**1. Introduction**

Stress is a general term for the defense response to external stimuli, such as pain, cold, and infection, and can include mental tension and worry [1]. Recently, there has been considerable interest in the effects of psychological stress on the body, particularly in light of the discovery of a neural pathway that transmits stress signals from specific areas of the cerebral cortex, which processes psychological stress and emotions, to the hypothalamus, which controls the sympathetic nervous system [2]. The human body is constantly exposed to stress and works to maintain a constant internal environment via the control of the autonomic nervous, endocrine, and immune systems for protection [3]. However, excessive stress weakens the body's stress resistance, resulting in physical and psychological symptoms, including a deterioration in the quality of sleep and reduced work efficiency [4].

The autonomic nervous system, which includes the sympathetic and parasympathetic nervous systems, plays several important roles in the regulation of biological functions, such as respiration/circulation, digestion/absorption, secretion, and metabolism in a coordinated manner to maintain homeostasis [5]. For example, an imbalance in the autonomic nervous system may prevent the activation of the sympathetic nervous system, even during activities when it is typically activated, or the activation of the parasympathetic nervous system, even during recovery and rest. These conditions

lead to an inability to concentrate and various ailments, such as poor sleep quality, reduced immunity, poor bowel movements, as well as various diseases [5]. Eliminating the underlying stress is an e ffective approach to balancing the autonomic nervous system. However, adjustments in lifestyle and nutrition are also e ffective.

*Euglena gracilis* (*Euglena*) is a single-celled microalga with plant and animal properties. *Euglena* consists of various nutrients, such as vitamins, minerals, amino acids, and fatty acids, and are used as nutritional and general supplements [6]. Previous studies have confirmed that *Euglena* intake e ffectively suppresses elevated blood glucose levels [7], suppresses fat accumulation [8,9], attenuates lifestyle-related disease symptoms [10], promotes immune function [11,12], and improves bowel movements [13]. Autonomic nervous system activity plays an important role in maintaining homeostasis and is involved in biological processes such as lipolysis, blood glucose regulation, immune function, and digestion. For example, the autonomic nervous system of the pancreas contributes to the regulation of blood glucose levels by regulating insulin secretion [14]; in white adipose tissue the autonomic nervous system is involved in lipolysis [14,15]; in the spleen, it is involved in immunity [16,17], and in the stomach and intestines it is involved in the promotion of peristaltic movements [18]. These findings led us to hypothesize that various beneficial health e ffects of *Euglena* intake are mediated by its regulation of the autonomic nervous system balance.

In this study, we investigated the e ffects of the ingestion of *Euglena*-containing food for 12 weeks on the autonomic nervous system, psychological factors, and quality of sleep in men and women aged 20 to 64 years su ffering from decreased motivation and a decline in sleep quality.

#### **2. Materials and Methods**
