# Chemicals from Brominated Flame Retardants: Analytical Methods, Occurrence, Transport and Risks
## 2 Analytical Methods for the Detection of BFRs in Environmental and Biological Samples
### Sample Collection and Extraction Methods
#### 2.1.1 Biotic Samples
Serum and Urine SamplesThe analysis of BFRs in serum and urine samples have been reported by many researchers in a bid to understand the metabolism, transport, and elimination of BFRs in the human body. Several extraction techniques can be used to extract BFRs from serum and urine samples. Methods frequently used include LLE [27; 28], SPE [29], QuEChERS [30], and ASE [31], while recent advancements in extraction methods have seen the use of novel microextraction techniques such as Solid-Phase Microextraction (SPME) and Liquid-Phase Microextraction (LPME). The extraction of Serum and urine samples using hollow fiber liquid-phase microextraction (HF-LPME) have been reported by [PERSON] et al. [32]. [PERSON] et al. [32] developed a method that employed the use of nitrogen-doped carbon nanotubesN-CNTs) reinforced hollow fiber liquid-phase microextraction (HF-LPME) combined with high-performance liquid chromatography (HPLC) for the simultaneous determination of TBBPA and decabromodiphenyl oxide (BDE209) in serum and urine samples. The solvents used for the extraction were toluene-nocanol (1:1) and toluene-ethyl acetate (1:1). The hollow fiber membrane blocked the interference of macromolecules in the sample during the extraction process which played a role in purification. The study results showed quantitation limits (LOQs) ranges of 0.375-2.8 ng/mL and 1.25-9.4 ng/mL, respectively. The average recoveries for the two target analytes at three spiked levels ranged from 84.5-11%. The researchers concluded that the method could not only enrich TBBPA and BDE209 but also remove protein from biological samples at the same time. Food and FeedSolid food samples are usually blended into particulate size to increase the surface area, and liquid samples are homogenized to ensure the uniform distribution of BFRs throughout the sample prior to extraction. Soxhlet Extraction (SE) is widely used for extractions of BFRs from fat-containing food samples to extract lipophilic BFRs [33]. However, this method can be time-consuming and may require large amounts of solvent. Alternative methods include SPE [34], ASE [35], matrix solid-phase dispersion (MSPD) [36], pressurized liquid extraction (PLE) [37], and QuEChERS methods [38]. ASE is increasingly becoming the routine extraction method for the analysis of BFRs in food and feed [39; 40]. However, multiple steps (combined extraction and clean-up) may be applied simultaneously when using ASE, consequently reducing analysis time and solvent consumption. This can be observed from the analysis of brominated flame retardants (BFRs) in animal-derived food samples from Nigeria [40]. The recovery from the analysis ranged between 31 and 135%, and the method LOD ranged between 0.042 and 2.0 (ng/g). [PERSON] et al. [36] designed a novel pretreatment method for the extraction of polybrominated diphenyllethers (PBDEs) using matrix solid phase dispersion (MSPD) and depth purification using dispersive liquid-liquid micro-extraction (DLLME) from vegetables. The results showed satisfactory recoveries of spiked samples (82.9-113.8%, except for BDE-183 (58.5-82.5%)) and matrix effects (\(-\)3.3-18.2%). The limits of detection and the limits of quantification were in the range of 1.9-75.1 ug kg\({}^{-1}\) and 5.7-25.3 ug kg\({}^{-1}\), respectively. A novel and reliable analytical method was developed and validated for the simultaneous determination of 1,3,5-tris(2,3-dibromopropyl)-1,3,5-triazine-2,4,6(1H,3H,5H)-trione (TDBP-TAZTO), and 2,4,6-tris(2,4,6-tribromophenoxy)-1,3,5-triazine (TTBP-TAZ) in environmental samples using high-performance liquid chromatography coupled to a tandem mass spectrometer [37]. The extraction efficiency of sediment and fish samples was tested with different techniques (pressurized liquid, solid-liquid, ultrasound-assisted, and Soxlet extraction). Additionally, cleanup using modified multilayer silica gel (sediment) and gel permeation chromatography as well as Florisil columns (fish) with several solvent mixtures were performed.