 Liquid: liquid extraction (LLE)**

**4. Isolation of a mycotoxin from real samples**

and temperature ≥15°C [18]. OTA is carcinogenic and neurotoxic for humans, and immunotoxic for animals [19]. OTA can cause various forms of kidney, liver, and brain diseases in both humans and animals, although the trace amount of OTA

Zearalenone (ZEN) produced by *Fusarium* or *Giberella* species grown on crops (maize, barley, oats, wheat, rice, also bread) is a potent estrogen metabolite causing

A variation of chromatographic techniques based on small disposable cartridges packed with silica gel or bonded phase, which in the stationary phase is the basic principle of solid-phase extraction. The sample loaded in one solvent under low pressure and rinsed to remove the most of contaminant are moved and eluted in another solvent. These cartridges have a high capacity for small binding molecules. Different bonding phase such as silica gel, aminopropyl, florisil, phenyl, ion exchange materials, anionic and cationic to affinity materials including immunoadsorbents and molecular imprinted polymers (MIPs) are available in SPE cartridges [22]. OTA formation occurs in some Spanish sweet, which going drying process. C-18 column had been shown successful recovery above 90% of OTA, which enables to be isolated from the matrix [23]. Silica gel frequently used in SPE because the surface of silica particles is heterogeneous with a variety of silanol group which can bind target compound through multiple electrostatic interactions. Generally, silica gel was used directly or after modification, and it is a hydrophobic phase which used in environmental and food analysis of toxin, which performed both polar and non-polar solvents. Previous research conducted by Leitner et al. [24] showed that the use of C-18 reverse-phase in the extraction of OTA from wine and offer good result with

Liquid–liquid extraction (LLE) or also known as solvent extraction agitating different solubility of toxin in the aqueous phase and an immiscible organic phase to extract the compound into one solvent and leaving the rest of matrix in others phase. A solvent such as hexane and cyclohexane are used to remove non-polar contaminant or molecule, for example, lipids, and cholesterol [25]. The common goal of LLE is sample clean-up and analyte component pre-concentration. Sample clean-up requires high selectivity of partitioning analyte component over potential interferents while analyte component pre-concentration require high distribution ratio to analyte can be extracted from a large volume of sample too small volume of extractant. Two bulk-liquid phases at least which are an aqueous phase that contains dissolved sample an organic extractant phase. The variety of condition will decide either the agitated mixture become the dispersed phase and another continuous phase. The thermodynamic driving force is resulting from the movement of chemical species from one bulk phase to another in two ways either by the difference in chemical potential for neutral species or electrochemical for ionic species [26]. Lately, Ezekiel et al. [27] used acetonitrile/water/acetic acid 79:20:1, (v/v/v) in a 50 mL polypropylene for the metabolites extraction and determination

**88**

of apparent recoveries.

Supercritical fluid extraction (SFE) had been used for years for industrial-scale separation and isolation of variety compound. SFE also has been utilizing in the field of food science to isolate not only natural food component but also unnatural compound like organic contaminants. SFE was developing and used as an alternative to extraction using liquid solvents. SFE considered an up-and-coming technique for the future because supercritical fluids have useful physical properties such as high viscosity and high diffusion constant for sample extraction which result in faster mass transport than regular and shorter the time for extraction. Using compressible gas like carbon dioxide (CO2), the solvation power can be changed by altering the density or decrease the pressure to atmospheric pressure [28].

Most common supercritical fluid (SF) used is SC-CO2, which is a suitable substituent for halogenated solvents. This is because the carbon dioxide is non-toxic, non-flammable, not significantly contribute to global warming and might be the cheapest solvent except for water. The usage of SFE to extract mycotoxin are very limited until recently because of the relative polar nature of mycotoxin and relative non-polar nature of food commodities such as nut and nut product. Taylor et al. [29] investigated the use of analytical SFE to remove aflatoxin Bi from field inoculated corn samples. Modification using a combination of various pressures "(2000- 15,000 psi), temperatures (40–80°C), the quantity of SC-CO2 (50–500 ml), and organic modifiers were used to optimize the extraction method. Optimal conditions were 5000 psi at 80°C with 15% modifier (acetonitrile/methanol 2:1) and a liquid carbon dioxide volume of 100 ml. The result gained from the extraction was 94.6% (RSD 6.2%, n = 5) of aflatoxin Bi could be recovered from ground corn contaminated at a level of approximately 500 μg/kg when using these settings.
