Estimation of Analgetic Effect and Cooperability of Pediatric Patients During Application of Gaseous Mixture of Oxygen:Nitrogen Suboxide 50%:50% for Minimal Invasive Procedures
Conditions: Pain, Anxiety, Sedation
Sponsor: Mother and Child Health Institute of Serbia Dr Vukan Cupic

1 
 
 
Study protocol 
 
 
Observation of clinical effects of oxygen: nitrogen 
50%:50% gas mixture application for minor invasive 
procedures in pediatric patients 
 
Correspondence 
Ana Mandras, MD PhD 
Department of Anesthesiology 
Institute for mother and child health care,  
Radoja Dakica 6-8 
11070 Belgrade,  
Serbia  
Email: ana_mandras@yahoo.com 
 
 
 
Date  
30.09.2024. 
 
 
 
 

2 
 
Summary 
Rationale 
Reducing pain and anxiety, avoiding psychological trauma associated with medical procedures 
are the focus of pediatric anesthesiologist in everyday practice. Nitrous oxide is a useful option 
for children who require procedural sedation. It is easy to apply, has satisfactory sedative and 
analgetic effects. Side effects are transient and vanish after discontinuation. It has been used for 
decades in pediatric emergency department as well for procedural sedation and analgesia.   
Objective 
The aim of this study is to evaluate the analgetic and anxiolytic profile of oxygen: nitrogen 
50%:50% gas mixture for minor invasive procedures in pediatric patients.  
Study design 
Single center prospective observational cohort study. 
Study population 
Patients from 1 to 16 years of age admitted to department of anesthesiology and intensive care 
therapy, Institute for mother and child health care. Inclusion criteria: age 1-16 years; patients 
scheduled for following procedures : venepunction, arterial line placement, nasogastric tube 
placement, urinary catheter placement, removal of thoracic drain, skin suture. Exclusion criteria, 
presence one of the following conditions: age <1 year or > 16 years; pneumothorax; 
pneumoperitoneum; pneumocephalus; otitis media; obstructive lung disease; upper airway 
infection; methylenetetrahydrofolate reductase (MTHF) deficiency; vitamin B12 deficiency; 
megaloblastic anemia; folate deficiency (Vitamin B9 deficiency); bleomycin sulphate therapy; 
neuropathy, pregnancy.  
Main study endpoints 
Primary endpoint is evaluation of pain intensity following the implemented procedure. 
Secondary endpoints are evaluation of anxiolytic effect, achieved level of sedation and 
observation of possible side effects such as hypotension, bradycardia and desaturation. 
Nature and extent of the burden and risks associated with participation, benefit and group 
relatedness  
The risks of prospective collection of demographic data, pain and anxiety scores during and after 
gas mixture application are negligible and the burden is minimal. 
 
 

3 
 
1. INTRODUCTION 
Separation of parents, anticipation of unknown procedure and pain related to it create fear and 
anxiety in children of all ages. Half of admitted children cry, fight with medical staff and panic 
when preparing for unpleasant procedures like venipunctures, bladder catheterizations, lumbar 
punctures, bone marrow aspirations, nasogastric tube placement, laceration repairs, and others. 
One of the most difficult challenges professionally and emotionally is learning to handle pain 
and stress related emotions in pediatric patients. Pain is defined by the International Association 
of the Study of Pain as “an unpleasant sensory and emotional experience associated with, or 
resembling that associated with, actual or potential tissue damage.” Expanded definition includes 
items that provide further context to the complex topic of pain: (1) pain is always a personal 
experience; (2) pain is different from nociception; (3) pain is learned through life experiences; 
(4) a person’s report of pain should be respected; (5) pain can have adverse effects on function 
and well-being; and (6) verbal description is one of several behaviors used to express pain.1 Pain 
and its’ psychological components in children are often under-recognized and undertreated. 
Children with serious medical conditions as well as healthy children have to undergo significant 
amounts of painful medical procedures throughout childhood (e.g. vaccinations, venipuncture, 
heel lance, insertion of a peripheral venous catheter, bone marrow aspirations, lumbar punctures, 
etc).  In the vast majority of infants, no analgesic strategies are employed.2 Research has shown 
that exposure to pain and stress related to it early in life has long-lasting consequences in terms 
of increases in the risk for developing problems in adulthood (chronic pain, anxiety and 
depressive disorders). Therefore, adequate management of infant and child pain is imperative.3,4 
Reducing pain and anxiety, avoiding psychological trauma associated with medical procedures 
are the focus of pediatric anesthesiologist in everyday practice. Nitrous oxide (N2O) is an 
effective sedative and analgetic agent for mildly to moderately painful pediatric procedures, 
particularly for short ones.5,6 It has been used for decades in Europe and USA for pediatric 
procedural sedation and analgesia (PSA) without serious adverse effects. European Society of 
Anesthesiology Task Force on Nitrous Oxide issued in 2019. a narrative review concerning use 
of N2O procedural sedation and analgesia (PSA). This review states that N2O can be efficiently 
and safely used for procedural sedation in pediatric population for a variety of procedures of 
light to moderate pain intensity.7 
 
 
 
 
 

4 
 
2. OBJECTIVES & HYPOTHESES 
 
2.1. Objectives 
The aims of this study are: 
- to evaluate the efficacy of oxygen: nitrogen 50%:50% gas mixture in achieving analgesia in 
pediatric patients requiring minor invasive procedures  
- to evaluate the efficacy of oxygen: nitrogen 50%:50% gas mixture in achieving desired 
sedation in pediatric patients requiring minor invasive procedures   
- to evaluate safety profile of oxygen : nitrogen 50%:50% gas mixture in achieving desired 
sedation in pediatric patients requiring  minor invasive procedures  
Minor invasive procedure investigated in this study are venepunction, arterial line placement, 
nasogastric tube placement, urinary catheter placement, removal of thoracic drain and kin suture  
2.2. Hypotheses  
- application of oxygen: nitrogen 50%:50% gas mixture will achieve pain relief in pediatric 
patients requiring minor invasive procedures   
- application of oxygen: nitrogen 50%:50% gas mixture will achieve anxiolysis in pediatric 
patients requiring minor invasive procedures 
 
3. STUDY DESIGN 
Prospective, single center observational study of a random-sample cohort of pediatric patients 
undergoing any in-hospital minor invasive procedure during a continuous 24 months period of 
recruitment starting October 2024.  
 
 
 
 
 
 

5 
 
4. STUDY POPULATION  
 
4.1. Population (base) 
Pediatric patients undergoing in hospital minor invasive procedures. 
4.2. Inclusion criteria 
Patients are eligible if: 
1 Age 1-16 years 
2 Veinpuncture 
3 Arterial line placement 
4 Nasogastric tube placement 
5 Urinary catheter placement 
6 Removal of thoracic drain 
7 Skin suture 
4.3. Exclusion criteria 
The following patients will be excluded if any of the following conditions is present 
1 Age <1 year or > 16 years 
2 Pneumothorax 
3 Pneumoperitoneum 
4 Pneumocephalus 
5 Otitis media  
6 Obstructive lung disease 
7 Upper airway infection 
8 Methylenetetrahydrofolate reductase (MTHF) deficiency 
9 Vitamin B12 deficiency 
10 Megaloblastic anemia 

6 
 
11 Folate deficiency (Vitamin B9 deficiency) 
12 Bleomycin sulphate therapy 
13 Neuropathy  
14 Pregnancy 
 
4.4. Sample size calculation 
Based on previous published data estimated number of patients required to evaluate analgetic 
and anxiolytic effect as well as safety profile is between 32 and 90.8,9,10 We aim to observe 300 
occasions that required oxygen: nitrogen 50%:50% gas mixture in order to draw unbiased 
conclusions.  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

7 
 
5. METHODS  
 
5.1. Study endpoints 
5.1.1. Primary endpoint 
Primary endpoint is evaluation of pain intensity using age specific pain scales. 
5.1.2. Secondary endpoints: 
Secondary endpoints are evaluation of cooperability of a child, anxiolysis, hemodynamic 
stability during inhalation and oxygenation as well as potential side effects. 
Clinical outcome measures:  
1 pain assessment as no/mild pain, assessed using age-based pain scales: FLACC up to 4 years, 
FACES 4-9 years and Numeric Pain Rating Scale 9-16 years;  
2 anxiolytic effect defined as child’s cooperabiliy, that is executes orders or refuses to execute 
orders;  
2 Level of sedation (desired level would be mild sedation defined as Ramsey score 2) 
3 side effects: desaturation, bradycardia, hypotension 
4 heart rate (HR), blood pressure (BP), hemoglobin oxygen saturation (SpO2); monitored before, 
during and 30 minutes post gas mixture application    
Desaturation is defined as SpO2 < 95% on room air. Hypotension, hypertension, bradycardia, 
tachycardia are defined in accordance to age.   
 
 
 
 
 
 
 
 

8 
 
6. PROTOCOL 
Application of gas mixture will be conducted via suitable face mask, flow calculated to achieve 
ventilation minute volume (MV) in accordance to age, MV (L/min) = tidal volume (Vt) x 
respiratory rate (RR).  
6.1 Management of side effects: 
1 desaturation – supplemental oxygen via face mask 
2 hypotension – iv fluids, vasoactive drugs if needed 
3 bradycardia – atropine 0,2 mg iv 
Important note – side effects such as hypotension, bradycardia are not expected to occur 
regarding good pharmacodynamic profile of nitrous oxide on cardiovascular system. However, 
implementing and monitoring these possible effects during the study increases the safety profile 
of the study  
7. STATISTICS 
Descriptive statistics will be used to describe baseline characteristics. Categorical variables will 
be expressed as n (%), continuous variables as median [IQR]. Depending on data distribution 
parametric or nonparametric test will be used to explain relationship regarding outcome 
variables.  
 
 
 
 
 
 
 
 
 
 
 

9 
 
8. SAFETY REPORTING  
8.1. Definition in matter of Pharmacovigilance 
8.1.1. Adverse event  
An adverse event is an undesirable experience occurring during the administration of a medicine 
for which the causal relationship with the administration of the medicine does not have to be 
proven. An adverse event is any unintended and undesirable signal (e.g. abnormal laboratory 
findings), symptom or disease that is time-related to the administration of a medicine. 
A serious adverse event is any undesirable reaction/event with the following results: 
Death 
Immediate life-threatening situation, 
Permanent or significant disability/incapacity, 
Requires hospitalization or prolongation of existing hospitalization, 
Is a congenial anomaly or birth defect, 
Other medically significant condition. 
8.1.2. Interaction 
Interaction is a change in the pharmacokinetic or pharmacodynamic properties of a medicine 
caused by the concurrent administration of another medicine, food, or any other substance. 
 8.1.3. Special situations: 
-  Overdose is the administration of a medicine in a single or daily dose in an amount that 
exceeds the maximum recommended individual or daily dose according to the Summary of 
Product Characteristics and Patient Information Leaflet, including cumulative effects due to 
overdose. 
-  Unauthorized administration (Off label use) of a medicine is the administration of a medicine 
with marketing authorization, that is administered in a therapeutic indication, dosage or manner 
not listed in the Summary of Product Characteristics, or not approved. 
- Abuse of a medicine is the permanent or occasional, intentional excessive use of a medicine 
followed by harmful physical or psychological effects. 
-  Medical error is any unintentional error in prescribing, issuing or administration of a medicine 
by a healthcare professional or patient. 

10 
 
-   Occupational exposure to a medicine is the work-related exposure of a person to a medicine. 
-  Misuse of a medicinal product are situations where a medicinal product is intentionally and 
inappropriately used not in accordance with the terms of the marketing authorisation 
-  Lack of efficacy is an effect which is less than the expected action of the Product 
8.2. Reporting of adverse effect and special situations 
The post authorization safety study sponsor is responsible to fulfill with its legal obligation with 
regard Pharmacovigilance. Specifically, the sponsor must identify and register the adverse 
reactions and special situations derived during the study implementation and to report the cases 
associated with suspected adverse reactions to the Medicines and Medical Devices Agency of 
Serbia (ALIMS) within the following deadlines: 
For serious adverse events: immediately and no later than 15 days from on the day of receipt of 
the first information 
For non serious adverse event: no later than 90 days from on the day of receipt of the first 
information 
The post authorization safety study sponsor will also inform the Marketing Authorisation Holder 
(MAH) involved about any adverse event or special situation occurred during the 
implementation of the present study, which is or not associated with a suspected adverse 
reaction, within 1 working day in written form from on the day of receipt of the first information. 
 
 
 
 
 
 
 
 
 
 
 

11 
 
9. ETHICAL CONSIDERATIONS 
9.1. Regulation statement 
The study will be conducted according to the principles of the Declaration of Helsinki (revision 
Fortaleza, Brazil, October 2013), with Good Clinical Practice (GCP). 
9.2. Recruitment and consent 
The investigator will enroll patients in the study. Informed consent by the patient and/or parents 
will be obtained by the investigator. 
9.3. Benefits and risks assessment, group relatedness 
This study does not result in any risk or burdens to patients. 
9.4. Incentives 
There is no financial incentive for subjects to participate in this study. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

12 
 
References  
1. Raja SN, Carr DB, Cohen M, et al. The revised International Association for the Study of 
Pain definition of pain: Concepts, challenges, and compromises. Pain 2020;161(9):1976-
82 
2.  Roofthooft DW, Simons SH, Anand KJ, Tibboel D, van Dijk M. Eight years later, are we 
still hurting newborn infants? Neonatology 2014;105(3):218-226. 
3. Victoria NC, Murphy AZ. Exposure to Early Life Pain: Long Term Consequences and 
Contributing Mechanisms. Curr Opin Behav Sci 2016;7:61-68. 
4. Brattberg G. Do pain problems in young school children persist into early adulthood? A 
13-year follow-up. Eur J Pain 2004;8(3):187-199. 
5. Amit Sethi, Sumati Bhalla, Kunal Gupta. Basic Properties of Nitrous Oxide Gas. In: 
Kunal Gupta, Dimitrios Emmanouil, Amit Sethi.editors. Nitrous Oxide in Pediatric 
Dentistry, Springer Nature Switzerland, 2020;62-74 
6. Zier JL, Liu M. Safety of high-concentration nitrous oxide by nasal mask for pediatric 
procedural sedation: experience with 7802 cases. Pediatr Emerg Care 2011;27:1107-12 
7. Wolfgang Buhre, Nicola Disma, Jan Hendrickx, Stefan DeHert, Markus W. Hollmann et 
al.  European Society of Anaesthesiology Task Force on Nitrous Oxide: a narrative 
review of its role in clinical practice. British Journal of Anaesthesia 2019;122 (5): 
587e604 
8. Ekbom K, Kalman S, Jakobsson J, Marcus C. Efficient intravenous access without 
distress: a double-blind randomized study of midazolam and nitrous oxide in children and 
adolescents. Arch Pediatr Adolesc Med 2011; 165: 785-791  
9. Jimenez A, Blazquez D, Cruz J. Use of combined transmucosal fentanyl, nitrous oxide, 
and hematoma block for fracture reduction in a pediatric emergency department. Pediatr 
Emerg Care 2012; 28: 676-679 
10. Lee JH, Kim K, Kim TY, Jo YH, Kim SH, Rhee JE, Heo CY, Eun SC. A randomized 
comparison of nitrous oxide versus intravenous ketamine for laceration repair in children. 
Pediatr Emerg Care 2012; 28: 1297-1301