#METABOLOMICS WORKBENCH epannkuk_20210518_084258_mwtab.txt DATATRACK_ID:2646 STUDY_ID:ST001806 ANALYSIS_ID:AN002928 PROJECT_ID:000000 VERSION 1 CREATED_ON May 24, 2021, 9:34 am #PROJECT PR:PROJECT_SUMMARY An important component of ionizing radiation (IR) exposure after a radiological PR:PROJECT_SUMMARY incident may include low-dose rate (LDR) exposures either externally or PR:PROJECT_SUMMARY internally, such as from 137Cs deposition. LDR exposures can have different PR:PROJECT_SUMMARY effects compared to acute high-dose rate exposures from a health and PR:PROJECT_SUMMARY biodosimetry perspective. In this study, a novel irradiation system, VAriable PR:PROJECT_SUMMARY Dose-rate External 137Cs irradiatoR (VADER), was used to expose male and female PR:PROJECT_SUMMARY mice to a variable LDR over a 30-day time span to cumulative doses of 1 (only in PR:PROJECT_SUMMARY males), 2, 2.8, 4.1, 8.8 (only in males), or 9.7 Gy to simulate fall-out type PR:PROJECT_SUMMARY exposures. Urine and serum from mice exposed to an acute dose (~0.8 Gy/min) of PR:PROJECT_SUMMARY x-rays were collected in parallel. Radiation markers were identified by global PR:PROJECT_SUMMARY mass spectrometry based metabolomics and the machine learning algorithm Random PR:PROJECT_SUMMARY Forests. PR:INSTITUTE Georgetown University PR:FIRST_NAME Evan PR:ADDRESS 3970 Reservoir Rd, NW New Research Building E504 PR:EMAIL elp44@georgetown.edu PR:PHONE 2026875650 PR:PROJECT_TITLE Effect of external low-dose rate radiation on mouse biofluid metabolomics PR:LAST_NAME Pannkuk #STUDY ST:STUDY_TITLE Effect of external low-dose rate radiation on mouse biofluid metabolomic ST:STUDY_TITLE signatures (part V) ST:STUDY_SUMMARY An important component of ionizing radiation (IR) exposure after a radiological ST:STUDY_SUMMARY incident may include low-dose rate (LDR) exposures either externally or ST:STUDY_SUMMARY internally, such as from 137Cs deposition. LDR exposures can have different ST:STUDY_SUMMARY effects compared to acute high-dose rate exposures from a health and ST:STUDY_SUMMARY biodosimetry perspective. In this study, a novel irradiation system, VAriable ST:STUDY_SUMMARY Dose-rate External 137Cs irradiatoR (VADER), was used to expose male and female ST:STUDY_SUMMARY mice to a variable LDR over a 30-day time span to cumulative doses of 1 (only in ST:STUDY_SUMMARY males), 2, 2.8, 4.1, 8.8 (only in males), or 9.7 Gy to simulate fall-out type ST:STUDY_SUMMARY exposures. Urine and serum from mice exposed to an acute dose (~0.8 Gy/min) of ST:STUDY_SUMMARY x-rays were collected in parallel. Radiation markers were identified by global ST:STUDY_SUMMARY mass spectrometry based metabolomics and the machine learning algorithm Random ST:STUDY_SUMMARY Forests. ST:INSTITUTE Georgetown University ST:LAST_NAME Pannkuk ST:FIRST_NAME Evan ST:ADDRESS 3970 Reservoir Rd, NW New Research Building E504 ST:EMAIL elp44@georgetown.edu ST:PHONE 2026875650 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:GENDER Male #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Raw file names and additional sample data SUBJECT_SAMPLE_FACTORS 1 114V Factor:LDR | Factor:D5 RAW_FILE_NAME=NEG_012 SUBJECT_SAMPLE_FACTORS 3 93A Factor:Control | Factor:D3 RAW_FILE_NAME=NEG_013 SUBJECT_SAMPLE_FACTORS 4 63A Factor:HDR | Factor:D1 RAW_FILE_NAME=NEG_014 SUBJECT_SAMPLE_FACTORS 5 113V Factor:LDR | Factor:D5 RAW_FILE_NAME=NEG_015 SUBJECT_SAMPLE_FACTORS 6 123V Factor:LDR | Factor:D20 RAW_FILE_NAME=NEG_016 SUBJECT_SAMPLE_FACTORS 9 159V Factor:Control | Factor:D30 RAW_FILE_NAME=NEG_019 SUBJECT_SAMPLE_FACTORS 11 99A Factor:Control | Factor:D5 RAW_FILE_NAME=NEG_020 SUBJECT_SAMPLE_FACTORS 13 153V Factor:Control | Factor:D20 RAW_FILE_NAME=NEG_021 SUBJECT_SAMPLE_FACTORS 14 151V Factor:Control | Factor:D20 RAW_FILE_NAME=NEG_022 SUBJECT_SAMPLE_FACTORS 16 89A Factor:Control | Factor:D2 RAW_FILE_NAME=NEG_023 SUBJECT_SAMPLE_FACTORS 17 87A Factor:Control | Factor:D2 RAW_FILE_NAME=NEG_029 SUBJECT_SAMPLE_FACTORS 18 90A Factor:Control | Factor:D2 RAW_FILE_NAME=NEG_030 SUBJECT_SAMPLE_FACTORS 20 115V Factor:LDR | Factor:D5 RAW_FILE_NAME=NEG_031 SUBJECT_SAMPLE_FACTORS 21 72A Factor:HDR | Factor:D3 RAW_FILE_NAME=NEG_032 SUBJECT_SAMPLE_FACTORS 22 186V Factor:Control | Factor:D3 RAW_FILE_NAME=NEG_033 SUBJECT_SAMPLE_FACTORS 23 77A Factor:HDR | Factor:D5 RAW_FILE_NAME=NEG_036 SUBJECT_SAMPLE_FACTORS 25 100A Factor:Control | Factor:D5 RAW_FILE_NAME=NEG_037 SUBJECT_SAMPLE_FACTORS 27 172V Factor:LDR | Factor:D2 RAW_FILE_NAME=NEG_038 SUBJECT_SAMPLE_FACTORS 30 130V Factor:LDR | Factor:D30 RAW_FILE_NAME=NEG_039 SUBJECT_SAMPLE_FACTORS 31 126V Factor:LDR | Factor:D30 RAW_FILE_NAME=NEG_040 SUBJECT_SAMPLE_FACTORS 33 75A Factor:HDR | Factor:D3 RAW_FILE_NAME=NEG_046 SUBJECT_SAMPLE_FACTORS 34 171V Factor:LDR | Factor:D2 RAW_FILE_NAME=NEG_047 SUBJECT_SAMPLE_FACTORS 37 158V Factor:Control | Factor:D30 RAW_FILE_NAME=NEG_048 SUBJECT_SAMPLE_FACTORS 38 176V Factor:Control | Factor:D2 RAW_FILE_NAME=NEG_049 SUBJECT_SAMPLE_FACTORS 39 156V Factor:Control | Factor:D30 RAW_FILE_NAME=NEG_050 SUBJECT_SAMPLE_FACTORS 40 152V Factor:Control | Factor:D20 RAW_FILE_NAME=NEG_053 SUBJECT_SAMPLE_FACTORS 42 70A Factor:HDR | Factor:D2 RAW_FILE_NAME=NEG_054 SUBJECT_SAMPLE_FACTORS 44 95A Factor:Control | Factor:D3 RAW_FILE_NAME=NEG_055 SUBJECT_SAMPLE_FACTORS 45 157V Factor:Control | Factor:D30 RAW_FILE_NAME=NEG_056 SUBJECT_SAMPLE_FACTORS 47 5V Factor:LDR | Factor:D1 RAW_FILE_NAME=NEG_057 SUBJECT_SAMPLE_FACTORS 48 178V Factor:Control | Factor:D2 RAW_FILE_NAME=NEG_063 SUBJECT_SAMPLE_FACTORS 49 67A Factor:HDR | Factor:D2 RAW_FILE_NAME=NEG_064 SUBJECT_SAMPLE_FACTORS 50 112V Factor:LDR | Factor:D5 RAW_FILE_NAME=NEG_065 SUBJECT_SAMPLE_FACTORS 51 2V Factor:LDR | Factor:D1 RAW_FILE_NAME=NEG_066 SUBJECT_SAMPLE_FACTORS 52 96A Factor:Control | Factor:D5 RAW_FILE_NAME=NEG_067 SUBJECT_SAMPLE_FACTORS 53 160V Factor:Control | Factor:D30 RAW_FILE_NAME=NEG_070 SUBJECT_SAMPLE_FACTORS 55 97A Factor:Control | Factor:D5 RAW_FILE_NAME=NEG_071 SUBJECT_SAMPLE_FACTORS 58 84A Factor:Control | Factor:D1 RAW_FILE_NAME=NEG_072 SUBJECT_SAMPLE_FACTORS 59 122V Factor:LDR | Factor:D20 RAW_FILE_NAME=NEG_073 SUBJECT_SAMPLE_FACTORS 61 174V Factor:LDR | Factor:D2 RAW_FILE_NAME=NEG_074 SUBJECT_SAMPLE_FACTORS 63 88A Factor:Control | Factor:D2 RAW_FILE_NAME=NEG_080 SUBJECT_SAMPLE_FACTORS 65 173V Factor:LDR | Factor:D2 RAW_FILE_NAME=NEG_081 SUBJECT_SAMPLE_FACTORS 66 69A Factor:HDR | Factor:D2 RAW_FILE_NAME=NEG_082 SUBJECT_SAMPLE_FACTORS 69 154V Factor:Control | Factor:D20 RAW_FILE_NAME=NEG_083 SUBJECT_SAMPLE_FACTORS 70 82A Factor:Control | Factor:D1 RAW_FILE_NAME=NEG_084 SUBJECT_SAMPLE_FACTORS 71 68A Factor:HDR | Factor:D2 RAW_FILE_NAME=NEG_087 SUBJECT_SAMPLE_FACTORS 72 71A Factor:HDR | Factor:D3 RAW_FILE_NAME=NEG_088 SUBJECT_SAMPLE_FACTORS 73 183V Factor:LDR | Factor:D3 RAW_FILE_NAME=NEG_089 SUBJECT_SAMPLE_FACTORS 74 3V Factor:LDR | Factor:D1 RAW_FILE_NAME=NEG_090 SUBJECT_SAMPLE_FACTORS 75 155V Factor:Control | Factor:D20 RAW_FILE_NAME=NEG_091 SUBJECT_SAMPLE_FACTORS 76 175V Factor:LDR | Factor:D2 RAW_FILE_NAME=NEG_097 SUBJECT_SAMPLE_FACTORS 78 62A Factor:HDR | Factor:D1 RAW_FILE_NAME=NEG_098 SUBJECT_SAMPLE_FACTORS 79 190V Factor:Control | Factor:D3 RAW_FILE_NAME=NEG_099 SUBJECT_SAMPLE_FACTORS 80 141V Factor:Control | Factor:D5 RAW_FILE_NAME=NEG_100 SUBJECT_SAMPLE_FACTORS 81 85A Factor:Control | Factor:D1 RAW_FILE_NAME=NEG_101 SUBJECT_SAMPLE_FACTORS 82 177V Factor:Control | Factor:D2 RAW_FILE_NAME=NEG_104 SUBJECT_SAMPLE_FACTORS 83 78A Factor:HDR | Factor:D5 RAW_FILE_NAME=NEG_105 SUBJECT_SAMPLE_FACTORS 84 64A Factor:HDR | Factor:D1 RAW_FILE_NAME=NEG_106 SUBJECT_SAMPLE_FACTORS 85 184V Factor:LDR | Factor:D3 RAW_FILE_NAME=NEG_107 SUBJECT_SAMPLE_FACTORS 89 143V Factor:Control | Factor:D5 RAW_FILE_NAME=NEG_108 SUBJECT_SAMPLE_FACTORS 90 94A Factor:Control | Factor:D3 RAW_FILE_NAME=NEG_114 SUBJECT_SAMPLE_FACTORS 91 4V Factor:LDR | Factor:D1 RAW_FILE_NAME=NEG_115 SUBJECT_SAMPLE_FACTORS 92 180V Factor:Control | Factor:D2 RAW_FILE_NAME=NEG_116 SUBJECT_SAMPLE_FACTORS 93 80A Factor:HDR | Factor:D5 RAW_FILE_NAME=NEG_117 SUBJECT_SAMPLE_FACTORS 95 121V Factor:LDR | Factor:D20 RAW_FILE_NAME=NEG_118 SUBJECT_SAMPLE_FACTORS 96 66A Factor:HDR | Factor:D2 RAW_FILE_NAME=NEG_121 SUBJECT_SAMPLE_FACTORS 97 187V Factor:Control | Factor:D3 RAW_FILE_NAME=NEG_122 SUBJECT_SAMPLE_FACTORS 101 86A Factor:Control | Factor:D2 RAW_FILE_NAME=NEG_123 SUBJECT_SAMPLE_FACTORS 104 92A Factor:Control | Factor:D3 RAW_FILE_NAME=NEG_124 SUBJECT_SAMPLE_FACTORS 105 185V Factor:LDR | Factor:D3 RAW_FILE_NAME=NEG_125 SUBJECT_SAMPLE_FACTORS 106 179V Factor:Control | Factor:D2 RAW_FILE_NAME=NEG_131 SUBJECT_SAMPLE_FACTORS 110 144V Factor:Control | Factor:D5 RAW_FILE_NAME=NEG_132 SUBJECT_SAMPLE_FACTORS 111 189V Factor:Control | Factor:D3 RAW_FILE_NAME=NEG_133 SUBJECT_SAMPLE_FACTORS 112 124V Factor:LDR | Factor:D20 RAW_FILE_NAME=NEG_134 SUBJECT_SAMPLE_FACTORS 113 33V Factor:Control | Factor:D1 RAW_FILE_NAME=NEG_135 SUBJECT_SAMPLE_FACTORS 114 73A Factor:HDR | Factor:D3 RAW_FILE_NAME=NEG_138 SUBJECT_SAMPLE_FACTORS 117 81A Factor:Control | Factor:D1 RAW_FILE_NAME=NEG_139 SUBJECT_SAMPLE_FACTORS 118 128V Factor:LDR | Factor:D30 RAW_FILE_NAME=NEG_140 SUBJECT_SAMPLE_FACTORS 119 127V Factor:LDR | Factor:D30 RAW_FILE_NAME=NEG_141 SUBJECT_SAMPLE_FACTORS 120 111V Factor:LDR | Factor:D5 RAW_FILE_NAME=NEG_142 SUBJECT_SAMPLE_FACTORS 121 65A Factor:HDR | Factor:D1 RAW_FILE_NAME=NEG_148 SUBJECT_SAMPLE_FACTORS 123 1V Factor:LDR | Factor:D1 RAW_FILE_NAME=NEG_149 SUBJECT_SAMPLE_FACTORS 126 181V Factor:LDR | Factor:D3 RAW_FILE_NAME=NEG_150 SUBJECT_SAMPLE_FACTORS 127 142V Factor:Control | Factor:D5 RAW_FILE_NAME=NEG_151 SUBJECT_SAMPLE_FACTORS 130 182V Factor:LDR | Factor:D3 RAW_FILE_NAME=NEG_152 SUBJECT_SAMPLE_FACTORS 131 188V Factor:Control | Factor:D3 RAW_FILE_NAME=NEG_155 SUBJECT_SAMPLE_FACTORS 133 145V Factor:Control | Factor:D5 RAW_FILE_NAME=NEG_156 SUBJECT_SAMPLE_FACTORS 134 83A Factor:Control | Factor:D1 RAW_FILE_NAME=NEG_157 SUBJECT_SAMPLE_FACTORS 135 76A Factor:HDR | Factor:D5 RAW_FILE_NAME=NEG_158 SUBJECT_SAMPLE_FACTORS 136 129V Factor:LDR | Factor:D30 RAW_FILE_NAME=NEG_159 SUBJECT_SAMPLE_FACTORS 137 34V Factor:Control | Factor:D1 RAW_FILE_NAME=NEG_165 SUBJECT_SAMPLE_FACTORS 139 61A Factor:HDR | Factor:D1 RAW_FILE_NAME=NEG_166 SUBJECT_SAMPLE_FACTORS 141 125V Factor:LDR | Factor:D20 RAW_FILE_NAME=NEG_167 SUBJECT_SAMPLE_FACTORS 143 91A Factor:Control | Factor:D3 RAW_FILE_NAME=NEG_168 SUBJECT_SAMPLE_FACTORS 144 32V Factor:Control | Factor:D1 RAW_FILE_NAME=NEG_169 SUBJECT_SAMPLE_FACTORS 145 79A Factor:HDR | Factor:D5 RAW_FILE_NAME=NEG_172 SUBJECT_SAMPLE_FACTORS 146 35V Factor:Control | Factor:D1 RAW_FILE_NAME=NEG_173 SUBJECT_SAMPLE_FACTORS 147 98A Factor:Control | Factor:D5 RAW_FILE_NAME=NEG_174 SUBJECT_SAMPLE_FACTORS 149 74A Factor:HDR | Factor:D3 RAW_FILE_NAME=NEG_175 #COLLECTION CO:COLLECTION_SUMMARY Serum was collected after irradiation CO:SAMPLE_TYPE Blood (serum) #TREATMENT TR:TREATMENT_SUMMARY The VADER was designed to deliver controlled dose rates in the range 0.1 – 1 TR:TREATMENT_SUMMARY Gy/day to a cohort of up to 15 mice. The VADER uses ~0.5 Ci of retired 137Cs TR:TREATMENT_SUMMARY brachytherapy seeds that are arranged in two platters placed above and below a TR:TREATMENT_SUMMARY “mouse hotel”. The platters can be placed ~0.5 – 60 cm above and below the TR:TREATMENT_SUMMARY mouse hotel allowing implementation of time-variable dose rates. Offline TR:TREATMENT_SUMMARY dosimetry of the VADER was performed annually using a NIST traceable 10x6-6 TR:TREATMENT_SUMMARY ionization chamber (Radcal Corp., Monrovia, CA). Dose uniformity across the TR:TREATMENT_SUMMARY surface was measured using EBT3 film (Ashland, Covington, KY, USA) and the TR:TREATMENT_SUMMARY variation was 15% across the hotel. A lead and high-density concrete brick TR:TREATMENT_SUMMARY shield ensured minimal radiation doses to occupationally exposed personnel TR:TREATMENT_SUMMARY (operators) inside (< 0.1 mGy/wk) and outside the room (< 0.02 mGy/wk). The TR:TREATMENT_SUMMARY mouse hotel consists of an acrylic box (35 x 35 x 12 cm) allowing housing of ≤ TR:TREATMENT_SUMMARY 15 mice with bedding material and food/water ad libitum. Temperature (20 – TR:TREATMENT_SUMMARY 25°C), humidity (40 – 60%), airflow and lighting were fully controlled to TR:TREATMENT_SUMMARY required animal care standards (temperature/humidity sensor, HWg HTemp, TruePath TR:TREATMENT_SUMMARY Technologies Victor, NY). Environmental controls and monitoring were integrated TR:TREATMENT_SUMMARY into the mouse hotel for easy replacement in case of radiation damage. Mice were TR:TREATMENT_SUMMARY monitored in real time using a 180° fisheye ELP USB camera (Amazon). #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Samples were prepared and analyzed as previously described.18, 19 Briefly, serum SP:SAMPLEPREP_SUMMARY (5 μl) was deproteinized (195 μl 66% cold acetonitrile [ACN]) with internal SP:SAMPLEPREP_SUMMARY standards (2 μM debrisoquine [M+H]+ = 176.1188; 30 μM 4-nitrobenzoic acid SP:SAMPLEPREP_SUMMARY [M-H]- = 166.0141), vortexed, incubated on ice (10 min), and centrifuged for 10 SP:SAMPLEPREP_SUMMARY min (max speed, 4 °C). SP:PROCESSING_STORAGE_CONDITIONS -80℃ #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY Mobile phases consisted of the following: solvent A (water/0.1% formic acid CH:CHROMATOGRAPHY_SUMMARY [FA]), solvent B (ACN/0.1% FA), solvent C (isopropanol [IPA]/ACN (90:10)/0.1% CH:CHROMATOGRAPHY_SUMMARY FA). The gradient for urine was (solvent A and B) 4.0 min 5% B, 4.0 min 20% B, CH:CHROMATOGRAPHY_SUMMARY 5.1 min 95% B, and 1.9 min 5% B at a flow rate of 0.5 ml/min. The gradient for CH:CHROMATOGRAPHY_SUMMARY serum was (solvent A, B, and C) 4.0 min 98:2 A:B, 4.0 min 40:60 A:B, 1.5 min CH:CHROMATOGRAPHY_SUMMARY 2:98 A:B, 2.0 min 2:98 A:C, 0.5 min 50:50 A:C, and 1.0 min 98:2 A:B at a flow CH:CHROMATOGRAPHY_SUMMARY rate of 0.5 ml/min. CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Waters Acquity CH:COLUMN_NAME Waters Acquity BEH C18 (50 x 2.1mm, 1.7 um) #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Waters Synapt G2 S QTOF MS:INSTRUMENT_TYPE QTOF MS:MS_TYPE ESI MS:ION_MODE NEGATIVE MS:MS_COMMENTS Negative and positive electrospray ionization (ESI) data-independent modes were MS:MS_COMMENTS used for data acquisition with leucine enkephalin ([M+H]+ = 556.2771, [M-H]- = MS:MS_COMMENTS 554.2615) as Lock-Spray®. Operating conditions for ESI were: capillary voltage MS:MS_COMMENTS 2.75 kV, cone voltage 30 V, desolvation temperature 500°C, desolvation gas flow MS:MS_COMMENTS 1000 L/Hr. MS:MS_RESULTS_FILE ST001806_AN002928_Results.txt UNITS:peak area Has m/z:Yes Has RT:Yes RT units:Minutes #END