Evaluación estadística y recolección de datos

This study reports no significant changes in bone resorption markers, urinary DPD and plasma CTx (figures 4.2 and 4.3). The current study group had similar mean values of urinary DPD and plasma CTx compared to postmenopausal women of the same age. Baseline plasma CTx in this study was 0.476-0.502 r 0.018 ng/ml. A previous study reported plasma CTx of 0.43 r 0.15 ng/ml in healthy postmenopausal women (57.5 r 4.7 years, n=27) (Valderas et al., 2009). This study reported average baseline urinary DPD of 6.61-7.82 r 1.16 mmol/mmol creatinine. Previously, healthy postmenopausal women (62.1 r 2.4 years, n=42) had an average of 6.99 r 0.72 mmol/mmol creatinine (Arjmandi et al., 2003).

The results regarding DPD are in contrast to a recent meta-analysis (Taku et al., 2010b): 50-90 mg/day of aglycone isoflavone for 10 weeks to 12 months, decreased urinary DPD by 18.5% in postmenopausal women. The current study was a six-week intervention and may have been be too short to assess a change in DPD.Two previous short-term isoflavone interventions, each of four-week duration, found a significant decrease in DPD in women receiving aglycone isoflavone supplementation, 14 mg/day (Mori et al., 2004), and 38.4 mg/day Uesugi et al., 2002), which are lower doses than the dose used in the current study. However, the perimenopausal group in the trial by Uesugi and colleagues (2002) may have been more responsive to isoflavone treatment (Ma et al., 2011) compared to the 1-10 year postmenopausal women in this study. Furthermore, the trial by Mori et al. (2004) was considered low quality (Taku et al., 2010a). In regards to equol production and bone resorption markers, Wu et al. (2006) measured urinary DPD in response to 75 mg/day isoflavone conjugates, and found no change in DPD in either equol producers or non-producers. Equol levels reached 81

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ng/mL in the equol producers (mean age 55 years) (Wu et al., 2006). Interestingly, BMD loss was inhibited at the subtotal body and hip exclusively in the equol producers (Wu et al., 2006). Urinary DPD decreased by 24% in non-equol producers after treatment with a pure S-equol supplement, 10 mg/day for 12 months (Tousen et al., 2011). Serum equol reached 85.1 nmol/L (20.7 ng/ml) during supplementation (Tousen et al., 2011).

This study adds to the limited literature regarding the effect of isoflavone supplementation on plasma CTx in postmenopausal women. There have been no meta-analyses in this area as there are only two relevant studies that have measured this parameter (Albertazzi, Steel & Botazzi, 2005; Marini et al., 2008). In these studies, plasma CTx did not respond to isoflavone supplementation. Pure genistein supplements were used in the studies by Albertazzi et al. (2005) and Marini and colleagues (2008) whereas this study used a mixed supplement. Marini and colleagues (2008) found that when analysing each year separately that there was a significant decrease in plasma CTx over years 2-3, but there was high participant dropout for the third year that would bias the results. More long-term RCT are needed to elucidate the relationship between isoflavone and plasma CTx in postmenopausal women. It is important that these future studies measure equol production.

One potential effect on bone turnover not estimated in this study was the effect of the vitamin C content of kiwifruit on calcium absorption from the diet. Vitamin C has been shown to facilitate calcium absorption and retention in an animal study (Morel & Wolber, unpublished data, as cited in Wolber et al., 2013, P249). Given that consumption of two green kiwifruit daily provided an additional 412-464% of the RDI for vitamin C, the kiwifruit and isoflavone treatment had the potential to increase the fractional absorption of intestinal calcium. However, serum calcium and PTH were not measured, so the effect of kiwifruit on calcium retention is unknown. Nonetheless, this effect seems unlikely as neither of the bone resorption markers decreased in response to the kiwifruit and isoflavone treatment.

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There are several potential reasons for the lack of association between isoflavone supplementation and bone resorption observed in this study. Firstly, the intervention duration was too short: meta-analysis by Taku et al., (2011) found significant change in urinary DPD following isoflavone interventions ≥10 weeks. In addition, the equol producer subgroup (n=10, 30% of participants) was underpowered to detect a change in bone turnover markers (discussed further in the limitations section). At least 27 participants were required to detect a significant change in urinary DPD (≥1.79 nmol/mmol creatinine) and at least 20 participants were required to detect a significant change in CTx (≥0.09 nmol/L).

Tousen and colleagues (2011) report a significant decrease in urinary DPD in response to equol supplementation in non-equol producing postmenopausal Japanese women. Serum equol levels in the equol producers of this study were comparatively low and may explain the absence of an effect on bone turnover. The average serum equol obtained in the current study was 16.6 nmol/L, which is equivalent to 3.93 ng/ml, whereas in the study by Tousen et al. (2011) serum equol reached 20.7 ng/ml in equol producers. Moreover, Tousen et al. (2011) conducted a longer intervention with a larger sample size (12 months; n=23).

Finally, the women in this study ranged from 1-10 years postmenopause, and based on serum E2 levels it is estimated that participants were on average 8-10 years postmenopausal (Sowers et al., 2008). Bone turnover and bone loss is most rapid in the in early menopause (1-5 years since the onset of menopause) and isoflavone intervention has been shown to be more effective at preserving bone health in early postmenopausal women (Ma et al., 2008). The current study group may have been less responsive to an isoflavone intervention.

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