Recently, web-based dietary assessment tools for the targeted population have been developed and used to estimate the dietary intake level in several epidemiological studies. This study aimed to examine the validity of estimating energy and nutrient intake by the web-based 24 h dietary recall (Web24HR), which we developed for the Japanese population. Overall, 228 adults aged ≥20 years who agreed to participate were included. Web24HR was administered three times per person: twice within 3 weeks and once 3 months later. The data on 3-day weighed food records (WFR) at 3-month intervals in the four seasons were collected using the reference method. The intake of energy and nutrients between Web24HR and WFR were compared using Pearson’s correlation coefficients and the Bland–Altman analysis. As results, the correlations were moderate for both men (median r = 0.51) and women (median r = 0.38) except for iodine, retinol, retinol equivalents, and β-tocopherol. The Bland–Altman method revealed that the bias in intake was within ±10% for most nutrients, except for cholesterol, iodine, vitamin C, and the water content, in both sexes. Additionally, monounsaturated fatty acids in men and β-cryptoxanthin in women exhibited an underestimation of more than 10%. In conclusion, the Web24HR intake assessment showed moderate correlations for most nutrients in both sexes. The bias in intake was within ±10% for most nutrients, but there were discrepancies for some nutrients. This tool’s performance is comparable to Japan’s standard dietary exposure assessment methods and will be helpful for future applications in epidemiological studies, though caution is needed for certain nutrient assessments.
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