The benefits of a Mediterranean Diet (MedDiet) in the management of diabetes have been reported, but the contribution of polyphenol-rich citrus fruit has not been studied widely. Here, we report the sub-study findings of a previously conducted MedDiet intervention clinical trial in patients with type 2 diabetes mellitus (T2DM), where we aimed to measure the diet intervention effects on plasma citrus bioflavonoids levels and biomarkers of inflammation and oxidative stress. We analysed plasma samples from 19 (of original 27) participants with T2DM who were randomly assigned to consume the MedDiet intervention or their usual diet for 12 weeks and then crossed over to the alternate diet. Compared with baseline, MedDiet significantly increased levels of the citrus bioflavonoids naringin, hesperitin and hesperidin (by 60%, 58% and 39%, respectively, p < 0.05) and reduced plasma levels of the pro-inflammatory cytokine IL-6 (by 49%, p = 0.016). Oxidative stress marker 8-hydroxy-2′-deoxyguanosine (8-OHdG) decreased by 32.4% (p = 0.128). Usual diet did not induce these beneficial changes. The reduced inflammatory profile of T2DM participants may, in part, be attributed to the anti-inflammatory actions of citrus bioflavonoids. Together with indications of improved oxidative stress, these findings add to the scientific evidence base for beneficial consumption of citrus fruit in the MedDiet pattern.

The most potent of the indole diterpenes, lolitrem B, is found in perennial ryegrass (Lolium perenne L.) infected with the endophyte Epichloë festucae var. lolii (also termed LpTG-1). Ingestion causes a neurological syndrome in grazing livestock called ryegrass staggers disease. To enable the rapid development of new forage varieties, the toxicity of lolitrem B and its biosynthetic intermediates needs to be established. However, most of these indole diterpenes are not commercially available; thus, isolation of these compounds is paramount. A concentrated endophyte-infected perennial ryegrass seed extract was subjected to silica flash chromatography followed by preparative HPLC and purification by crystallization resulting in lolitrem B and the intermediate compounds lolitrem E, paspaline and terpendole B. The four-step isolation and purification method resulted in a 25% yield of lolitrem B. After isolation, lolitrem B readily degraded to its biosynthetic intermediate, lolitriol. We also found that lolitrem B can readily degrade depending on the solvent and storage conditions. The facile method which takes into consideration the associated instability of lolitrem B, led to the purification of indole diterpenes in quantities sufficient for use as analytical standards for identification in pastures, and/or for toxicity testing in pasture development programs.

The neuroactive mycotoxin lolitrem B causes a neurological syndrome in grazing livestock resulting in hyperexcitability, muscle tremors, ataxia and, in severe cases, clonic seizures and death. To define the effects of the major toxin lolitrem B in the brain, a functional metabolomic study was undertaken in which motor coordination and tremor were quantified and metabolomic profiling undertaken to determine relative abundance of both toxin and key neurotransmitters in various brain regions in male mice. Marked differences were observed in the duration of tremor and coordination between lolitrem B pathway members, with some showing protracted effects and others none at all. Lolitrem B was identified in liver, kidney, cerebral cortex and thalamus but not in brainstem or cerebellum which were hypothesised previously to be the primary site of action. Metabolomic profiling showed significant variation in specific neurotransmitter and amino acid profiles over time. This study demonstrates accumulation of lolitrem B in the brain, with non-detectable levels of toxin in the brainstem and cerebellum, inducing alterations in metabolites such as tyrosine, suggesting a dynamic catecholaminergic response over time. Temporal characterisation of key pathways in the pathophysiological response of lolitrem B in the brain were also identified.