The results of the present study supported our hypothesis that metabolic inflexibility, indicated by a pragmatic proxy measure of fasting resting energy expenditure to food quotient (RER:FQ) ratio, was associated with insulin resistance and family history of diabetes mellitus, in otherwise apparently healthy women, independent of body fatness and age. Insulin resistance was associated with lower rates of whole body fat oxidation in relation to habitual dietary fat intake. This extends the findings of Ukropcova , who demonstrated impaired metabolic flexibility in response to short-term high-fat feeding, in healthy individuals, as measured by changes in resting energy expenditure during sleep or in response to a hyperinsulinaemic, euglycaemic clamp. Similar to the current study, this effect was most marked in those with a family history of Type 2 diabetes mellitus. However, unlike previous studies, the present study demonstrated these relationships using simple fasted resting measures of substrate oxidation, against self-reported, habitual dietary fat intake, without any manipulation of diet, or induced glycaemia .
The link between substrate oxidation or metabolic flexibility and family membership, as well as, family history of diabetes, has been demonstrated using 24-hr resting energy expenditure [14, 30], fasting substrate oxidation  and suppression of fat oxidation in response to insulin-induced glycaemia and in response to high-fat feeding, Ukropcova [17, 19], hypothesized that patterns of fat oxidation are genetically or epigenetically determined, in that, the differences in metabolic phenotype were independent of body fat percentage. Taken together, these results suggest that metabolic inflexibility is linked to genetic or epigenetic phenomenon, and is not secondary to insulin resistance, or the result of obesity, per se.
In previous studies, metabolic inflexibility has been shown to be concomitant with reduced skeletal muscle mitochondria size and density, [17, 35] with an associated decrease in capacity for complete oxidative disposal of fat and loss of insulin sensitivity. Meex,  demonstrated that, 3 months of exercise training improved insulin sensitivity in both type 2 diabetics and non-diabetic individuals, along with metabolic flexibility, particularly in relation to insulin-stimulated, oxidative, glucose disposal. In addition, mitochondrial function was increased in both groups. This is consistent with the results of the current study, in which the total physical activity levels and RER:FQ ratio contributed to the overall variance in HOMA-IR, or insulin resistance (Table 5). However, we found no direct correlation between self-reported physical activity and resting energy expenditure or the RER:FQ ratio. The lack of a direct association between physical activity, fasting substrate oxidation and metabolic flexibility, as measured in the present study, may be related to the fact physical activity levels were generally low, and the data were not normally distributed. Alternatively subjects may have over reported physical activity levels or the GPAQ may not have been sufficiently sensitive to distinguish between groups. Not unexpectedly, we found that the insulin resistant group reported lower levels of physical activity and a higher dietary fat intake , and thus, may have additionally confounded this relationship in bivariate analyses.
One unexpected finding in the present study, was the inverse associations between serum free fatty acids concentrations and insulin resistance, in these non-diabetic, otherwise healthy women. While there are numerous studies that have shown that elevated free-fatty acid levels may be implicated in the aetiology of insulin resistance and are associated with Type 2 diabetes mellitus [37, 38], there is also some indication that the sequelae of insulin resistance may be tissue- and substrate-specific [16, 39]. As such, it is possible that in these non-diabetic subjects, the increased circulating insulin concentrations, were still effective in the suppression of lipolysis or in the non-oxidative disposal of fatty acids .
In a cohort study, Bickerton,  compared men in the highest and lowest quartiles for fasting plasma insulin and found that there were no differences between groups with respect to circulating free fatty acid concentrations. However, those with the highest fasting insulin also had higher triglyceride concentrations. In a smaller metabolic study, insulin-resistant men were compared to weight-matched, BMI-matched controls. Despite higher insulin levels, the insulin-resistant group had similar fasting free fatty acid concentrations when compared to control subjects, and systemic fatty acid production and appearance were lower. These authors suggest that there are ‘altered metabolic partitioning’ of fatty acids (reduced oxidation, increased re-esterification), in response to the insulin resistant state. This is consistent with Forbes , who found that lipolysis, palmitate appearance and palmitate oxidation were actually lower in non-obese women at risk for diabetes compared to controls. While at the same time, they demonstrated higher plasma insulin levels, suggesting increased capacity for storing fat.
The influence of diet on insulin sensitivity is mediated by both energy and nutrient content, in particular by different types of dietary fatty acids [42, 43]. The higher dietary fat intake (Table 3) of the insulin resistant group compared to the insulin sensitive group directly relates to insulin sensitivity . We also found that higher dietary ω-3 fatty acids, specifically C20:5, C22:5, and C22:6, were strongly associated with increased fat oxidation (data not shown). Thus, specific dietary fatty acids may modulate the relationship between substrate oxidation and insulin resistance, by altering fatty acid partitioning, towards either storage or oxidation .
One limitation of the present study was that total fat intake was assessed by a quantified validated food frequency questionnaire (FFQ) , rather than an objective biomarker, which may have introduced the potential for bias, as over and under reporters were excluded from the analysis. However, a study in a sample of American men comparing fatty acid intake assessed by subcutaneous fat aspirate , 2-weeks diet records, and an amended version of the food frequency questionnaire used in the US Nurses’ Health Study  suggested that the estimates of fat intake from the food frequency questionnaire were as valid as those from the diet records.