Oxyhydrogen Nanobubble Infusion as a Complementary Therapy in Patients With Parkinsonism
Phase: PHASE2
Conditions: Parkinson Disease
Sponsor: dr. Siti Nurlaela, SpN

 
 
 
Research Protocol 
 
 
Oxyhydrogen Nanobubble Infusion as a Complementary Therapy in Patients with 
Parkinsonism 
 
 
Ethical Approval Letter No. 291/EC/KEPK/08/2024 
 
 
 
 
Nurlaela S*, Sumitro SB **, Sabarudin A**, Marhaendroputro EA***, Hernowo AT**, 
Sulastri EE**, Widayati A*, Harbiyanti NT*, Sari AN*, Nanda CF****, Hannan H**** 
 
*Neurology department of Brawijaya University Hospital, Malang, East Java, 
Indonesia 
**Indonesia Molecule Institute, Malang, East Java, Indonesia 
***Neurology department of Saiful Anwar General Hospital, Faculty of Medicine of 
Brawijaya University, Malang, East Java, Indonesia 
**** General practitioner of Brawijaya University Hospital, Malang, East Java, 
Indonesia 
 
 
 
 
 
Brawijaya University Hospital 
18 August 2024 
 

 
 
 
Background 
This study was conducted at Brawijaya University Hospital in Malang, Indonesia, involving 35 
patients with parkinsonism. The study used a prospective cohort study design. The effects of 
oxyhydrogen nanobubble (OHNB) infusion on the subjects were observed without comparison to 
a control group (placebo). Pre- and post-test results were evaluated before and after the OHNB 
infusion. 
The variables analyzed in this study were the degree of parkinsonism using the Hoehn and Yahr 
scale, cognitive function using the MOCA-Ina test, and quality of life for patients with parkinsonism 
using the PDQ-39 questionnaire. 
The hypothesis of this study is that OHNB infusion can improve disease severity, cognitive 
function, and quality of life in patients with parkinsonism.. 
OHNB is an infusion that containing H2 and O2 gas molecules in the form of nanobubbles. H2 
and O2 gas molecules encapsulated in nanobubbles are more stable and can persist for longer 
periods in solution (Li et al., 2022) and more easily penetrate the blood-brain barrier (Owen et al., 
2016; Alghamdi et al., 2022). H2 nanobubbles have superior antioxidant properties than H2 water 
alone (Kato et al., 2015). H2 have anti-inflammatory effects, thus reducing cellular inflammation 
(Cheng et al., 2023), reducing oxidative stress and protecting the brain from ischemic injury 
(Ohsawa et al., 2007), mitigating acute neurotoxic effects, and addressing behavioral disorders 
caused by MPTP exposure on dopaminergic neurons (Fujita et al., 2009). O2 is required to 
activate the HIF-1α pathway which can maintain neuronal integrity.(Pinilla et al, 2021) 
 
Study Purpose 
The aim of this study was  to determine the effect of oxyhydrogen nanobubble infusion on the 
degree of disease, cognitive function, and quality of life of patients with parkinsonism. 
 
Study Design 
The study used a prospective cohort study design. The effects of oxyhydrogen nanobubble 
(OHNB) infusion on the subjects were observed without comparison to a control group 
(placebo). Pre- and post-test results were evaluated before and after the OHNB infusion. 
. 
 

 
 
 
Inclusion Criteria 
• 
Patients who meet the criteria for parkinsonism 
• 
18-70 years old 
 
Exclusion Criteria 
• 
Patient is unwilling to undergo examination. 
• 
Pregnancy. 
• 
History of malignancy and/or chemotherapy. 
• 
History of alcohol or drug use that can induce extrapyramidal symptoms. 
• 
Hyperthyroidism. 
• 
Impaired kidney function. 
• 
Impaired liver function. 
• 
Patients with heart failure requiring fluid restriction. 
• 
MRI results suggest a brain tumor causing parkinsonism symptoms. 
• 
Catalase enzyme deficiency (acatalasemia). 
 
Study Protocol 
In the initial phase of the study, patients with Parkinsonism underwent a brief history taking to 
determine whether they met the inclusion criteria. If the patient met the inclusion criteria, informed 
consent was obtained. 
Next, the patient was evaluated to determine whether they met the exclusion criteria, including 
pregnancy, a history of malignancy and/or chemotherapy, and a history of drug or alcohol use 
that could induce extrapyramidal syndrome symptoms. If the exclusion criteria were met, the 
patient was excluded from the study. 
Patients included in this study then underwent a complete physical examination (internal status 
and neurological status), complete blood count, SGOT/SGPT, Urea/Creatinine, random blood 
sugar, blood electrolyte levels, thyroid function, chest x-ray, ECG, and head MRI. If exclusion 
criteria were found, namely hyperthyroidism, impaired heart function  requiring fluid restriction, 
impaired liver function (increased SGOT/SGPT > 1.5 x normal), impaired kidney function 
(creatinine > 1.5), or head imaging results showed a brain tumor, then the patient was excluded 
from the study. This initial pre-treatment data was also used as the baseline data for the study 
subjects. 

 
 
 
During the blood draw, a catalase enzyme test was also performed by spraying hydroxypropyl 
methylcellulose onto the patient's blood on a glass slide. If bubbles/foam appear in the patient's 
blood upon application, it indicates the presence of catalase. However, if no bubbles/foam appear, 
it indicates the patient has acathalasemia and is excluded from the study. 
Patients who meet the inclusion criteria and do not meet the exclusion criteria will then undergo 
a parkinsonism severity assessment based on the Hoehn and Yahr scale, the MoCA-Ina test, and 
the PDQ 39 questionnaire as pre-treatment data. 
Patients will then receive 12 oxyhydrogen nanobubble infusions, administered twice weekly. 
Evaluation of the effects of the OHNB infusions will be conducted twice: after the sixth infusion 
(following the 15 ml dose) and after the 12th infusion. Patients will undergo a complete physical 
examination (internal and neurological status), complete blood count (CBC), SGOT/SGPT, 
Urea/Creatinine, random blood sugar, blood electrolyte levels, parkinsonism severity based on 
the Hoehn and Yahr scale, the MoCA-Ina test, and the PDQ 39 questionnaire as post-treatment 
data. 
 
Data Analysis 
Data regarding age, gender, duration of parkinsonism symptoms, types of medications 
consumed, comorbidities (hypertension, coronary heart disease, metabolic syndrome), degree of 
parkinsonism according to the Hoehr and Yahr classification, cognitive function according to the 
MoCA-Ina examination, and PDQ 38 examination results in this study will be presented in a 
frequency distribution table. Then, the table will be analyzed descriptively. 
Data regarding parkinsonism severity examination based on Hoehn and Yahn scale, MoCA-Ina 
examination, and PDQ 39 questionnaire examination PDQ-39 pre and post treatment are 
numerical data consisting of three groups of data, namely pre-treatment data, evaluation data 
after the 7th injection (mid-treatment data), and evaluation data after the 12th injection (post-
treatment data). will be analyzed statistically using one-way ANOVA. The data will then be 
analyzed whether they are normally distributed or not. If the data are normally distributed, they 
will be analyzed using Repeated ANOVA Test, and if they are not normally distributed, they will 
be analyzed using Friedman Test. 
 
 

 
 
 
Expected Results 
Oxyhydrogen nanobubble infusion can improve disease severity, cognitive function, and quality 
of life in patients with parkinsonism. 
 
Ethical Considerations  
This research upholds research ethics so that it can provide values that benefit the wider 
community. The results of this study are expected to have a positive impact on the Indonesian 
health system, making it more independent and accessible to the people. However, a potential 
risk posed by this study is the possibility of a healing crisis, in which participants experience a 
decline in physical condition such as fever, chills, and pain. After experiencing a healing crisis, 
participants can experience the benefits of oxyhydrogen nanobubble infusion, such as feeling 
much healthier, more energetic, and refreshed than before the infusion. This risk can be managed 
with the guidance and prescription of medication provided by a doctor, thereby minimizing the 
occurrence and severity of a healing crisis. 
 
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Mitochondria-Targeting Nutrient via Activating Keap1-Nrf2 Antioxidant System. 
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