# Chemical Compositional Changes in Over-Oxidized Fish Oils
## Discussion
### Specific Groups of Secondary Oxidation Products and Biological Activity
#### 4.4.2 Isoprostanoids  
The present study reports for the first time that several isoprostanoids in unesterified free form are present in refined fish oils, and that specific isoprostanoids are formed during the development of rancidity. Isoprostanoids, and specifically F\({}_{2}\)-isoprostanes, were originally identified as esterified products within phospholipids in peroxididized biological membranes [80]. Isoprostanoids can also be formed from unesterified omega-3 LCPUFA under peroxid oxidative conditions [81; 82]. Given the absence of active deacylating enzymes in refined oils, either the isoprostanoids are released by hydrolysis during oxidation or they are uniquely formed from free fatty acids in the oils (although we do not know what the levels are in this study). It is also possible that isoprostanoids formed during oxidation will be present in esterified form within the triglycerides that make up the bulk of the hoki liver oil and anchovy oil. This will need to be further investigated. Interestingly, all the tested isoprostanoids were also present at baseline, which suggests that these are also formed during the extraction and refining of the oil, and/or they originate from the fish from which the oil was originally extracted. Since isoprostanoids are chemically relatively stable at ambient conditions, it will be interesting to evaluate if specific members could be suitable markers for secondary oxidation in omega-3 EPA/DHA oils, including low-grade oxidation in ingredient oils and finished products. Recently, neuroprostanes derived from DHA and isoprostanes from EPA were shown to be present in two encapsulated marine dietary supplement oils: calanus oil and a fish oil composed of a mixture of sardine, anchovy, mackerel, and herring oil [83]. Several phytoprostanes, isoprostanes, and neuroprostanes identified in the over-oxidized hoki liver oil at day 30 were present at higher levels than before the exposure to oxygen and light or at day 5 or day 15. Several of these substances were also present in considerably higher levels than in anchovy oil. However, strictly taken, at day 15, none of the measured isoprostanoids had increased compared to baseline, which in many cases had initially decreased from baseline to day 5. Between day 5 and day 30, a range of isoprostanoids were formed increasingly, ultimately reaching levels markedly surpassing baseline, typically about 3.5 times. The most abundant isoprostanoids formed from day 5 onwards were 5-_epi_-5-F\({}_{34}\)-IsoP (0.32 ng/g oil) and 5-F\({}_{34}\)-IsoP (0.1 ng/g oil), which were both derived from EPA, and the DHA-derived 4(\(RS\))-4-F\({}_{44}\)-NeuroP (0,2 ng/g oil), 20-_epi_-20-F\({}_{44}\)-NeuroP (0.023 ng/g oil), and 14(\(RS\))-14-F\({}_{44}\)-NeuroP (0.014 ng/g oil). Thus, specific isoprostanoids could be considered of interest to further evaluate to contributing to, or modulating, the postnatal and maternal toxicity as observed by [PERSON] et al. The abundance of individual PUFA species in each oil was related to the concentrations of isoprostanoids formed. EPA and DHA are clearly a chemical substrate for the formation of specific isoprostanoids, but also other major LCPUFA species present in fish oils, such as AA and ALA, as well as LCPUFA found in lower abundance, such as adrenic acid and DPA n-3, were converted to the corresponding isoprostanes and phytoprostanes (and related furans). In vivo, tissue levels of EPA- and DHA-derived isoprostanes and neuroprostanes increase following dietary supplementation with EPA and DHA [37]. While all the measured isoprostanoids were found in relatively low concentrations, some of these substances have potent biological activity. For example, 15-F\({}_{24}\)-IsoP has potent vasomotor activity, and it activates platelets [84]. 5-_epi_-5-F\({}_{34}\)-IsoP regulates neurotransmitter release via activity on the prostanoid EP1 receptor [36]. With regard to the perinatal period, F\({}_{2}\)-isoprostanes are strongly related to the consequences of intra-uterine hypoxia and neonatal morbidity [85].