Study of Drug Exposure in Systemic Circulation of Primatene Mist (0.25mg) by Oral Inhalation, Versus Epinephrine Injection (0.30mg) by Intramuscular Injection and ProAir (0.18mg) by Oral Inhalation in Healthy Individuals: A Randomized, Safety Evaluator-blind, Three-Treatment, Crossover, Fasting Study
Phase: PHASE4
Conditions: Pharmacokinetics, Epinephrine, Albuterol, Asthma, Anaphylaxis
Sponsor: Amphastar Pharmaceuticals, Inc.



Amphastar Pharmaceuticals, Inc. 
 
Protocol No. API-E004-CL-I 
Primatene Mist, Epinephrine HFA MDI (E004)                                         
Version 1.1 (11/ 08 /2019) 
 
 
 
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SYNOPSIS 
 
General Information 
 
 
 
(1) Protocol No.  
 
 
API-E004-CL-I 
            (2) Title                                               Study of Drug Exposure in Systemic Circulation of 
Primatene® Mist (0.25mg) by Oral Inhalation, versus 
Epinephrine Injection (0.30mg) by IM and ProAir® 
(0.18mg) by Oral Inhalation, in Healthy Individuals 
(3) Subtitle 
A Randomized, Safety Evaluator-blind, Three-treatment, 
Crossover, Fasting Study  
(4) Study Drug  
Primatene® Mist (E004), Epinephrine HFA MDI Inhalation 
Aerosol 0.125mg/inhalation, NDA 205920 
(5) Comparator Treatments 
Epinephrine Injection (Generic of EpiPen®), Auto-Injector, 
0.3 mg by 
 
 
ProAir® HFA MDI Inhalation Aerosol by 
 
 (0.090mg/inhalation)  
(6) Delivery Path    
 
 
Oral Inhalation (E004, ProAir)  
 
 
 
 
Intramuscular Injection (Epinephrine Injection) 
(7) Manufacturer  
 
 
  
(8) Sponsor 
 
  
 
Amphastar Pharmaceuticals, Inc.  
(9) Objectives 
To assess the drug exposure profile in systemic circulation 
of Primatene® Mist by inhalation, versus Epinephrine by 
IM and ProAir® HFA by inhalation in healthy adults 
 
(10) Phase                   
 
IV 
 
 
I. 
Study Design 
 
This is a randomized, safety evaluator-blind, three-treatment, crossover, fasting study in healthy 
adults (18 - 50 years of age). The design includes a Screening Visit, three Dosing Visits and a 
follow-up phone evaluation. All study subjects must be properly consented, and screened against 
the inclusion and exclusion criteria prior to randomization.   
 
1) 
Screening Visit (3 – 7 days before Visit-1):  
Prospective subjects will sign informed consent and be assigned a screening ID, EIS-mm, 
where EI represents “E004-CL-I Study”, S stands for screening, and mm is the screening 
sequential number. Subjects will be screened per inclusion/exclusion criteria. The 

Amphastar Pharmaceuticals, Inc. 
 
Protocol No. API-E004-CL-I 
Primatene Mist, Epinephrine HFA MDI (E004)                                         
Version 1.1 (11/ 08 /2019) 
 
 
 
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following screening activities will be conducted: (i) physical examination (PE), (ii) 
measurements of vital signs, (iii) ECG, (iv) urinalysis (UA), (v) urine drug screen, (vi) 
alcohol screen (urine or breathalyzer), (vii) urine or serum pregnancy test (all female 
subjects), and (viii) fasting (≥ 8 hours) blood tests [comprehensive metabolic panel, 
complete blood count (CBC) with differentials, thyroid function tests, HIV, hepatitis B and 
C tests]. 
 
The subject’s recent medication history will include a review for any use of medicines as 
listed in the prohibited drug list, Appendix III. Qualified subjects will be scheduled for 
Dosing Visit. 
 
Per FDA guidance for Metered Dose Inhaler (MDI), each subject will be trained on the 
correct self-administration of MDI using placebo, a MDI training unit that contains no 
active drug, to assure a relatively consistent inspiratory flow rate and inspiratory duration [1]. 
A video on how to use the MDI will be provided at each training session at Screening and 
dosing visits. Subjects will be retrained on the use of the MDI at each dosing visit. 
Vitalograph AIM™ (Aerosol Inhalation Monitor) will be used to monitor the duration of the 
inhalation during the training at Screening and the dosing visit. In order to be enrolled into 
the study, subject must demonstrate proficiency in the use of MDI after training.  
 
2) 
Dosing Visits (separated by 2 to 7 day between the three visits):   
Subject qualification will be confirmed at the beginning of the Dosing Visit. After satisfying 
the enrollment criteria, each subject will be assigned a unique study subject ID as EIPK-nn, 
where nn is the sequential number based on the order of enrollment.  
 
At each dosing visit, subjects will either self-administer the MDI inhaler provided 
(Primatene Mist for Treatment-A, or ProAir HFA for Treatment-C), or receive an IM 
injection (0.30mg Epinephrine injection for Treatment-B)  
 
Fasting (≥ 10 hours) is required prior to study drug dosing. Meals will be served after the 4 
hour post-dose PK sampling point and after completion of End-of-Study (EOS) blood draw. 
 
Record concomitant drugs or ongoing medications since the Screening Visit, and ensure that 
all prohibited drugs washout have been met. 
 
One (1) baseline and twenty-five (25) serial post-dose PK blood samples will be collected 
for PK analysis.  
 

Amphastar Pharmaceuticals, Inc. 
 
Protocol No. API-E004-CL-I 
Primatene Mist, Epinephrine HFA MDI (E004)                                         
Version 1.1 (11/ 08 /2019) 
 
 
 
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At each Dosing Visit, vital signs will be measured at baseline, 5±2 min, 15±5 min, 60±5 
min, 120±15 min and 24 ±2 hrs after dosing. ECG will be measured at baseline, 20±5 min, 
120±15 min and 24 ±2 hrs after dosing. 
 
Serum potassium and glucose levels will be measured at baseline, 10±2 min, 60±5 min and 
24 ± 2 hrs post-dose. 
 
An EOS evaluation will be performed, which includes reviews and collection of vital signs, 
ECG, clinical lab tests (≥8 hrs fasting metabolic panel, CBC with differentials, and UA), 
and PE. 
 
3) 
A Follow-up Evaluation:  
Subjects who experienced any serious adverse event (SAE) or other adverse events (AE) 
that requires follow-up will remain under medical supervision until the AE is resolved or 
stabilized. A follow-up phone call will be made to all subjects within 7 days after the last 
dosing visit, or at subject discontinuation from the study.  
 
 
II. 
Study Subjects 
A sufficient number of candidates will be screened and enrolled to target approximately 24 
evaluable subjects.  
 
All subjects will be enrolled based on the following inclusion/exclusion criteria. 
 
Inclusion Criteria: 
(1) 
Agree to participate; understand and sign informed consent; 
(2) 
Male and female adults, aged 18 to 50 years, inclusive at Screening;  
(3) 
Generally healthy and medically stable, with no clinically significant abnormalities 
based on physical examination and laboratory tests as determined by the Investigators; 
(4) 
Have good venous access; 
(5) 
Have normal resting blood pressure and normal heart rate (HR) without history of 
syncope; a subject with out of range blood pressure may be enrolled in the study at the 
discretion of the Principal Investigator;  
(6) 
Have a body mass index (BMI) of 18.0 – 30.0 kg/m2;  
(7) 
Female candidates must be >1 year post-menopausal or practicing a clinically 
acceptable form of birth control and confirmed by negative urine or serum pregnancy 
test at Screening;  
(8) 
Negative HIV-Ab, HBs-Ag and HCV-Ab; 
(9) 
Negative alcohol test (urine or breathalyzer); 
(10) Negative drug screening results;  

Amphastar Pharmaceuticals, Inc. 
 
Protocol No. API-E004-CL-I 
Primatene Mist, Epinephrine HFA MDI (E004)                                         
Version 1.1 (11/ 08 /2019) 
 
 
 
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(11) Currently non-smoker; have not used any tobacco products for at least three (3) months 
prior to Screening; and 
(12) Demonstrate proficiency in the use of MDI and a consistent inhalation time >2.0 
seconds after training, for at least three (3) times, with a maximum of 5 attempts.  
 
Exclusion Criteria: 
1) Concurrent clinically significant cardiovascular, hematological, renal, neurologic, 
hepatic, endocrine, psychiatric, or malignant diseases. 
2) Known intolerance or hypersensitivity to any component of the study drugs (i.e., 
Epinephrine, Albuterol Sulfate or any sympathomimetic drugs, HFA-134a, thymol, 
ethanol, ascorbic acid, nitric acid, and hydrochloric acid). 
3) Upper or lower respiratory tract infection, or other systemic infection within 6 weeks 
prior to Screening; 
4) Clinically significant abnormalities in the screening/baseline ECG; prolonged corrected 
QT interval (QTcF) on ECG: men >450ms, women: >470ms; single or multiple 
premature ventricular contractions (PVC); 
5) Abnormal thyroid function test (if TSH is out of range, refer to T3/T4 for thyroid 
function assessment); 
6) Subject has been on other investigational drug/device studies within 30 days of Screening 
Visit or planned participation in another investigational drug trial at any time during this 
trial; 
7) Women who are pregnant or lactating or planning a pregnancy during the study period; 
8) Subject has donated or lost > 500 mL of blood within 3 months of Screening;  
9) Evidence of alcohol or drug abuse or dependency within 6 months prior to screening; or 
10) Use of any of the prohibited drugs listed in Appendix III without appropriate washout.  
 
 
III. 
Study Drugs and Dosing Regimens  
 
Qualified subjects will participate in three (3) dosing visits separated by a period of 2 – 7 days. 
Each subject will receive one study treatment at each dosing visit per the randomized sequences, 
as described in Table A. 
 
MDIs will be dispensed by the designated site professional, and the correctness of the initial dose 
indicator mark shall be verified prior to dispensing. The reading on the dose indicator of each 
MDI unit will be recorded prior to and after dosing in the study source documents. IM injection 
will be dispensed using aseptic techniques by the designated site personnel.  
 
Due to the difference in dosing position for IM and oral inhalation, and to preserve blinding from 
evaluators, subjects will be escorted to a separated dosing area for study drug administration. 

Amphastar Pharmaceuticals, Inc. 
 
Protocol No. API-E004-CL-I 
Primatene Mist, Epinephrine HFA MDI (E004)                                         
Version 1.1 (11/ 08 /2019) 
 
 
 
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Table A.  Study Arms and Treatments 
 
 
 
Subjects will be retrained on the use of the MDI at each Dosing Visit. Vitalograph AIM™ 
(Aerosol Inhalation Monitor) will be used to monitor and measure the duration of the inhalation 
during the training at Screening and prior to dosing. 
 
Each MDI must be primed at the study site prior to the first use by releasing 4 test sprays (E004 
for Treatment-A), or 3 test sprays (ProAir for Treatment-C) into the air away from the face, 
shake the inhaler well before each spray. Priming will be performed in an area that is well 
ventilated (preferably a fume hood, or an open space), and physically separated from the study 
evaluation area.  
 
Subjects will self-administer two (2) inhalations (within 30 seconds interval between each 
inhalation) of E004 for Treatment-A, or two (2) inhalation of ProAir for Treatment-C. The study 
drug administration process will also be recorded, and the video will be used to investigate 
apparent irregularities. Time of the start of each inhalation will be recorded. The used MDI 
inhaler will be collected at the end of each Dosing Visit.  
 
IM injection (0.30mg of epinephrine solution in 0.30 mL) for Treatment-B will be given on the 
thigh. Subjects will be lying down in a supine position. The 0.30 mL dose will be given 
perpendicularly as a single deep IM injection into anterolateral aspect of the thigh (vastus 
lateralis site). Epinephrine injection (Treatment B) will be administered as follows [2]: 
 
1) Doser or designated site personnel will first pull off the blue safety release to unlock the 
auto injector.  
2) Subject will hold the leg firmly in place.  
Items
Treatment-A
Treatment-B
Treatment-C
Study Drug Name
Primatene Mist, E004
Epinephrine Injection Auto-
Injector (Generic of EpiPen®)
ProAir
Manufacturer
API
Epinephrine
Epinephrine
Albuterol Sulfate
Dosage Form
Microcrystalline Suspension
HFA MDI
Sterile Solution
Microcrystalline Suspension
HFA MDI
Strength
0.125 mg/inh
0.3mg/0.3mL
0.09 mg/inh
Dose Regimen
      2 inhalations, 0.25mg   
IM Injection of 0.30 mg 
Epinephrine in 0.30 mL            2 inhalations, 0.180 mg   
Route of Administration
Oral Inhalation
 IM Injection
Oral Inhalation

Amphastar Pharmaceuticals, Inc. 
 
Protocol No. API-E004-CL-I 
Primatene Mist, Epinephrine HFA MDI (E004)                                         
Version 1.1 (11/ 08 /2019) 
 
 
 
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3) Doser will place the orange tip against the middle of the outer thigh (upper leg) at a right 
angle (perpendicular) to the thigh. 
4) Swing and push the injector firmly until it ‘clicks.’ The click signals that the injection has 
started. 
5) Hold firmly in place for 3 seconds.  
6) Remove the injector from the thigh and massage (apply gentle pressure) with dry gauze 
to the injection area for 10 seconds. The orange tip automatically extends out after use. 
7) The injection site will be observed for any medication leakage for approximately 30 
seconds after the dosing is completed. If there is observed leakage, PK samples will not 
be collected, subject will be early terminated (ET). 
 
The study medication administration should be conducted between 07:00 and 11:00 AM on the 
Dosing Visit. 
                   
IV. 
PK Blood Sampling  
 
1) 
Sampling Schedules and Time Window  
A total of twenty-six (26) PK blood samples will be collected from each Dosing Visit. The blood 
sampling schedule and allowed time window are listed in Table B.  
 
Table B. PK Blood Sampling Schedule (~5 mL per sample) 
 
 
 
Seq. 
#
Time window
Sample 
No.,"XX"
Seq. 
#
Time window
Sample 
No.,"XX"
1
within 30 min 
pre-dosing
01
14
40 min
14
2
1 min
02
15
50 min
15
3
2 min
03
16
60 min
16
4
3 min
04
17
70 min
17
5
5 min
05
18
80 min
18
6
7 min
±1 min
06
19
90 min
19
7
9 min
07
20
120 min
20
8
12 min
08
21
4 hrs
21
9
15 min
09
22
6 hrs
22
10
18 min
10
23
8 hrs
23
11
21 min
11
24
12 hrs
24
12
25 min
12
25
18 hrs
±20 min
25
13
30 min
13
26
24 hrs
26
±2 min
±10 min
±15 min
±5 min
±1 min
± 0.5 min
Scheduling
Baseline 
Scheduling

Amphastar Pharmaceuticals, Inc. 
 
Protocol No. API-E004-CL-I 
Primatene Mist, Epinephrine HFA MDI (E004)                                         
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One (1) pre-dose baseline sample will be collected within 30 minutes before dosing. Time 0 is 
defined as the beginning of the last inhalation of the MDI, or at the completion of IM 
administration. 
 
 
2) 
Sampling Procedure  
A catheter will be inserted into one of the antecubital region veins for blood sampling. 
Alternatively, if there is a clot in the catheter, collection from a vein of the hand or forearm for 
easier access or via venipuncture is also acceptable. After each blood sample collection, a 
catheter flush will be performed to avoid catheter clot formation and prevent interference 
between samples.   
 
At each sampling point, the first 1 mL of blood will be discarded for waste (utilizing a disposable 
tube), and blood samples (~5 mL) will be collected in ice-chilled potassium-EDTA sample tubes, 
mixed well immediately following collection, kept on ice or refrigerated and centrifuged within 
20 minutes of collection. All blood samples will be centrifuged at 4°C, 2,000-3,000 g for 20 
minutes for plasma isolation. Isolated plasma will be distributed into two (2) aliquots, 
approximately 1.5 mL in the first aliquot and the remainder in the second aliquot. The plasma 
sample vials will be frozen immediately on dry ice and then stored (within 60 minutes following 
plasma separation) in a freezer at -20C or lower until analysis. 
 
 
3) 
PK Sample ID  
 
The sample ID for PK blood samples (before centrifugation) is defined as:    
      
                                                            “EIPK-nn-V-XX” 
 
where EIPK-nn is the subject ID, V is the dosing visit code, V=1 to 3, and XX is the two-digit 
time-point sequential number of the sample: 01, 02, 03, … 26 as listed in Table B. 
 
The sample ID for PK plasma samples (after centrifugation) is defined as “EIPK-nn-V-XX-k” 
and k is the aliquot number (A or B). The PK plasma sample ID labels, with the size of 
1/2”×1¾” (Avery label 5667), should be printed in blue font as demonstrated below: 
 
            
 
 
4) 
Sample Transportation and Analysis 
 
The chain of custody, temperature control, and related documents for sample transportation must 
EIPK-nn-V-XX-k 

Amphastar Pharmaceuticals, Inc. 
 
Protocol No. API-E004-CL-I 
Primatene Mist, Epinephrine HFA MDI (E004)                                         
Version 1.1 (11/ 08 /2019) 
 
 
 
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be recorded. During transportation of PK samples from the site to Amphastar, the temperature 
should be below -20ºC. PK samples will be analyzed with an established LC/MS/MS method for 
detection of Epinephrine and Albuterol. 
 
V. 
Study Evaluations 
 
1) Primary Endpoints 
 
AUC0-6h, defined as area under the curve (AUC) concentration curve over time, 
from time 0 to 6 hours post-dose for epinephrine; 
 
AUC0-24h, defined AUC of plasma Albuterol concentration curve over time 0 to 24 
hours post-dose; 
 
AUC0-∞, defined as AUC in the plot of plasma Albuterol, or Epinephrine versus 
time from time 0 to infinity; and 
 
Cmax, defined as the maximum plasma concentration of Albuterol, or Epinephrine. 
 
2) Secondary Endpoints 
 
tmax of Albuterol, or Epinephrine, defined as the time, at which the Cmax is observed; 
and 
 
T1/2, terminal elimination half-life of each analyte. 
 
3) Additional Evaluations 
 
Relative Bioavailability of Treatment-A (E004 inhalation) over Treatment-B (IM 
epinephrine injection); 
 
Epinephrine concentrations versus time; and 
 
The mean and standard deviation of study endpoints will be estimated. The 
variances within and between subjects will be estimated. 
 
4) Statistical Analysis: 
All endpoint will be listed and compared between treatments. 
 
 
VI. 
Safety Evaluations 
 
1) Monitor changes in vital signs and ECG will be analyzed between Screening, designated 
post-dose time points and EOS;  
2) Heart rate, and systolic and diastolic blood pressure will be measured at baseline, 5±2 min, 
15±5 min, 60+5 min, 120±15 min and 24 ±2 hours after dosing; 
3) Monitor serum potassium and glucose at study baseline, designated post-dose time points and 
EOS; 
4) Physical examinations at Screening, and at the end of the last Dosing Visit, or study 
termination; 

Amphastar Pharmaceuticals, Inc. 
 
Protocol No. API-E004-CL-I 
Primatene Mist, Epinephrine HFA MDI (E004)                                         
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5) The results of laboratory tests, including CBC (with differential), metabolic panel and UA, 
will be assessed and compared between data at Screening and that at EOS; 
6) All subjects will be queried for ADEs. All ADEs must be recorded and assessed. 
 
 

Amphastar Pharmaceuticals, Inc. 
 
Protocol No. API-E004-CL-I 
Primatene Mist, Epinephrine HFA MDI (E004)                                         
Version 1.1 (11/ 08 /2019) 
 
 
 
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Table of Contents 
 
1. INTRODUCTION AND RATIONALE ................................................................................ 14 
1.1. 
Introduction of the Drug Product ..................................................................................... 14 
1.2. 
Mechanism of Action ...................................................................................................... 15 
1.3. 
Indications and Dose ....................................................................................................... 15 
1.4. 
Pharmacokinetics (PK) and Pharmacodynamics (PD) Profile ........................................ 15 
1.5. 
Contraindications, Warnings, Precautions, and Adverse Reactions ................................ 17 
1.5.1. 
Contraindications ......................................................................................................... 17 
1.5.2. 
Warnings and Precautions............................................................................................ 17 
1.5.3. 
Adverse Reactions ....................................................................................................... 18 
1.6. 
Drugs with Strong Impact on PK or PD .......................................................................... 18 
2. OBJECTIVES ......................................................................................................................... 20 
2.1. 
Objective .......................................................................................................................... 20 
2.2. 
Primary Endpoints ........................................................................................................... 20 
2.3. 
Secondary Endpoint ......................................................................................................... 20 
3. INVESTIGATORS AND STUDY SITES ............................................................................. 21 
3.1. 
Principal Investigator(s) .................................................................................................. 21 
3.2. 
Investigational Site(s) ...................................................................................................... 21 
3.3. 
Laboratory ....................................................................................................................... 21 
4. MATERIALS AND METHODS ........................................................................................... 22 
4.1. 
Study Design.................................................................................................................... 22 
4.2. 
Study Subjects ................................................................................................................. 22 
4.2.1. 
Sample Size .................................................................................................................. 22 
4.2.2. 
Subject Enrollment....................................................................................................... 22 
4.2.3. 
Inclusion Criteria: ........................................................................................................ 22 
4.2.4. 
Exclusion Criteria ........................................................................................................ 23 
4.3. 
Randomization Procedures .............................................................................................. 23 
4.3.1. 
Subject Identification Assignment ............................................................................... 23 
4.3.2. 
Subject Randomization ................................................................................................ 24 
4.4. 
Dispensing, Returning, and Storage of Material and Drug Supplies ............................... 24 
4.5. 
Study Drugs Administration and Training ...................................................................... 25 
4.5.1. 
Training for Correct Use of MDI Inhaler .................................................................... 25 
4.5.2. 
Preparation and Training of MDIs at the Dosing Visit ................................................ 25 
4.5.3. 
Detailed MDI Dosing Procedure ................................................................................. 26 

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Protocol No. API-E004-CL-I 
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4.5.4. 
IM Injection Procedure ................................................................................................ 27 
4.6. 
Blinding Methodology and Procedure ............................................................................. 27 
4.7. 
Unblinding ....................................................................................................................... 27 
5. STUDY PROCEDURE ........................................................................................................... 29 
5.1. 
Global Plan ...................................................................................................................... 29 
5.2. 
Screening Procedure and Scheduling .............................................................................. 29 
5.2.1. 
Schedule and Purpose of Screening Visit .................................................................... 29 
5.2.2. 
Informed Consent Form ............................................................................................... 29 
5.2.3. 
Medical History, PE, and General Safety (Screening Activities) ................................ 29 
5.2.4. 
Instruction to Subjects at the End of Screening Visit .................................................. 30 
5.3. 
Dosing Visits ................................................................................................................... 30 
5.3.1. 
General Profile for Dosing Visits ................................................................................ 30 
5.3.2. 
Pre-dose Activities ....................................................................................................... 31 
5.3.3. 
Instructions to the Subjects before Dosing at Dosing Visit ......................................... 31 
5.3.4. 
Study Medication Administration ................................................................................ 32 
5.3.5. 
PK Blood Sampling ..................................................................................................... 32 
5.3.6. 
PK Blood Sample ID and Labeling ............................................................................. 34 
5.3.7. 
Safety Evaluations/Monitoring .................................................................................... 34 
5.4. 
Activity at End of Study (EOS) ....................................................................................... 34 
5.5. 
Follow-up Activities ........................................................................................................ 35 
5.6. 
PK Sample Transfer and Laboratory Analysis ................................................................ 35 
5.7. 
Summary and Schema of Study Activities ...................................................................... 35 
5.7.1. 
Data collection ............................................................................................................. 36 
5.7.2. 
Schema of Study Activities .......................................................................................... 36 
5.8. 
Chart for Study Activities ................................................................................................ 38 
6. STATISTICAL ANALYSIS ................................................................................................... 39 
6.1. 
Populations ...................................................................................................................... 39 
6.1.1. 
Per-Protocol Population (PPP) ..................................................................................... 39 
6.1.2. 
Treated Population (TP) ............................................................................................... 39 
6.1.3. 
Intent-to-treat Population (ITT) ................................................................................... 39 
6.2. 
PK Results Evaluations ................................................................................................... 40 
6.2.1 
Calculation of Primary Endpoints ................................................................................ 40 
6.2.2 
Calculation of Secondary Endpoints ............................................................................ 40 
6.2.3 
Missing Data Handling................................................................................................. 40 
6.2.4 
Relative Bioavailability ................................................................................................ 40 
6.3. 
Safety Assessment ........................................................................................................... 40 
6.4. 
Interim Analyses .............................................................................................................. 41 

Amphastar Pharmaceuticals, Inc. 
 
Protocol No. API-E004-CL-I 
Primatene Mist, Epinephrine HFA MDI (E004)                                         
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7. STUDY MANAGEMENT ...................................................................................................... 42 
7.1. 
Discontinued Subjects or Withdrawal ............................................................................. 42 
7.2. 
Adverse Events ................................................................................................................ 42 
7.2.1. 
Types of Adverse Events ............................................................................................. 42 
7.2.2. 
Definition of Serious and Unexpected Adverse Events ............................................... 43 
7.2.3. 
Obligations of AE Reporting to Sites, FDA and IRB .................................................. 44 
7.2.4. 
Follow-up of Adverse Events ...................................................................................... 45 
7.3. 
Clinical Laboratory Test and Normal Laboratory Ranges .............................................. 45 
8. STUDY MONITORING, DOCUMENTATION AND RECORD KEEPING .................. 46 
8.1. 
Data Quality Assurance and Monitoring ......................................................................... 46 
8.2. 
Electronic Case Report Forms (eCRFs) .......................................................................... 46 
8.3. 
Clinical Trial Drug Accountability Data ......................................................................... 46 
8.4. 
Trial Documents .............................................................................................................. 47 
8.5. 
Reporting and Publication ............................................................................................... 47 
8.6. 
Archive ............................................................................................................................ 47 
9. ETHICS .................................................................................................................................... 48 
9.1. 
Institutional Review Board (IRB) or Ethics Committee Approval ................................. 48 
9.2. 
ICH GCP Guidelines and Declaration of Helsinki .......................................................... 48 
9.3. 
Informed Consent ............................................................................................................ 48 
9.4. 
HIPAA Compliance ......................................................................................................... 48 
10. REFERENCES ........................................................................................................................ 49 
Appendix I. Approval by an IRB/Ethics Committee .................................................................. 50 
Appendix II. Body Mass Index (BMI) Table ............................................................................... 52 
Appendix III. Prohibited Drug List ............................................................................................. 53 
Appendix III. Prohibited Drug List (Continued) ........................................................................ 54 
Appendix IV. Blood Metabolic/CBC and Urinalysis Tests ........................................................ 55 
 
 
 
 
 
 

Amphastar Pharmaceuticals, Inc. 
 
Protocol No. API-E004-CL-I 
Primatene Mist, Epinephrine HFA MDI (E004)                                         
Version 1.1 (11/ 08 /2019) 
 
 
 
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1. INTRODUCTION AND RATIONALE 
 
1.1. Introduction of the Drug Product 
 
Epinephrine, also known as adrenaline, is an adrenal hormone and sympathetic 
neurotransmitter with both α- and β-adrenergic activities. Systemically administered 
epinephrine can result in a whole array of adrenergic responses, resembling those induced by 
activation of the sympathetic nervous system and the adrenal glands. Epinephrine 
administered by oral inhalation is associated with a rapid and effective delivery to the 
respiratory tract, making it ideal for rescue of acute asthma symptoms. It has been well 
established that orally inhaled epinephrine is associated with a minimal systemic exposure  [3], 
due partly to its mode of pulmonary delivery and partly to its post-absorption degradation by 
catechol-O-methyltransferase 
(COMT) 
and 
monoamine 
oxidases 
(MAO) 
in 
the 
gastrointestinal tract and liver [4]. In addition, its local α-adrenergic vasoconstrictive actions 
in the lungs have been postulated to further reduce its systemic exposure after inhalation.  
 
Primatene® MIST (E004) is a epinephrine inhalation aerosol, supplied in metered dose 
inhaler (MDI), which allows 0.125 mg per spray of epinephrine. Primatene® MIST is the 
only FDA-approved over-the-counter asthma inhaler that is indicated for temporary relief of 
mild symptoms of intermittent asthma. 
 
Epinephrine Injection by 
(therapeutically equivalent to EpiPen®) is a patient-actuated, 
auto-injection system that delivers a single dose of 0.30 mg epinephrine injection. Each auto-
injector contains 1 mL epinephrine solution. Approximately 0.7 mL remains in the auto-
injector after activation, but is not available for future use, and should be discarded. This 
auto-injector is used for immediate administration in patients who are determined to be at 
increased risk for anaphylaxis, including individuals with a history of anaphylactic reactions. 
 
PROAIR HFA Inhalation Aerosol is made of albuterol sulfate, a racemic salt, of albuterol. 
PROAIR is supplied as a pressurized aluminum canister, with a plastic actuator, a dose 
indicator and dust cap. Each canister contains 8.5 g of the formulation and provides 200 
actuations. Each actuation delivers 120 mcg of albuterol sulfate from the canister valve and 
108 mcg of albuterol sulfate from the actuator mouthpiece (equivalent to 90 mcg of albuterol 
base).  Albuterol HFA is prescribed to treat bronchospasm, and also commonly used as 
rescue inhalers (short-term bronchodilators) to alleviate asthma attacks. 
 
 
 
 
 

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1.2. Mechanism of Action 
 
Although systemically administered epinephrine binds to all α- and β-adrenergic receptors 
non-selectively, the inhaled epinephrine is delivered to the pulmonary airways and binds to 
the β2-adrenergic receptors which are the predominant type in the bronchial smooth muscles 
[3]. This leads to the activation of adenylcyclase, and to an increase in the intracellular 
concentration of cyclic-3’,5’-adenosine monophosphate (cAMP). This increase of cAMP in 
turn leads to the activation of protein kinase A, which then inhibits the phosphorylation of 
myosin and lowers intracellular ionic calcium concentrations, and results in relaxation of 
smooth muscles of all airways. Increased cAMP concentrations are also associated with the 
inhibition of release of inflammatory mediators from mast cells in the airway. These events 
lead to dilation of the tracheal bronchial lumens with substantial increase in air passage. 
Epinephrine acts as a functional antagonist to relax the airway smooth muscles irrespective 
of the spasmogen involved, thus protecting against most if not all bronchoconstrictor 
challenges. 
 
Similarly, Albuterol relaxes the smooth muscle of all airways, from the trachea to the 
terminal bronchioles.  
 
1.3. Indications and Dose 
 
Primatene Mist, Epinephrine inhalation aerosol is indicated for temporary relief of mild 
symptoms of intermittent asthma (wheezing, tightness of chest, and shortness of breath) in 
patients age 12 years and older The recommended dose is 1-2 actuations every 3 hours. 
 
Epinephrine injection is indicated in the emergency treatment of allergic reactions (Type I) 
including anaphylaxis. 
 
PROAIR HFA Inhalation Aerosol is a beta2-adrenergic agonist indicated for treatment or 
prevention of bronchospasm in patients 4 years of age and older with reversible obstructive 
airway disease, as well as for prevention of exercise-induced bronchospasm in patients 4 
years of age and older. 
 
 
1.4. Pharmacokinetics (PK) and Pharmacodynamics (PD) Profile 
 
Pharmacokinetics: 
 
The absorption of inhaled epinephrine is rapid. In a multiple-dose systemic absorption of 
inhaled epinephrine study which enrolled six normal healthy volunteers, a rapid rise of 
plasma epinephrine levels with short duration were observed after administration of 15 puffs 

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followed by 30 puffs, with a >5 minute interval, at 160 mcg per puff. One minute after the 
initial 15 inhalations (2.4 mg) of epinephrine, the mean base-line epinephrine levels rose 
from 0.23 (±0.02) to a peak of 1.50(±0.61) nmol/L (i.e., 275 pg/ml). One minute after the 
subsequent 30 inhalations (a dose of 4.8 mg epinephrine), the epinephrine level rose to a 
peak of 4.22 (1.93) nmol/L (i.e., 772 pg/ml). The plasma epinephrine fell to baseline at 20 
min after 30 inhalations of epinephrine [2]. 
 
Overall systemic sympathomimetic effects are minimized due to a very low systemic 
exposure (~5% bioavailability) after oral inhalation [3]. 
 
The systemic levels of albuterol are low after inhalation of recommended doses. The terminal 
plasma half-life of albuterol delivered by PROAIR HFA was approximately 6 hours.  
 
Information from published literature suggests that the primary enzyme responsible for the 
metabolism of albuterol in humans is SULTIA3 (sulfotransferase). The primary route of 
elimination of albuterol is through renal excretion (80% to 100%) of either the parent 
compound or the primary metabolite. Less than 20% of the drug is detected in the feces.  
 
Pharmacodynamics: 
 
In a double-blind, randomized, two-period crossover study with mild-to-moderate asthma 
patients, the FEV1 measurements in subjects receiving inhalation of epinephrine were compared 
with those receiving placebo inhalations. Clinically significant improvement in FEV1, defined as 
an increase of ≥15% above baseline, has been observed. The results also showed that at 15 
seconds after one inhalation, 11 of the 24 subjects receiving epinephrine, but only 1 of the 23 
subjects receiving placebo, exhibited significant improvement in FEV1 (P < 0.004). Mean 
volume (and %) increments (± 1 SEM) in FEV1 at 15 sec after 1 inhalation were 390±60 ml 
(16.8±2.6%) after epinephrine, and -120±7 ml (-6.1±2.9%) after placebo (P < 0.0001). Mean 
peak volume (and %) increments in FEV1 were 800±80 ml (34.5±3.3%) after epinephrine, and 
180±60 ml (6.9±2.5%) after placebo (P<0.0001). The same study also demonstrated that the 
mean time (±SD) to peak increase in FEV1 (Tmax) was 7.5±7.8 min. With 2 inhalations 
administered 1 min apart, average Tmax was less than 10 min [5]. 
 
Epinephrine injection is mostly indicated for emergency treatment for anaphylactic reactions. 
When given subcutaneously or intramuscularly, epinephrine has a rapid onset and short 
duration of action [2]. Through its action on alpha-adrenergic receptors, epinephrine lessens 
the vasodilation and increased vascular permeability that occurs during anaphylaxis, which 
can lead to loss of intravascular fluid volume and hypotension. Through its action on beta-
adrenergic receptors, epinephrine causes bronchial smooth muscle relaxation and helps 
alleviate bronchospasm, wheezing and dyspnea that may occur during anaphylaxis. 
Epinephrine injectionalso alleviates pruritus, urticaria, and angioedema and may relieve 

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gastrointestinal and genitourinary symptoms associated with anaphylaxis because of its 
relaxer effects on the smooth muscle of the stomach, intestine, uterus and urinary bladder. 
 
1.5. Contraindications, Warnings, Precautions, and Adverse Reactions  
 
1.5.1. Contraindications 
 
Epinephrine Inhalation Aerosol is contraindicated in patients who are taking a prescription 
drug that contains phenylephrine, pseudoephedrine, ephedrine, or caffeine (for allergy, 
cough-cold, or pain), monoamine oxidase inhibitor (MAOI) (in certain drugs for depression, 
psychiatric or emotional conditions, or Parkinson’s disease), or for 2 weeks after stopping 
the MAOI drug.  
 
Albuterol Sulfate HFA is contraindicated in patients with a history of hypersensitivity to 
albuterol and any other Albuterol Sulfate HFA components.  
 
1.5.2. Warnings and Precautions 
 
Primatene Mist should be taken with precaution under certain conditions: 
 ever been hospitalized for asthma 
 heart disease 
 high blood pressure 
 diabetes 
 trouble urinating due to an enlarged prostate gland 
 thyroid disease 
 seizures 
 narrow angle glaucoma 
 
Epinephrine injection should be used with precaution: 
 more than two sequential doses of epinephrine should only be administered under 
direct medical supervision. 
 Epinephrine injection should only be injected into the anterolateral aspect of the 
thigh 
 rare cases of serious infections at the injection site may occur 
 may trigger allergic reactions associated with sulfite in certain susceptible individuals 
 patients with heart disease, hyperthyroidism, diabetes,  Parkinson’s disease (may notice 
a temporary worsening of symptoms), elderly individuals, and pregnant women should 
be carefully instructed in regard to the circumstances under which epinephrine should 
be used. 
 

Amphastar Pharmaceuticals, Inc. 
 
Protocol No. API-E004-CL-I 
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PROAIR should be taken with precaution and discontinue use under the following 
circumstances: 
 paradoxical bronchospasm may occur 
 deterioration of asthma 
 not to substitute for use of anti-inflammatory agents 
 cardiovascular effects may occur 
 exceeding  recommended dose may be fatal 
 immediate hypersensitivity reactions may occur 
 hypokalemia and changes in blood glucose may occur. 
 
 
1.5.3. Adverse Reactions 
 
The clinically observed adverse events are mainly related to its systemic adrenergic actions 
exerted by the small fraction of epinephrine that enters into the systemic blood circulation 
and interacts with the α- and β-adrenoceptors on the endothelia cells. The common and 
expected adverse event include: Increased blood pressure and heart rate (which could lead 
to more serious problems such as heart attack and/or stroke), nervousness, sleeplessness, 
rapid heartbeat, tremor, seizure, etc. Rarely, bronchial irritation and edema may occur. In 
some patients, severe prolonged asthma attacks may be precipitated. Rebound 
bronchospasm may occur when the effects of epinephrine end. 
 
Adverse reactions to epinephrine include anxiety, apprehensiveness, restlessness, tremor, 
weakness, dizziness, sweating, palpitations, pallor, nausea and vomiting, headache, and/or 
respiratory difficulties. 
 
Most common adverse reactions associated with the use of PROAIR are headache, 
tachycardia, pain, dizziness, pharyngitis, and rhinitis. 
 
Use of Albuterol Sulfate HFA may be associated with the following: 
 paradoxical bronchospasm; 
 cardiovascular effects; 
 immediate hypersensitivity reactions; and 
 hypokalemia 
 Hyperglycemia. 
 
1.6. Drugs with Strong Impact on PK or PD 
 
Drugs with impact on PK: Short-acting sympathomimetic aerosol bronchodilators and 
adrenergic drugs. 

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Drug with Impact on PD and safety: (i) Non-selective α-blockers; (ii) Beta-blockers; (iii) 
Diuretics, or non-potassium sparing diuretics may potentiate hypokalemia or ECG changes; 
(iv) Digoxin; (v) Monoamine oxidase (MAO) inhibitors; (vi) Sympathomimetics; (vii) 
Thyroid hormones; and  (viii) tricyclic antidepressants.  
 
A list of drugs, which have strong impact on PK or PD profile for epinephrine and Albuterol, 
is provided in Appendix III. 
 
 

Amphastar Pharmaceuticals, Inc. 
 
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2. OBJECTIVES 
 
2.1. 
Objective  
 
To assess the drug exposure profile in systemic circulation of Primatene® Mist by inhalation, 
versus Epinephrine by IM, and ProAir® HFA by inhalation in healthy adults 
 
2.2. Primary Endpoints   
 
 
AUC0-6h, defined as area under the curve (AUC) concentration curve over time, 
from time 0 to 6 hours post-dose for Epinephrine; 
 
AUC0-24h, defined AUC of plasma Albuterol concentration curve over time 0 to 24 
hours post-dose; 
 
AUC0-∞, defined as AUC in the plot of plasma Albuterol, or Epinephrine versus 
time from time 0 to infinity; and 
 
Cmax, defined as the maximum plasma concentration of Albuterol, or Epinephrine. 
 
 
2.3. Secondary Endpoint 
 
tmax of Albuterol, or Epinephrine, defined as the time, at which the Cmax is observed; 
and 
 
T1/2, terminal elimination half-life of each analyte. 
 
 
 

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3. INVESTIGATORS AND STUDY SITES 
 
3.1. Principal Investigator(s)  
 
Principal Investigators (PI) responsible for the conduct of this study will be board certified 
physicians who are familiar with the study medications and trained to handle any untoward 
events related to medications.  
 
The PI is responsible for the global aspects of the research study. For safety monitoring, a 
study physician or healthcare professional, who is certified for Advanced Cardiac Life 
Support, will be available at the study site at the time of study drug administration and during 
the entire time of PK blood sampling period. The Institutional Review Board (IRB) 
acknowledges one PI per study site. Qualifications for the PI and Co-Investigator(s) are listed 
on the Form FDA 1572. 
 
3.2. Investigational Site(s) 
 
The detailed information about the study site can be found on Form FDA 1572, including: 
Name, address, and statement of qualifications of each Principal Investigator; name of each 
sub-Investigator working under the supervision of the Investigator; name and address of the 
research facilities to be used; name and address of each reviewing IRB per US 21 CFR 
312.23 section 6(iii)(b). 
 
3.3. Laboratory 
 
Samples of blood for evaluation of hematology and chemistry (metabolic panels) will be 
analyzed by the clinical laboratory with certification from a recognized accreditation agency 
(e.g., College of American Pathology or Clinical Laboratory Improvement Amendments 
certification). Refer to Appendix IV for a list of parameters analyzed, and the laboratory 
manual for details regarding specimen sample collection, processing, and shipment 
procedures. PK samples will be processed and analyzed by R&D Department, Amphastar 
Pharmaceuticals, Inc. 
 
 

Amphastar Pharmaceuticals, Inc. 
 
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4. MATERIALS AND METHODS 
 
4.1. Study Design 
 
This is a randomized, safety evaluator-blind, three-treatment, crossover, fasting study in 
healthy adults (18 - 50 years of age). 
 
4.2. Study Subjects 
 
4.2.1. Sample Size 
 
A total of at least 24 evaluable healthy adult volunteers will take part in this crossover 
study. 
 
4.2.2. Subject Enrollment 
 
A sufficient number of candidates will be screened to ensure 24 evaluable subjects will be 
enrolled into the study based upon the satisfaction of the inclusion and exclusion criteria. 
Informed consent will be obtained prior to performing any screening procedures. 
 
4.2.3. Inclusion Criteria:  
 
Candidates will be qualified only if they meet all of the following criteria: 
(1) 
Agree to participate; understand and sign informed consent; 
(2) 
Male and female adults, aged 18 to 50 years, inclusive at Screening;  
(3) 
Generally healthy and medically stable, with no clinically significant abnormalities 
based on physical examination and laboratory tests as determined by the 
Investigators; 
(4) 
Have good venous access; 
(5) 
Have normal resting blood pressure and normal heart rate (HR) without history of 
syncope; a subject with out of range blood pressure may be enrolled in the study at 
the discretion of the Principal Investigator;  
(6) 
Have a body mass index (BMI) of 18.0 – 30.0 kg/m2;  
(7) 
Female candidates must be >1 year post-menopausal or practicing a clinically 
acceptable form of birth control and confirmed by negative urine or serum 
pregnancy test at Screening;  
(8) 
Negative HIV-Ab, HBs-Ag and HCV-Ab; 
(9) 
Negative alcohol test (urine or breathalyzer); 
(10) Negative drug screening results;  
(11) Currently non-smoker; have not used any tobacco products for at least three (3) 
months prior to Screening; and 

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(12) Demonstrate proficiency in the use of MDI and a consistent inhalation time >2.0 
seconds after training, for at least three (3) times, with a maximum of 5 attempts.  
 
4.2.4. Exclusion Criteria 
 
Candidates will be excluded from the study for any of the following reasons: 
 
(1) 
Concurrent clinically significant cardiovascular, hematological, renal, neurologic, 
hepatic, endocrine, psychiatric, or malignant diseases. 
(2) 
Known intolerance or hypersensitivity to any component of the study drugs (i.e., 
Epinephrine, Albuterol Sulfate or any sympathomimetic drugs, HFA-134a, thymol, 
ethanol, ascorbic acid, nitric acid, and hydrochloric acid). 
(3) 
Upper or lower respiratory tract infection, or other systemic infection within 6 weeks 
prior to Screening; 
(4) 
Clinically significant abnormalities in the screening/baseline ECG; prolonged 
corrected QT interval (QTcF) on ECG: men >450ms, women: >470ms; single or 
multiple premature ventricular contractions (PVC); 
(5) 
Abnormal thyroid function test (if TSH is out of range, refer to T3/T4 for thyroid 
function assessment); 
(6) 
Subject has been on other investigational drug/device studies within 30 days of 
Screening Visit or planned participation in another investigational drug trial at any 
time during this trial; 
(7) 
Women who are pregnant or lactating or planning a pregnancy during the study 
period; 
(8) 
Subject has donated or lost > 500 mL of blood within 3 months of Screening;  
(9) 
Evidence of alcohol or drug abuse or dependency within 6 months prior to 
screening; or 
(10) Use of any of the prohibited drugs listed in Appendix III without appropriate 
washout.  
 
 
4.3. Randomization Procedures 
 
4.3.1. Subject Identification Assignment 
 
Subject will sign informed consent and be assigned a screening ID, as  
 
EIS-mm, 
 
where EI  represents “E004-CL-I”, S stands for screening, and mm is the screening 
sequential number. The subject will be evaluated per inclusion/exclusion criteria. The 

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following procedures will be performed at Screening: (i) physical examination (PE), (ii) 
measurements of vital signs, (iii) ECG, (iv) urinalysis (UA), (v) urine drug screen, (vi) 
alcohol screen (urine or breathalyzer), (vii) urine or serum pregnancy test (female subjects), 
and (viii) fasting (≥ 8 hours) blood tests [comprehensive metabolic panel, complete blood 
count (CBC) with differentials, thyroid function tests, HIV, hepatitis B and C tests]. 
 
After satisfying the enrollment criteria, subjects will be assigned a unique subject ID as  
  
EIPK-nn, 
 
where nn is the sequential number in order of enrollment.  
 
The same Subject ID will be used throughout the Dosing Visits. The Subject ID numbers 
will be recorded on the study source documents, the case report forms, and other 
appropriate study documents. In addition, all subjects will be identified by their first, 
(middle), and last initials. 
 
4.3.2. Subject Randomization 
 
The Investigator must ensure that all inclusion/exclusion criteria are met, and will endorse 
the subject enrollment at the beginning of Dosing Visit 1.  
 
A set of computer generated randomization codes will be supplied by Amphastar IT 
department in accordance with the Standard Operating Procedure (SOP). A 3 × 3 Latin 
Square method will be used for randomization. The randomization code will dictate the 
sequence of the three (3) crossover treatments for individual subject. 
 
4.4. Dispensing, Returning, and Storage of Material and Drug Supplies 
 
All study drug units are prepared and packaged as individual units by the Amphastar 
manufacturing team under GMP conditions and QA oversight. Study drugs will be packaged 
and delivered to the investigational site. All study units are labeled with proper subject ID#, 
and Visit#, but are devoid of any treatment-identifying information.  
 
Study drugs will be properly labeled and delivered to the dosing area for immediate use. All 
study drug units must be returned to the site or designee when the visit is completed, and its 
use documented.   
 
At each Dosing Visit, one (1) study treatment (Treatment-A, B, or C) will be assigned to 
subjects per the randomization code (Table 1).  
 

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Table 1. Treatments and Dose 
 
 
 
Study drugs should be stored at USP defined, controlled room temperature, with excursions 
permitted from 15°-25°C (59°-77°F). The study drugs will be protected from environmental 
extremes and stored in a locked cabinet or room to which only designated persons have 
access. The inhalers will be stored with the mouthpiece down.  
 
4.5. Study Drugs Administration and Training 
 
4.5.1.  Training for Correct Use of MDI Inhaler 
 
During the Screening and at the beginning of every Dosing Visit, each subject will be trained 
on the correct use of the MDI inhalation aerosols in a standard fashion to assure a relatively 
consistent inspiratory flow rate and duration. A video on how to use the MDI will be 
provided at each training session. The training will use placebo, a MDI simulation unit that 
contains no active drug. Subjects have to demonstrate a consistent inhalation time >2.0 
seconds (at least 3 times, with a maximum of 5 attempts) after training. Vitalograph AIM™ 
(Aerosol Inhalation Monitor) will be used to monitor and measure the duration of the 
inhalation during the training at Screening and prior to dosing.  
 
4.5.2.  Preparation and Training of MDIs at the Dosing Visit  
 
Each MDI must be primed at the study site prior to the first use by releasing 4 test sprays 
(E004 for Treatment-A), or 3 test sprays (ProAir for Treatment-C) into the air away from the 
face, shaking well before each spray. Priming will be performed in an area that is well 
ventilated (preferably a fume hood, or an open space), and physically separated from the 
study evaluation area. Refer to the training video for details on the priming procedure. 
 
Items
Treatment-A
Treatment-B
Treatment-C
Study Drug Name
Primatene Mist, E004
Epinephrine Injection Auto-
Injector (Generic of EpiPen®)
ProAir
Manufacturer
API
Epinephrine
Epinephrine
Albuterol Sulfate
Dosage Form
Microcrystalline Suspension
HFA MDI
Sterile Solution
Microcrystalline Suspension
HFA MDI
Strength
0.125 mg/inh
0.3mg/0.3mL
0.09 mg/inh
Dose Regimen
      2 inhalations, 0.25mg   
IM Injection of 0.30 mg 
Epinephrine in 0.30 mL            2 inhalations, 0.180 mg   
Route of Administration
Oral Inhalation
 IM Injection
Oral Inhalation

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(1) Correct sitting position: In order to get a deep proper breath during dosing, the 
subject must sit on a stool, not a chair, with their posture straight in a relaxed calm 
state for at least 1 min prior to dosing. Do not slouch or lean forward or backward. 
(2) Finger placement for dosing: To ensure proper drug delivery, subject is required to 
hold the canister with the thumb at the bottom and place the index finger of the same 
hand on the top of the canister to release the drug. For each actuation, subject must 
fully depress the top of the metal canister, and hold it for at least one second 
before releasing. 
(3) Critical Timing: Subject begins a slow and deep inhalation before pressing the 
inhaler. A slight delay in the sequence can result in deposition of inhaled drugs on 
the back of the throat and tongue. 
(4) Deep inhalation as long as possible: After pressing the inhaler, subject should 
continue to inhale slowly and deeply for as long as possible (at least 3-5 seconds, 
refer to the training video). Keep tongue away from the opening of the mouthpiece. 
(5) After deep inhalation, hold the breath for approximately 10 seconds (5- 10 seconds) 
[6] to allow deposition of drug particles into the lower level of the lung. 
 
4.5.3. Detailed MDI Dosing Procedure 
 
(1) The designated study site personnel will verify if the canister is fully inserted into 
the actuator. 
(2) The designated study site personnel will remove the cap from the mouthpiece of the 
actuator, and check the mouthpiece for foreign objects prior to use;  
(3) Subjects will first shake and spray into the air one time to mix the medicine. 
(4) Subjects will exhale deeply through the mouth and empty  the lungs as completely as 
possible by exhaling fully (exhalation must be deep and slow, similar to the deep 
inhalation step); 
(5) Immediately after the full exhalation, subjects will hold the inhaler with the 
mouthpiece down, then places the mouthpiece fully into their mouth, with lips sealed 
around the mouthpiece; 
(6) While inhaling, subjects will fully depress the top of the canister with an index 
finger. Subjects should keep inhaling deeply and slowly for as long as possible (at 
least  3-5 seconds) through the mouth; 
(7) After breathing in fully, remove the inhaler from the mouth, subjects keep the mouth 
closed, hold the breath for approximately10 seconds (5- 10 seconds) [6]; 
(8) Subjects will exhale slowly through the nose (keep lips nearly closed); and 
(9) Repeat the above (1) to (7) steps for the second inhalation. The time interval 
between each actuation will be within 30 seconds. 
 
 

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4.5.4. IM Injection Procedure 
 
IM injection (0.30mg of epinephrine solution in 0.30 mL) for Treatment-B will be given on 
the thigh. Subjects will be lying down in a supine position. The 0.30 mL dose will be given 
perpendicularly as a single deep IM injection into anterolateral aspect of the thigh (vastus 
lateralis site). Epinephrine auto-injector (Treatment B) will be administered as follows [2]: 
 
(1) Doser or designated site personnel will quickly twist off the yellow cap off the auto- 
injector. 
(2) Subject will hold the leg firmly in place.  
(3) Doser or designated site personnel will first pull off the blue safety release to unlock 
the auto injector.  
(4) Doser will place the orange tip against the middle of the outer thigh (upper leg) at a 
right angle (perpendicular) to the thigh. 
(5) Swing and push the injector firmly until it ‘clicks.’ The click signals that the 
injection has started. 
(6) Hold firmly in place for 3 seconds. Time 0 begins once the holding is completed. 
(7) Remove the injector from the thigh and massage (apply gentle pressure) with dry 
gauze to the injection area for 10 seconds. The orange tip automatically extends out 
after use. 
(8) The injection site will be observed for any medication leakage for approximately 30 
seconds after the dosing is completed. If there is observed leakage, PK samples will 
not be collected, subject will be early terminated (ET). 
 
4.6. Blinding Methodology and Procedure 
 
The preparation area will be physically separated from the dosing area to ensure the evaluator 
remains blinded. All treatments will be prepared by qualified personnel at the study site prior 
to dosing at each study visit according to the order of treatment arms assigned to subjects per 
the randomization code. Subjects will be escorted to a separated dosing area for study drug 
administration to preserve blinding from evaluators. 
Study drugs will be dispensed by CRC or pharmacist per randomization.  
 
Safety evaluator(s) will remain blinded during the course of the study and will have the sole 
responsibility of evaluating any Adverse events. 
 
4.7. Unblinding  
 
Should any safety evaluator become “unblinded” to any subject’s treatment assignment 
during the study, the Investigator shall notify Amphastar Clinical Research Department 
within 24 hours. The Justification for Code Breaking Form will be signed and dated by the 

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Investigator, and the Amphastar Clinical Research Associate (CRA). The form will be placed 
back into the envelope, which is not to be resealed, and kept in study site regulatory files, 
separated from the other envelopes. Any subject whose envelope seal has been broken will 
be subject to QA audit and may be excluded from the BE data analyses. Subject should also 
be reminded to maintain the identity/configuration of the study drug provided to them and 
must not disclose this information to the blinded study team or other subjects in order to 
maintain the blinding of the evaluators. 
 
 

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5. STUDY PROCEDURE 
 
5.1. Global Plan 
 
The design includes (i) a Screening Visit (3-7 days prior to Visit-1), (ii) three (3) Dosing 
Visits separated by a 2-7 day washout period, and (iii) a post-study follow-up evaluation 1-7 
days after Dosing Visit-3 or study termination. 
 
5.2. Screening Procedure and Scheduling 
 
5.2.1. Schedule and Purpose of Screening Visit 
 
The Screening Visit takes place 3-7days prior to Dosing Visit-1. During Screening, subject 
qualification will be determined based on the Inclusion/Exclusion Criteria (Sections 4.2.3 
and 4.2.4), the physical examination, and laboratory test results. These data will be 
documented on the subject source documents. 
 
5.2.2. Informed Consent Form 
 
The Informed Consent document will be used to explain the purpose, risks, and benefits of 
study participation to the subject in simple terms at the Screening Visit. The Investigator 
must obtain a signed informed consent form from each subject before proceeding to any 
screening and study activities. A copy of the signed Informed Consent Form should be 
given to the subject. The Informed Consent Form includes compliance with the privacy 
provisions of HIPAA. 
 
5.2.3. Medical History, PE, and General Safety (Screening Activities) 
 
The Screening Visit entails the following screening activities: 
 
(1) 
Interview for demographic information and medical history; 
(2) 
Body Mass Index (BMI) needs to meet the requirements in Section 4.2.3. BMI can 
be calculated and checked by the formula and chart in Appendix II, or by utilizing 
the NIH website; 
(3) 
Review medication history for any prohibited drugs (Appendix III). Ensure any 
appropriate washout has been observed; 
(4) 
A full physical examination and measurements of vital signs will be performed to 
ensure that the subject is generally healthy;  
(5) 
The subject has to rest in a secluded area for at least 10 minutes before 
measurements of vital signs. The blood pressure and heart rate measurement can be 
repeated twice, each separated by a 15 minute interval;  

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(6) 
12-lead ECG will be conducted (routine and QT/QTc) to ensure absence of overt 
cardiac illnesses;  
(7) 
Collect urine samples for urinalysis (routine and microscopic), alcohol/drug screens 
(urine or breathalyzer test allowed for alcohol) for all subjects, and pregnancy test 
for all female subjects (serum pregnancy test allowed); 
(8) 
Collect blood samples for CBC, thyroid function, HIV-Ab, HBs-Ag, and HCV-Ab 
tests; 
(9) 
Collect fasting (≥ 8 hours) blood samples for serum comprehensive metabolic panel 
(per Appendix IV); and 
(10) At the Screening Visit, subjects should be trained on the use of the MDI inhalation 
aerosols in a standard fashion to assure a relatively consistent inspiratory flow rate 
and inspiratory duration as described in Section 4.5. Eligible subjects must 
demonstrate the correct use of the MDI unit and a consistent inhalation time >2.0 
seconds after training, for at least three (3) times, with a maximum of 5 attempts. 
AIM™ (Aerosol Inhalation Monitor) will be used to monitor and measure the 
duration of the inhalation during the training at Screening and prior to dosing. 
 
Out of range lab results may be repeated once at the discretion of the Investigator. Subject 
enrollment will be confirmed at the beginning of Dosing Visit-1 (Section 5.3.1). The final 
decision of enrollment will be determined by the Investigator. 
 
5.2.4. Instruction to Subjects at the End of Screening Visit 
 
(1) Dates for Dosing Visit-1 Visit-2, and Visit-3 should be separated by 2 to 7 days. 
(2) Subject must be at least 10-hours fasting for each Dosing Visit. 
(3) Subjects must refrain from alcohol consumption and caffeine-containing (including 
decaffeinated) beverages or foods (coffee, tea, applicable soft drinks including cola 
and analogs, or chocolate) for 24 hours prior to each Dosing Visit. 
(4) At each Dosing Visit, subjects are required to check in at the site the night before. 
Dinner will be provided at study site.  
 
5.3. Dosing Visits 
 
5.3.1. General Profile for Dosing Visits 
 
The same procedures will be performed at each Dosing Visit, except that the subject will be 
given different study treatments based on the randomization code. 
 
At Dosing Visit-1 the following activities will be conducted: 
 
(1) Enrollment is verified at the beginning of Dosing Visit-1, by reviewing all 
inclusion/exclusion criteria and other requirements.  

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(2) Upon verification of subject enrollment, a Subject ID will be assigned and the 
randomization procedure will be completed (see Section 4.3.1 and 4.3.2) 
 
5.3.2. Pre-dose Activities  
 
At each Dosing Visit, the following review and verification will be performed prior to 
study drug administration: 
 
(1) A urine or serum pregnancy test must be performed on all female subjects. If it is 
positive, the subject will be terminated from the study; 
(2) Any concomitant medication since the last visit will be recorded, including name of 
medication, route of administration, dosage, indication, and start and end dates 
where available. If any medications listed in Appendix III were used after the last 
visit without the proper washout period, the subject will be terminated; 
(3) Perform alcohol and drug test; if any is positive, subject will be terminated; 
(4) Subjects will be trained again on the use of the MDI inhalation aerosols prior to each 
treatment session, as described in Section 4.5.1;  
(5) Placement of ECG leads; 
(6) Subjects need to rest for at least 10 minutes prior to the baseline vital signs 
evaluations. Vital signs should be taken from the opposite arm of the IV catheter arm 
if possible; Vital signs measurement may be repeated twice to PI discretion.  
(7) An indwelling IV catheter shall be placed into one of the antecubital region veins for 
blood drawing, or separate venipunctures may be used; and 
(8) One (1) baseline blood samples will be collected within 30 minutes before dosing. 
 
5.3.3. Instructions to the Subjects before Dosing at Dosing Visit 
 
(1) No food will be allowed for 10 hours prior to dosing and until 4 hour post-dose PK 
samples have been collected. 
(2) Meals will be provided at normal times after the 4 hour post-dose blood draw. 
(3) Water intake is not allowed 1 hour before and 1 hour after drug administration. 
(4) All physical activities during the Dosing Visits should be limited with the exception 
of bathroom visits until the completion of the 120-minute post-dose PK sampling at 
each Dosing Visit. 
(5) The subject will remain in a semi-recumbent or seated position for 120 minutes after 
drug dosing. After the 120-minute PK sampling, the subject has no movement 
restrictions, but should refrain from any excessive physical activity.  
(6) During each Dosing Visit, subjects should refrain from alcohol consumption and 
caffeine-containing (including decaffeinated) beverages or foods (coffee, tea, 
applicable soft drinks including cola and analogs, or chocolate). 
 
 
 

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5.3.4. Study Medication Administration 
 
At each dosing visit, subjects will either self-administer 2actuations from the MDI inhaler 
provided, or receive an IM injection (0.30mg Epinephrine injection for Treatment-B). 
 
IM Injection into the vastus lateralis site: 
 
IM injection (0.30mg of epinephrine solution in 0.30 mL) for Treatment-B will be given on 
the thigh. Subjects will be lying down in a supine position. The 0.30 mL dose will be given 
perpendicularly as a single deep IM injection into anterolateral aspect of the thigh (vastus 
lateralis site). Refer to Section 4.5.4 for detailed IM injection procedure. 
 
MDI Self-Administration: 
 
The designated unblinded dispenser at the study site will prime each unit prior to the first 
use (Section 4.5). Each subject will also be retrained on the use of the MDI inhalation 
aerosols as described in Section 4.5.1 to Section 4.5.3.  
 
Study drugs will be dispensed by the designated site professional, and the reading on the 
dose indicator of the MDI unit will be recorded prior to dosing. 
 
Subject will shake the inhaler well before each spray. Subjects will self-administer 2 
inhalations from the inhaler unit (within 30 seconds interval between each inhalation). The 
study drug administration process will be video-recorded and used for an investigation of 
apparent dosing irregularity. Refer to Section 4.5.3 for critical instructions for correct 
dosing techniques. 
 
The study medication administration should be conducted between 07:00 and 11:00 AM on 
the Dosing Visit. Time of the start of each inhalation will be recorded. 
 
Once dosing is completed, CRC or other site professionals will record the readings of each 
dose-indicator in the study source documents. The used MDI inhaler will be collected at the 
end of the Dosing Visit. 
 
 
5.3.5. PK Blood Sampling  
 
At each Dosing Visit, PK blood samples will be collected and plasma will be isolated for 
analyzing the concentrations of Albuterol, and epinephrine. A total of 26 blood samples 
will be drawn from each subject at each Dosing Visit as indicated in Table 2. 
 

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(1) A catheter will be inserted into one of the antecubital region veins for blood 
sampling. Alternatively, collection from a vein of the hand or forearm for easier 
access or via venipuncture is also acceptable;  
(2) One (1) pre-dose samples (baseline) will be collected within 30 minutes before 
dosing; 
(3) Time 0 is defined as the beginning of the last inhalation of the MDI, or after 
completion of IM injection, Section 4.5.3.  
 
Table 2. PK Blood Sampling Schedule  
 
 
(4) A catheter flush will be performed between each blood sample collection to avoid 
catheter clot formation and prevent interference between samples. At each sampling 
point, the first 1.0 mL portion of the blood (or saline flush) will be collected into a 
disposable tube and discarded;  
(5) At each PK sampling point, blood samples (~5 mL), will be collected in ice-chilled 
potassium-EDTA sample tubes, mixed well (gently invert the tube 8-10 times) 
immediately following collection, kept on ice or refrigerated and centrifuged within 
20 minutes of collection. 
(6) If a tube breaks, a new sample can be collected if it is within the allowed time 
window (Table 2). 
(7) Plasma isolation (i.e., centrifugation of the blood samples) will start within 20 
minutes of blood collection. Blood samples will be centrifuged at 4°C, 2,000-3,000 
g for 20 minutes for plasma isolation. 
(8)  Isolated plasma will be distributed into two (2) 2 mL cryo vials, approximately 1.5 
Seq. 
#
Time window
Sample 
No.,"XX"
Seq. 
#
Time window
Sample 
No.,"XX"
1
within 30 min 
pre-dosing
01
14
40 min
14
2
1 min
02
15
50 min
15
3
2 min
03
16
60 min
16
4
3 min
04
17
70 min
17
5
5 min
05
18
80 min
18
6
7 min
±1 min
06
19
90 min
19
7
9 min
07
20
120 min
20
8
12 min
08
21
4 hrs
21
9
15 min
09
22
6 hrs
22
10
18 min
10
23
8 hrs
23
11
21 min
11
24
12 hrs
24
12
25 min
12
25
18 hrs
±20 min
25
13
30 min
13
26
24 hrs
26
±2 min
±10 min
±15 min
±5 min
±1 min
± 0.5 min
Scheduling
Baseline 
Scheduling

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mL in the first vial and the remainder in the second vial. The plasma sample vials 
will be frozen immediately on dry ice (within 60 minutes following plasma 
separation) and then stored (within 60 minutes following plasma separation) in a 
freezer at -20C or lower until analysis. 
    
 
5.3.6. PK Blood Sample ID and Labeling  
 
The sample ID for PK blood samples (before centrifugation) is defined as  
                   
“EIPK-nn-V-XX” 
 
Where, EIPK-nn is the subject ID; V is the Dosing Visit code, V=1, 2 or 3; and XX is the 
two-digit time-point sequential number of the sample: 01, 02, 03... 26 as listed in Table 2. 
 
The sample ID for PK plasma samples (after centrifugation) is defined as “EIPK-nn-V-
XX-k” and k is the aliquot number (A or B). The PK plasma sample ID labels, with the 
size of 1/2”×1¾” (Avery label 5667), should be printed in blue font as demonstrated below: 
 
       
 
5.3.7. Safety Evaluations/Monitoring  
 
The following safety measurements will be performed during each Dosing Visit: 
 
(1) Vital signs (HR, SBP/DBP): at baseline, 5±2 min, 15±5 min, 60±5 min, 120±15 min 
and 24 ±2 hrs post-dose. 
(2) 12-lead ECG (routine QT and QTc analysis): at baseline, 20±5 min, 120±15 min and 
24 ±2 hours post-dose. 
(3) Serum potassium and glucose levels will be measured at baseline, 10±2 min, 60±5 
min, and 24 ± 2 hrs post-dose. 
(4) All subjects will be queried for adverse events (AEs) and AEs must be recorded with 
all related information.  
 
5.4. Activity at End of Study (EOS) 
 
The following EOS evaluations will be conducted at the end of study as follows: 
 
(1) The Investigator(s) will ensure that all study drugs and study records are 
accountable. 
(2)  At Visit-3, blood and urine samples will be taken for fasting comprehensive 
metabolic panel (Refer to Appendix IV). Lunch will be provided after the EOS 
EIPK-nn-V-XX-k 

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blood draw. The meals and fasting schedule is indicated in Table 3. 
 
Table 3. Meals and fasting schedule 
              
 
      * At Visit-3, breakfast will be provided after EOS blood draws 
 
(3) Conduct EOS physical examination between 16 and 26 hours post-dose. EOS Vital 
signs, ECG, CBC, fasting comprehensive metabolic panel and serum 
potassium/glucose will be measured at the end of PK sample collection (24 ± 2 hrs 
post-dose) at Visit-3. 
(4) Adverse events, whether reported by the subjects or observed by study staff, will 
need to be documented, and reported or followed up where applicable. 
(5) Review all safety and compliance data. 
(6) For Early Termination (ET), an ET evaluation will be conducted with the same 
safety assessments as listed (items 1-5 above) when appropriate and feasible.  
 
 
5.5. Follow-up Activities 
 
All treated subjects will be followed up by a phone call scheduled 1-7 days after the 
completion of Dosing Visit-3 or study termination. Subjects will be asked if they have 
experienced any adverse event after the last Dosing Visit. 
 
5.6. PK Sample Transfer and Laboratory Analysis 
 
The plasma samples will be transferred at temperature below -20˚C from the site to the 
Amphastar R&D Laboratory. The chain of custody, temperature control and related 
documents for sample transfer must be recorded and documented. 
 
All plasma samples should be stored at -80ºC or lower in the laboratory. Epinephrine and 
Albuterol concentration in the PK plasma samples will be analyzed by the R&D Department 
of Amphastar with validated methods under GMP. 
 
 
5.7. Summary and Schema of Study Activities  
Night before Dosing
Day 1
Day 2 (Visit-3 Only)
Morning
NA
Dosing
Breakfast will be provided 
after EOS blood draw
Mid-Day      
NA
Lunch                 
(≥ 4 hr post-dose)
NA
Evening
Dinner, then fast
overnight (≥ 10 hr)
Dinner,  then fast 
overnight(≥8hr)*
NA

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5.7.1. Data collection  
 
Data collection for this study is summarized as follows: 
 PK samples for evaluation of plasma epinephrine or Albuterol concentrations; 
 Safety data as vital signs, ECG, urinalysis, serum potassium, glucose and clinical 
laboratory testing, etc.; and 
 Adverse events reported. 
 
5.7.2. Schema of Study Activities 
 
A schematic outline of the study activities is provided in Table 4 below. 
 
 
 
 
 
 

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Table 4. Summary of Activities in All Visits and EOS 
 
 
 
1. The fasting comprehensive metabolic panel (≥8 hours) for EOS will be taken at the Study Visit-3 or study 
termination 
2. Vital signs, ECG  and serum potassium and glucose at 24 ± 2h at Visit-3 can be used for EOS 
 
 
 
Study Activities & Evaluations
Screen 
Visit
Dosing       
Visit-1  
Dosing       
Visit-2 and 3
EOS1 
Informed Consent / HIPAA 
X
Medical history / demographics
X
Physical Exam (PE)
X
X 
Lab: Urinalysis (UA)
X
X 
Lab: CBC/Comp. metab. Panel
X
X 
Lab: Thyroid Function 
X
Lab: HIV-Ab, HB-sAg, HCV-Ab
X
Inclusion/Exclusion
X
X
Confirmation of enrollment
X 
Assign subject ID
X 
Randomization
X 
Training/Dosing of MDI
X
X
X
Concomitant Medicines
Urine or serum pregnancy test
X
X
X
Urine drug screen
X
X
X
Alcohol screen (urine or breathalyzer)
X
X
X
Food / beverage restrictions
X
X
Meals served
X
X
PK Baseline samples
X (1)
X  (1)
PK samples
X (25)
X (25)
Vital signs
X
X (6)
X (6)
X 2
12-lead ECG
X
X (4)
X (4)
X 2
Serum potassium and glucose
X
X (4)
X (4)
X2 
Adverse event reporting
Record throughout study period
Record throughout study period

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5.8. Chart for Study Activities   
 
Study activities are summarized in Figure 1. 
 
Figure 1. Summary of Study Activities 
 
screen, EOS 
screen, Dosing Visits
0             5'    10'   15'      20'                            60'                       120'       
24h
Dosing Visits
Thyroid, HIV, HBs ( Screen only),  Physical Examination, CBC, Metabolic Panel, Urinalysis
EOS
Urine Alcohol/Drug Screen, Pregnancy test (female subjects)
PK Sampling Time
0*
1'2'3' 5' 7' 9'12' 15'18' 21'25' 30'  40'  50' 60' 70'  80'  90'  120'      4h        6h         8h       
12h         
18h
24hPost-dose time
Time window
Study drug dosing
* Baseline sample 
Vital Signs(V), ECG (E), Serum Potassium and Glucose  (KG) 
screen, Dosing Visits, EOS
V,E, KG       V     KG    V       E        
V, KG                     V,E 
V,E, KG
 5'                          10'                   15'
(±2') (±2')  ( 5')  ( 5')                          (±5')
( 15') 
( 2h)
 1'
 20'
 2'
 1'

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6. STATISTICAL ANALYSIS 
 
6.1. Populations 
 
6.1.1. Per-Protocol Population (PPP) 
 
The “Per Protocol” population is defined as all subjects who have received all study 
medications during the study and are evaluable for all treatments. The primary analyses 
will be performed based on “Per Protocol” population (PPP).  
 
An evaluable subject for primary analyses of this study must meet all of the following ten 
(10) items for both treatments: 
 
(1) 
Correct dose and administration;  
(2) 
Nineteen (19) or more of 25 post-dose PK data points (>75%) are available; 
(3) 
There are no more than four (4) consecutive missing PK data points; 
(4) 
The baseline PK data point is available; 
(5) 
For Treatment-A, at least four (4) of the five (5) PK data points at 1, 2, 3, 5,  and 
7 minutes post-dose are available; 
(6) 
For Treatment-B, at least four (4) of the five (5) PK data points at 9, 12, 15, 18, 
and 21 minutes post-dose are available;  
(7) 
For Treatment-C, at least four (4) of the five (5) PK data points at 25, 30, 40, 50, 
and 60 minutes post-dose are available;  
(8) 
For Treatment-A, at least three (3) of the PK data points at 25, 30, 40, and 50 
minutes post-dose are available.  
(9) 
For Treatment-B, at least three (3) of the PK data points at 80, 90, 120 minutes 
and 4 hours post-dose are available; and 
(10) 
For Treatment-C, at least three (3) of the PK data points at 6, 8, 12 and 18 hours 
post-dose are available. 
 
6.1.2. Treated Population (TP) 
 
The “treated” population is defined as all subjects who have been randomized and treated 
with any dose of the study drugs. 
 
The safety evaluation will be performed base on treated population (TP). 
 
6.1.3. Intent-to-treat Population (ITT) 
 
The “Intent-to-Treat” population is defined as all subjects who have been randomized. 
 

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6.2. PK Results Evaluations 
 
6.2.1 Calculation of Primary Endpoints 
 
 Cmax will be obtained directly by observing the plot of PK curve; 
 AUC0-6h and   AUC0-24h will be calculated with trapezoid method;  
 AUC0-∞ will be calculated with extrapolation method.  
 
6.2.2 Calculation of Secondary Endpoints 
 
 tmax is an observed values, at which Cmax is achieved.  
 
6.2.3 Missing Data Handling 
 
Missing data includes missing samples and missing data points for various cases, such as: 
 
(1) Missed scheduled points for sampling; 
(2) Tube breakage; 
(3) Inadequate amount of sample for analysis; 
(4) Unexpected reasons (i.e., medical condition such that the subject is unable to 
complete the study). 
 
A table of all missing samples will be provided with explanations. 
 
 
6.2.4 Relative Bioavailability 
The relative bioavailability of Treatment-A over Treatment-B (IM) can be defined as 
follows: 
      
       
      
 
      
 
  
 
6.3. Safety Assessment 
 
(1) Changes in vital signs and ECG will be analyzed between Screening, designated post-
dose time points and EOS;  
(2) Changes in serum potassium and glucose at study baseline, designated post-dose time 
points and EOS; 
(3) Physical examinations at Screening, and at the end of the last Dosing Visit, or study 
termination; 
(4) The results of laboratory tests, including CBC (with differential), metabolic panel and 
UA, will be assessed and compared between data at Screening and that at EOS; 

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(5) AEs will be observed and documented as described in Sections 5.3.7 and 7.2.1. AEs of 
each study treatment will be assessed per seriousness, severity, expected/unexpected, 
etc. The occurrence of AEs as absolute case number and relative ratio will be compared 
between each treatment.   
 
6.4. Interim Analyses 
 
No interim analyses are planned for this study.  
 
 

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7. STUDY MANAGEMENT 
 
7.1. Discontinued Subjects or Withdrawal 
 
Subjects can withdraw or be withdrawn from the study at any time. In addition, it may be 
necessary to discontinue subjects from the study due to medical safety considerations, 
noncompliance or administrative concerns.   
 
If for any reason a subject is withdrawn or discontinued before completing the study, the 
reasons for withdrawal or discontinuation must be entered on the End-of-Study and/or 
Subject Withdrawal or Discontinuation source document page.  
 
7.2. Adverse Events 
 
The Investigator will be required to provide the sponsor with any information concerning any 
findings that suggest any adverse event pertinent to the investigation. 
 
The study may be stopped at any time due to safety concerns. Subject safety will be 
constantly monitored throughout the study. However, if and when a significant safety 
concern arises, such as occurrence of serious and unexpected adverse drug events that are 
considered associated with the study drug(s) per regulatory guidelines and SOP, the study 
may be discontinued and safety will be reviewed.   
 
Any such safety analysis will be presented and discussed in the final study report.  
 
7.2.1. Types of Adverse Events 
 
 
The term “adverse event” could include any of the following events that arise, or increase 
in severity and/or frequency, during the course of the study: 
a) Any signs or symptoms whether thought to be related or unrelated to the condition 
under study. 
b) Any clinically significant laboratory abnormality. 
c) Any physical abnormality detected. 
 
These data will be recorded on the appropriate case report forms, including “Adverse Event 
Form”, regardless of whether they are thought to be associated with the study or the drug 
under investigation. Per FDA definition, “associated with the use of the drug” means that 
“there is a reasonable possibility that the experience may have been caused by the drug”. 
 
An adverse event is considered associated with the use of the drug if the attribution is 
possible, probable, or very likely according to definitions specified in the AE reporting 
SOP and the CRF for adverse event. 

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Signs or symptoms will be graded by the Investigator as mild, moderate or severe 
according to the following definitions: 
 
Severity 
  Definition 
Mild 
              Causing no limitation of usual activities 
Moderate 
Causing some limitation of usual activities 
Severe 
     Causing inability to carry out usual activities 
 
When a subject experiences any significant adverse event or laboratory abnormality, the 
event must be followed based on good clinical practice until it becomes resolved or stable. 
 
7.2.2. Definition of Serious and Unexpected Adverse Events 
 
According to the FDA guidelines, a serious adverse event will refer to any adverse drug 
experience occurring at any dose results in any of the following outcomes: 
 
a) Cancer [6]. 
b) Death. 
c) A life-threatening adverse drug experience. 
d) Hospitalization or prolongation of existing hospitalization. 
e) A persistent or significant disability/incapacity. 
f) A congenital anomaly/birth defect. 
g) Other important medical events, while not resulting in death, being life-threatening, 
or requiring hospitalization, may still be considered as serious adverse events when, 
based upon appropriate medical judgment, they may jeopardize the patient or subject 
and may require medical or surgical intervention to prevent one of the outcomes 
listed in this definition.   
 
An unexpected event is any adverse experience, the specificity or severity of which is not 
consistent with the approved labeling. “Unexpected”, thus refers to an adverse drug 
experience that has not been previously observed rather than from the perspective of such 
experience not being anticipated from the pharmacological properties of the pharmaceutical 
product. 
 
Expected adverse events of Primatene Mist include: headache, nausea, vomiting, 
nervousness, dizziness, shaking (tremor), trouble sleeping, stomach upset, sweating, dry 
mouth, bad taste, cough, sore throat, difficult or painful urination, fast heartbeat, weakness, 
high blood pressure, loss of appetite, loss of skin color, or dry throat. Severe adverse 
reactions are very rare. 
 
 
 

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7.2.3. Obligations of AE Reporting to Sites, FDA and IRB 
 
Sponsor Reporting to Investigator:  Sponsor shall promptly notify the Investigators of 
any material discoveries made during monitoring of the sites which could affect the safety 
of participants or influence the conduct of the study. 
 
Investigator Reporting to Sponsor:  Any serious events, whether or not unexpected or 
considered to be associated with the use of the drug, must be communicated by the 
Investigator to Amphastar Pharmaceuticals, Inc. immediately upon discovery of the event. 
The Clinical Research Associate (CRA) will then advise the Investigator regarding the 
nature of any further information or documentation that is required. 
 
Sponsor/Investigator Reporting to FDA: It is required to report to FDA any serious 
adverse event observed during the conduct of the study, regardless of whether the event is 
considered drug related.  
 
Each notification shall be made as soon as possible and in no event later than 15 calendar 
days after the sponsor’s initial receipt of the information. Each notification shall be 
submitted via the FDA Safety Reporting Portal (SRP). The FDA must be notified via 
telephone of the Sponsor’s initial notification of any fatal or life-threatening experiences 
associated with the use of the drug as soon as possible but in no event later than seven (7) 
calendar days. 
 
Sponsor/Investigator Reporting to IRB:  All serious adverse events or an unanticipated 
problem must be reported to the appropriate IRB/Ethics Committee promptly. 
 
1. 
Serious Adverse Event (SAE) Reporting Policy: 
An adverse event is considered serious if it results in any of the outcomes listed in 
Section 7.2.2.  Study sites are required to report any SAE to the IRB, regardless of 
whether the event is considered drug related. The specific reporting policy defined by 
the IRB will be followed for the reporting of SAE to IRB within the required time 
period. 
 
Note: IRB does not require the event to be unexpected. The IRB feels that this category 
is too subjective, and can change over time, to be a criterion for deciding whether the 
event is reportable or not. 
 
2. 
Unanticipated Problem/Event Report Form: 
In order for the event to be reportable to the IRB, it must meet all of the following 
criteria: 
 The event is unexpected (in terms of nature, severity, or frequency) given the 
research procedures described in the study Protocol and/or Informed Consent 

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Form. 
 The event is related or possibly related to participation in the research study. 
 The event suggests that the research places subjects or others at a greater risk of 
harm than was previously realized or anticipated. 
 
It is required to report unanticipated problem(s) to the IRB promptly after the 
Investigator becomes aware of them. The specific reporting policy defined by the IRB 
will be followed for the reporting of unanticipated problem to IRB within the required 
time period. 
 
Sponsors are specifically required to “keep each participating Investigator informed of 
new observations discovered by or reported to the sponsor on the drug, particularly 
with respect to adverse effects and safe use”.  
 
7.2.4. Follow-up of Adverse Events 
 
The Sponsor’s CRA will periodically request follow-up information of the AE from the PI. 
The CRA will ensure that all fatal or serious adverse events are reported to the IRB, per 
Amphastar SOP. All serious adverse events must be followed up for at least 30 days or 
until resolved or having become stable.  
 
The sponsor shall promptly investigate all safety information received by it from the 
clinical studies. Follow up information to a safety report shall be submitted as soon as the 
relevant information becomes available.   
 
7.3. Clinical Laboratory Test and Normal Laboratory Ranges 
 
Each standard clinical laboratory test, i.e., CBC, serum comprehensive metabolic panel and 
urinalysis, will be performed by a single laboratory for all subjects enrolled in the study. The 
laboratory that will perform these tests required by the protocol must transmit normal ranges 
of the clinical laboratory tests to Amphastar Pharmaceuticals, Inc. Urinary pregnancy and 9 
panel drug tests may be performed using a clinically accepted test kit. 
 
 
 

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8. STUDY MONITORING, DOCUMENTATION AND RECORD KEEPING 
 
8.1. Data Quality Assurance and Monitoring 
 
The trial will be monitored according to the current Amphastar SOP.  
 
The site monitor will conduct monitoring of the investigational activities for the purpose of 
ensuring subject safety and study compliance to applicable regulatory guidelines. The 
Investigator will permit Amphastar authorized monitors to access the subject source 
documents, eCRFs, clinical supplies dispensing and storage area and study documentation as 
frequently as necessary and agrees to assist the site monitors with their activities. The study 
site will make the eCRF available, provide missing or correct data and PI will sign the 
eCRFs. Personal or subject identifying information will be treated as confidential and will 
NOT be publicly accessible. 
 
The trial information may be reviewed by regulatory authorities or independent QA auditors. 
The study site may be inspected during or after completing the study. The Investigators agree 
to allow inspectors from regulatory agencies to have access to all trial records, including 
subject source documents. By participating in this study, the Investigator 
 
8.2. Electronic Case Report Forms (eCRFs) 
 
Electronic Data Capture (EDC) system will be used for recording of electronic CRF (eCRF) 
for the conduct of the study by Amphastar, eCRF will be used for the purpose of capturing 
data for all consented participants in the study. The 24 hour clock will be used for all time 
entries. 
 
Subject source documents are the study subject records maintained at the study site. In some 
cases, the source documents may be the hospital’s or the physician’s clinical records. 
Therefore, the information collected on the eCRFs must match those records. Amphastar 
personnel will review all eCRFs to ensure the data is verifiable and accurate. At all times, the 
PI has final responsibility for the accuracy and authenticity of all clinical and laboratory data 
entered on the eCRFs. The eCRFs will be completed as soon as possible after the data 
become available. 
 
 
8.3. Clinical Trial Drug Accountability Data 
 
The Study Site Research Pharmacist or designated dispenser acknowledges that the drug 
supplies are investigational and, as such, must be handled in accordance with the protocol 
and labeling. The Research Pharmacist or designated personnel will maintain adequate 

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records showing the receipt, dispensing, return, or other disposition of the investigational 
study material, including the date, quantity, lot or code number, and identification of subjects 
(number, initials) who received or returned study drug.  
 
The Research Pharmacist or designated dispenser and Amphastar’s unblinded site monitor 
will review study supply records, including dispensing and accountability records, at 
monitoring visits.  
 
It is the responsibility of the Investigator and research pharmacist/designated dispenser to 
ensure that, when the study is completed or discontinued, unused study supplies will be 
returned or disposed of, as directed by Amphastar’s site monitors or CRA. Unused and 
returned medication must be available for verification by Amphastar’s site monitors or CRA.  
 
8.4. Trial Documents 
 
The trial documents, including the protocol signature page signed by the Investigators, 
Investigators’ curriculum vitae, IRB approval notice and IRB member list, approved 
Informed Consent Form, laboratory verification and institutional accreditation, will be 
submitted to Amphastar Pharmaceuticals, Inc. 
 
8.5. Reporting and Publication 
 
The clinical trial report will be prepared according to the Amphastar SOP for Clinical Trial 
Reporting.  
 
8.6. Archive 
 
The Investigator will retain a copy of all study documents, including reports to the 
IRB/Ethics Committee and to the sponsor, in accordance with the FDA and local regulations. 
The FDA regulations and/or ICH guidelines state that the Investigator must maintain study 
documents for a set period of time. The Investigator will arrange for the retention of the 
Subject Identification for at least 15 years after the Final Study Report has been signed. All 
other documents should be kept for the maximum period of time permitted by the institution. 
 
 
 

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9. ETHICS 
 
9.1. Institutional Review Board (IRB) or Ethics Committee Approval 
 
This study will be conducted in conformance with all regulatory guidelines, particularly the 
FDA guidance on MDI and DPI drug products [1]. This trial can be conducted only after full 
approval of the final protocol (and its addendum) has been obtained from the involved 
Institutional Review Board (IRB) or Ethics Committee. The approval must be received by 
Amphastar before the initiation of the study. 
 
9.2. ICH GCP Guidelines and Declaration of Helsinki 
 
This trial will be undertaken in accordance with the current ICH Good Clinical Practice 
(GCP) guidelines and adheres to the core principles of the Declaration of Helsinki of the 
World Medical Association to assure that the rights, safety and well-being of the study 
subjects are protected. 
 
9.3. Informed Consent 
 
The Investigator must explain to subjects the purpose, methods, potential benefits and risks 
of the trial. Subjects will be informed that they are free not to participate in the trial and they 
may withdraw from the study at any time. Written informed consent from each subject 
enrolled in the study, in accordance with the Code of Federal Regulations Title 21, Part 50, 
Subpart B will be recorded by means of dated signatures of the subjects.  
 
Informed Consent will include authorization to use data from the trial for all purposes 
relating to approval of the drug under investigation. 
 
9.4. HIPAA Compliance 
 
The Investigator is responsible for compliance with the privacy provisions of the Health 
Insurance Portability and Accountability Act including obtaining informed consent for use of 
data for marketing purposes.  
 
 
 

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10. REFERENCES 
 
[1]  FDA Draft Guidance for Industry Metered Dose Inhaler (MDI) and Dry Powder Inhaler 
(DPI) Drug Products - Quality Considerations, 04/2018.  
[2]  Instructions for use from Epinephrine Injection prescribing information, Revised 9/2018.  
[3]  Warren JB, Doble N, etc: Systemic absorption of inhaled epinephrine. Clin Pharmacol Ther, 
40:673-678, 1986.  
[4]  Westfall TC, Westfall DP: Adrenergic agonists and antagonists, in Brunton LL, Lazo JS, 
Parker KL (eds): Goodman & Gilman’s The Pharmacological Basis of Therapeutics. 11th 
Ed. P 237-296., 2006.  
[5]  Dauphinee B, Tashkin DP, et al: Placebo-controlled evaluation of the speed of onset of 
epinephrine metered-dose aerosol (Primatene® Mist) in mild to moderate asthmatics. Am J 
Respir Crit Care Med, 149:A204, 1994.  
[6]  American Thoracic Society, "Using Your Metered Dose," Am J Respir Crit Care Med, vol. 
190, pp. 5-6, 2014.  
[7]  CDISC SDTM Implementation Guide (Version 3.2). 11/26/2013.  
[8]  Prescribing Information for ProAir HFA Inhalation Aerosol (
), revised 06/2016.  
[9]  FDA Guidance for Industry and Investigators: Safety Reporting Requirements for INDs and 
BA/BE Studies, December 2012.  
[10] Guidance for Industry: Handling and Retention of BA and BE Testing Samples and 
Retention 
http://www.fda.gov/downloads/RegulatoryInformation/Guidances/UCM126836.pdf.  
[11] FDA Draft Guidance on Albuterol Sulfate, Recommended Apr 2013; Revised Jun 2013; 
Dec 2016.  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

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Appendix I. Approval by an IRB/Ethics Committee 
 
In performing this study, both the Investigator and Sponsor endorse, as a minimum, the standards for 
conduct of clinical research activities as set forth in the FDA regulations and/or adhere to the core 
principles of the Declaration of Helsinki and local country laws and regulations. 
 
The Investigator will submit the protocol, any amendments, and informed consent for IRB/Ethics 
Committee approval prior to initiation of the study. This will be appropriately documented. The 
IRB/Ethics Committee must give their approval in writing. The names and qualifications of the members 
of the review committee will be recorded and submitted to the sponsor with the written approval for the 
conduct of the study. The members of the IRB/Ethics Committee approving the protocol must be 
independent of the Sponsor and the Investigator. The written approval will consist of written 
documentation from the IRB/Ethics Committee containing the approval information. 
 
Until written approval by the IRB/Ethics Committee has been received by the Investigator, no subject 
may undergo any procedures solely for the purpose of determining eligibility for this study. 
 
Protocol amendments must also be reviewed and approved by the IRB/Ethics Committee. Written 
approval from the IRB/Ethics Committee, or a designee, must be received by the sponsor, Amphastar 
Pharmaceuticals, Inc. before implementation. This written approval will consist of documentation from 
the IRB/Ethics Committee containing the approval information. 
 
 
 

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Protocol Signature Page 
 
I confirm that I have read this protocol, I understand it, and I will work according to this protocol and to 
the ethical principles stated in the ICH Good Clinical Practice guideline and adhere to the core principles 
of the Declaration of Helsinki, or the applicable local laws and regulations of the study site for which I am 
responsible, whichever provides the greater protection of the individual. I will abide by the publication 
rules set forth in my agreement with Amphastar Pharmaceuticals, Inc. I will accept the monitor's 
inspection and overseeing of the study. I will promptly submit the protocol to the applicable Institutional 
Review Board/Ethical Committee for approval. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Signature of Investigator  
 
 
 
 
Date 
 
 
 
 
 
 
 
 
 
  
Investigator Name (Print) 
 
 
 
 
 
 
 
 
 
Title 
 
 
 
 
 
 
 
 
 
Name of Facility 
 
 
 
 
 
 
 
 
 
Street Address 
 
 
 
 
 
 
 
 
 
City, State, ZIP Code 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Signature of Representative of   
 
 
             Date 
Amphastar Pharmaceuticals, Inc. 
 
 

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Appendix II. Body Mass Index (BMI) Table 
 
BMI Formula: 
 
BMI=Weight [in kilograms]/ (Height [in meters]) 2   Or 
 
BMI=Weight [in pounds] X703.7/ (height [in inches]) 2 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

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Appendix III. Prohibited Drug List 
 
*This drug list is provided based on the drug interaction listed in package insert of Epi-Pen. Some drugs that have 
been discontinued per FDA website will not be listed here. Asthma medications are also not listed because this study 
is for healthy volunteers only. 
Drug Category
Major 
Indications
Type of 
Impact
Description of 
Impact
Route
Brand Name
Generic Name
Washout 
Period*
1
Anafranil
Clomipramine 
2
Asendin
Amoxapine
3
Norpramin
Desipramine
4
Pamelor
Nortriptyline
5
Sinequan
Doxepin
6
Surmontil
Trimipramine
7
Tofranil
Imipramine
8
PO/IV Elavil
Amitriptyline
9
Desyrel
Trazodone
10
PO
Ludiomil
Maprotiline
11
Serzone
Nefazodone
12
Benadryl
Diphenhydramine
13
Chlortrimeton Chlorpheneramine
14
Cardura
Doxazosin
3 days
15
Dibenzyline
Phenoxybenzamin
5 days
16
Flomax
Tamsulosin
3 days
17
PO
Hytrin
Terazosin
3 days
18
Minipress
Prazosin
3 days
19
Rapaflo
Silodosin
3 days
20
Uroxatral
Alfluzosin
3 days
21
PO/IV Betapace
Sotolal
22
Blocadren
Timolol
23
PO
Bystolic
Nebivolol
24
Coreg
Carvediol
25
PO/IV Inderal
Propranolol
26
PO
Lopressor
Metoprolol
27
PO
Tenormin
Atenolol
28
PO/IV Trandate
Labetalol
29
PO
Zebeta
Bisprolol
30 Cardiac Glycoside
Congestive 
heart failure
Safety 
May induce 
cardiac 
arrhythmia with 
Primatene; 
Albuterol may 
↓digoxin level
PO
Lanoxin
Digoxin
10 days
31
Aldactone
Spironolactone
32
Bumex
Bumetanide
33
Diuril
Chlorothiazide
34
Demadex
Torsemide
35
PO
HydroDiuril
Hydrochlorothiazid
3 days
36
Hygroton
Chlorthalidone
37
Lasix
Furosemide
38
Lozol
Indapamide
3 days
39
Zaroxolyn
Metolazone
1 day
Diuretics
PO
1 day
1 day
ECG changes/ 
Hypokalemia/ 
hyperkalemia
Anti-depressant 
(Tricyclic)
Anti-depressant 
(Polycyclic)
Depression
Safety
May ↑the 
cardiovascular 
side effects
PO
4 weeks
Potentiate side 
effect of 
Primatene and 
Albuterol
Alpha1-blockers
Hypertension/ 
Prostate 
enlargement
PK
↑PK level by 
competing for α-
receptors
Antihistamine
Allergy
Safety
β1/β2-Blockers     
(Selective & Non-
selective)
Hypertension/ 
angina
Safety     
PD
↓Effect of 
Primatene and 
Albuterol, also 
affect safety in 
QTc prolongation
6 hours
4 weeks
Edema/ 
Hypertension
Safety

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Appendix III. Prohibited Drug List (Continued) 
 
 
*This drug list is provided based on the drug interaction listed in package insert of Epi-Pen. Some drugs that have 
been discontinued per FDA website will not be listed here. Asthma medications are also not listed because this study 
is for healthy volunteers only. 
 
 
Drug Category
Major          
Indications
Type of 
Impact
Description of 
Impact
Route
Brand Name
Generic Name
Washout 
Period*
40
Marplan
Isocaarboxazid
41
Nardil
 Phenelzine
42
Parnate
Tranylcypromine
43
Eldepryl
Selegiline
44
Zelapar
Selegiline
45
Xadago
Safinamide
46
Adderall
2 days
47
Adderall XR
5 days
48
Dexedrine
Dextroamphetamine 4 days
49
Ritalin
Methylphenidate
2 days
50
Ritalin LA
Methylphenidate
5 days
51
Vyvanse
Lisdexamfetamine
1 day
52
Anaphylaxis
IV/IM/
SC
Adrenalin
Epinephrine 
1 day
53
Afrin
Oxymetazoline
2 days
54
Neo-synephrinePhenlephrine
1 day
55
Advil Cold & 
Sinus
Ibuprofen/ 
Pseudoephedrine
56
Allegra D
Fexofenadine/ 
Pseudoephedrine
57
Claritin D
Loratadine/ 
Pseudoephedrine
58
Allergy/Nasal & 
Chest  
Congestion
Mucinex D
Guaifenesin/ 
Pseudoephedrine
59
Sudafed/PE
Phenylephrine
60
Sudafed/SA
Pseudoephedrine
61
Allergy/Nasal 
congestion
Zyrtec D
Cetirizine/ 
Pseudoephedrine
62
Nausea & 
Vomiting
Compazine
Prochlorperazine
63
Etrafon*
Perphenazine/ 
amitriptyline
64
Mellaril*
Thioridazine
65
Serentil*
Mesoridazine
Stelazine*
Trifluoperazine
66
PO/IV Thorazine*
Chlorpromazine
67
Thyroid        
hormones
Hypothyroidism 
Safety
↑ cardiac effects 
of Primatene 
and Albuterol
PO
Synthroid
Levothyroxine
6 weeks
14 days
Monoamine 
Oxidase-B Inhibitor
Parkinsonism
Oral Decongestant  
/Oral or Parenteral 
Sympathomimetics
Attention        
Deficit 
Hyperactivity 
Disorder         
(ADHD)
PK/       
Safety
↑PK level and 
↑cardiac side 
effects of 
Primatene and 
Albuterol
PO
Dextroamphetamine 
mixed salts
Nasal           
Congestion
Nasal
Allergy/nasal 
congestion
↑ PK level by 
competing α-
receptor and 
↑cardiac side 
effects of 
Primatene and 
Albuterol
PO
1 day
Nasal      
Congestion
Monoamine 
Oxidase-A 
Inhibitor
Major           
depression
PK/       
Safety
↑PK  and cardiac 
side effects of 
Primatene and 
Albuterol
PO
Phenothiazines
Safety
May  potentiate 
side effect of 
Primatene and 
Albuterol
PO
7 days
Schizophrenia

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Appendix IV. Blood Metabolic/CBC and Urinalysis Tests 
 
Based on the ICH guidelines (i) Guideline for Good Clinical Practice E6 (R1), version 4, dated 10 June 
1996; and (ii) Safety Pharmacology Studies for Human pharmaceuticals S7A, version 4, dated 8 
November 2000. For safety assessment, the following items will be conducted at screening.  
 
* Normal value ranges may vary slightly among different laboratories, the value listed here are only for 
reference, the actual values will be determined by study site.                                                                                                    
 
 
 
#
Test Categories
Description
#
Test Items
*Reference 
Normal Range
Footnote
1
Total protein  
6.2 to 8.3 g/dL
Albumin/globulin
2
Sodium
133 to 145 
3
Potassium
3.4 to 5.4 mEq/L
4
Calcium
8.4 to 10.6 mg/dL
5
Chloride
96 - 110 mmol/L
6
Glucose
60 to 99 mg/dL
7
CO2
22 to 32 mEq/L
8
BUN 
6 to 21 mg/dL
Blood urea nitrogen
9
Creatinine
0.4 to 1.54 mg/dL 
10 ALP
35 to 104 U/L
Alkaline Phosphatase
11 ALT
0 to 40 U/L
Alanine Transaminase
12 AST
10 to 35 U/L
Aspartate Transaminase  
13 Bilirubin
0.0 to 1.4 mg/dL
14 Color
yellow
15 Character (Clarity)
clear
16 Specific gravity
1.003-1.029
17 PH
4.5 - 8.0
18 Protein 
negative
19 Glucose
negative
20 Nitrite
negative
21 Ketone
negative
22 Leukocyte esterase
negative
23 Blood
negative
24 Bilirubin
negative
25 Crystals
negative
26 Bacteria/Yeast
negative
27 RBC/WBC/Epithelial cells
negative
28 Casts
negative
29 RBC
3.9-5.2 ×106/µl  
Red blood cells
30 MCV  
80 to 100 fL
Mean Corpuscular Volume
31 MCH 
26 to 34 pg/cell
Mean Corpuscular Hemoglobin
32 MCHC
32 to 37 g/dL
Mean Corpuscular Hemoglobin 
Concentration
33 Hemoglobin
11.0-15.5 g/dL
34 Hematocrit
34.0-45.0 %
35 Platelet
150-450 ×103/µl
36 WBC  
4.3-11.0 ×103/µl
White Blood Cell
1
Comprehensive 
Metabolic Panel 
≥ 8 hr 
fasting
2
Urine analysis, 
routine and 
microscopic
fasting not 
required
3
 CBC with 
differentials 
ibid

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Appendix IV. Blood Metabolic/CBC and Urinalysis Tests (Continued) 
 
 
* Normal value ranges may vary slightly among different laboratories, the value listed here are only for 
reference, the actual values will be determined by study site.                                                                                                    
** All female subjects 
 
 
 
 
 
 
 
 
 
 
 
#
Test Categories
Description
#
Test Items
*Reference 
Normal Range
Footnote
4 Urinary or serum 
pregnancy test** 
ibid
37 Pregnancy
negative
38 amphetamines
negative
39 cannabinoids
negative
40 methadone
negative
41 barbiturates
negative
42 PCP
negative
43 opiates
negative
44 benzodiazepines
negative
45 cocaine/metabolites
negative
46 methamphetamines
negative
47 alcohol
negative
48 HIV Ab 
negative
Human Immunodeficiency Virus Antibody
49 HBs Ag
negative
Hepatitis B Surface Antigen
50 HCV Ab
negative
Hepatitis C Virus Antibody
51 Free T3
200-500 pg/dL
Triiodothyronine
52 Total T4
4.5-11.2 μg/dL
Thyroxine
53 TSH
0.4 - 4.0 mIU/L
Thyroid Stimulating Hormone
5
Urinary 
alcohol/nine 
panel drug 
screen tests 
ibid
* Normal value ranges may vary among different laboratories, the value listed here are only for reference, the actual values will 
be determined by study site.                                                                                                                                                      
** all female subjects
6
Tests for 
Infectious 
diseases
fasting not 
required
7 Thyroid Function fasting not 
required