Using electrocardiogram (ECG) and 3-axis acceleration data collected from the Allostatic State Mapping by Ambulatory ECG Repository (ALLSTAR) big Database, we analyzed the relationship between the length of sitting time and heart rate variability (HRV) indices for older adults (more than 65 y.o.). The analysis data is from 2019 to March 2021. The number of subjects for each year is 2019 (n = 23547), 2020 (n = 25402), and 2021 (n = 6205). Inclusion criteria for data analysis were sinus rhythm of 80% or higher and recording time of 80% or higher. The posture discrimination algorithm was applied to 3-axis acceleration data to calculate the ratio of sitting position during daily life. Sitting high ratio group and low ratio group were separated by the median daily sitting ratio as threshold, and compared HRV indices of both groups. HRV indices used time domain indices and frequency domain indices. As a result, R-R interval and respiratory cycle were significantly lower (P < 0.001) and frequency component power was significantly increased (P < 0.001) in the sitting high ratio group. Sitting for long time will be able to occur health risks such as myocardial infarction and cerebrovascular disease. Understanding the length of sitting time and its effect on HRV will lead to improvements in the lifestyle habits of older adults.