Hydrogen-rich bath with nano-sized bubbles improves antioxidant capacity based on oxygen radical absorbing and inflammation levels in human serum

Abstract

This study compared the effects of hydrogen-water (HW) bath on the oxygen radical absorption-based antioxidant capacity and the inflammatory indicator, C-reactive protein (CRP), in serum between healthy volunteers and inflammatory/collagen disease-patients. The HW bath apparatus supplied nano-bubbles with a diameter of 110 ± 10 nm and 338-682 μg/L of dissolved hydrogen after 120 minutes electrolysis, and nano-bubbles increased to 9.91 × 10⁷/mL along with the increase of correlative dissolved hydrogen. Ten-minute HW bath increased the oxygen radical absorption-based antioxidant capacity to 110.9 ± 9.2% at post-bathing 120 minutes, although unaltered with 10-minute normal water bath at 40°C in healthy subjects. The CRP level was repressed to 70.2 ± 12.1% at 120 minutes after HW bath, although rather increased for normal water bath. In the patients with connective tissue diseases, the CRP level was repressed to 3-24% upon 9 days to 4 months of HW bathing. In another six patients with diverse autoimmune-related diseases, upon daily HW bathing as long as 2-25 months, the pre-bathing CRP level of 5.31 mg/dL decreased to 0.24 mg/dL being within the standard-range, with relief of visible inflammatory symptoms for some cases. Thus, the HW bath with high-density nano-bubbles has beneficial effects on serum antioxidant capacity, inflammation, and the skin appearance. The study was approved by the Committee of Ethics, Japanese Center of Anti-Aging Medical Sciences (Authorization No. H-15-03-2, on January 15, 2019), which was a non-profitable organization officially authenticated by the Hiroshima Prefecture Government of Japan.

Keywords: C-reactive protein; anti-inflammatory effect; antioxidant capacity; electrolysis; human serum; human skin; hydrogen-rich water; nano-bubble.

Figure 1A

Electrode of a hydrogen-water-producing apparatus, a photograph of its operating state and structural drawing of the electrode. Note: (A) Macroscopically visible bubbles emerging from the electrode when the apparatus Lita Life Ver. 2. is running. Scale bar: 5 cm. (B) The three-blades-composed, lattice-shaped and platinum-plated electrode.

Figure 2

Flow chart of analysis on heathy and disease volunteers. Note: For analysis of short-term (5 days) effect of HW bath, six healthy subjects were divided as three controls (regular water bath) and three HW (hydrogen-rich water bath). They were measured for ORAC and CRP. For analysis of medium term (9 days–4 months) effect of HW bath, three diseased females used the HW bath, and were measured CRP. For analysis of long term (4 months–2 years 1 month) effect of HW bath, one diseased male and five diseased females used the HW bath, and were measured CRP. Among them, one female was taken photographs of her hand and one male was taken photographs of his abdomen stoma. CRP: C-reactive protein; HW: hydrogen-rich water; ORAC: oxygen radical absorption capacity.

Figure 3

Visualization of nano-bubbles in warm tap water and hydrogen-water made via electrolysis. Note: Tap water before electrolysis has only few nano-bubbles (upper-left), whereas after 60 minutes electrolysis, hydrogen nano-bubbles emerge continuously and become detectable (other three pictures) due to the laser beam-induced halation under a microscope/video camera. Scale bar: 1 μm.

Figure 4

The concentration and size-dependent distribution of nano-bubbles in the electrolyzed water. Note: The bath water (40°C) was electrolyzed in a 200-L-poured bathtub. The increased bubbles at indicated pre-/post-electrolytic times were evaluated by Nano-Sight analysis based on tracking of laser-beam-induced scattered-light, and were regarded as hydrogen nano-bubbles.

Figure 5

Properties of hydrogen-water generated by an electrolysis apparatus. Note: (A) Dependency of mean bubble size on the electrolysis time was measured. Preparation method of the bath water was the same as that in Figure 4. (B) The relation between the number of nano-bubbles and the dissolved hydrogen concentration after 30 minutes (~150 μg/L), 60 minutes (~250 μg/L), and 120 minutes (~400 μg/L). (C) Dissolved oxygen concentration depending on the electrolysis time was measured. (D) Time course of the hydrogen nano-bubble concentration, and comparison of the instruments. Concentrations of hydrogen nano-bubbles were serially measured using the Lita Life Ver. 2 instrument and other instruments (made by A and B companies). Data are expressed as mean ± SD, and were analyzed by Student’s t-test.

Figure 6

Effects of hydrogen-rich water (HW) bath on the antioxidant capacity oxygen radical absorption-based antioxidant capacity (ORAC) in the human serum. Note: (A) The time course of ORAC values in the control group (n = 3, regular tap water bath, 40°C, 10-minute bathing). (B) The time course of ORAC values in the HW group (n = 3, hydrogen-rich water bath, 40°C, 10-minute bathing). (C) The time course of ORAC values in the control group (n = 3) and in the HW group (n = 3) are expressed as mean ± SD, and were analyzed by the Student’s t-test. *P < 0.05.

Figure 7

Effects of short-term hydrogen-rich water (HW) bath on the inflammation indicator C-reactive protein (CRP) levels in the healthy human serum. Note: (A) The time course of CRP values in the control group (n = 3, regular tap water bath, 40°C, 10-minute bathing). (B) The time course of CRP values in the HW group (n = 3, hydrogen-rich water bath, 40°C, 10-minute bathing). (C) The time course of CRP values in the control group (n = 3) and in the HW group (n = 3) are expressed as mean ± SD, and analyzed by the Student’s t-test.

Figure 8

Effects of medium-term daily-usage of hydrogen-rich water (HW) bath on CRP levels in serum of human with disease. Note: The effects of daily repeating usage of HW bath on the CRP level in the serum of patients with connective tissue disease were examined. Serum samples from three patients were obtained before and after the examination period for 9 days to 4 months. crp: c-reactive protein.

Figure 9

Effects of long-term daily-usage of hydrogen-rich water (HW) bath on the C-reactive protein (CRP) levels in the serum of patients with connective-tissue disease. Note: The effects of HW bath on the CRP levels in the human serum of six patients. In all subjects the CRP values decreased within or nearly equal to the upper limit of the standard value (0.30 mg/dL, indicated by the dotted lines).

Figure 10

Effects of long-term daily-usage of hydrogen-rich water (HW) bath on the appearance of the skin of higher-CPR patients. Note: The prominent inflammation regions on the back of right and left hands of the subject suffering from collagen disease and dermatomyositis (Table 1) became smaller with slightness (A and B). The inflammatory symptoms of another subject suffering from acute aortic dissection and enema (Table 1) before HW bathing (C, upper photograph, Inflammation) were eased after HW bathing (C, lower photograph). In C, regions of enema and immobilizer resin are also indicated.

Figure 11

Schematic illustration of oxygen radical absorption-based antioxidant capacity increase-based antioxidant activity and C-reactive protein (CRP) decrease-based anti-inflammatory activity which were concurrently executed by hydrogen-rich water bath. Note: Under inflammation and oxidative stress, the native CRP pentamer can be converted into monomeric CRP, which invades into the blood vessel and causes inflammatory response. By taking hydrogen-rich water bath, hydrogen bubbles could rapidly infiltrate into the dermis and be absorbed into the blood stream, react with the reactive oxidative species and protect cells and tissues from inflammation/ oxidative stress-induced damages.