Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA162285

Local Sample ID:	QC.2_4
Subject ID:	SU001801
Subject Type:	Mammal
Subject Species:	Sus scrofa
Taxonomy ID:	9823
Age Or Age Range:	0-125 days
Weight Or Weight Range:	1-80 kg
Gender:	Male
Animal Animal Supplier:	Swine Innovation Centre Sterksel, the Netherlands
Animal Housing:	Metabolism cages
Animal Light Cycle:	12h light / 12h darknes
Animal Feed:	Experimental diets (see publication)
Animal Water:	Ad libitum
Animal Inclusion Criteria:	Birth weight

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Subject:

Subject ID:	SU001801
Subject Type:	Mammal
Subject Species:	Sus scrofa
Taxonomy ID:	9823
Age Or Age Range:	0-125 days
Weight Or Weight Range:	1-80 kg
Gender:	Male
Animal Animal Supplier:	Swine Innovation Centre Sterksel, the Netherlands
Animal Housing:	Metabolism cages
Animal Light Cycle:	12h light / 12h darknes
Animal Feed:	Experimental diets (see publication)
Animal Water:	Ad libitum
Animal Inclusion Criteria:	Birth weight

Factors:

Local Sample ID	MB Sample ID	Factor Level ID	Level Value	Factor Name
QC.2_4	SA162285	FL019174	QC	Diet
QC.2_4	SA162285	FL019174	QC	Wght_birth_cat

Collection:

Collection ID:	CO001794
Collection Summary:	In both balance periods, at d4 between 0900 and 1000 h three 1.5 ml urine samples per pig were taken from the buckets with urine collected from 0800 h that morning. Before taking the urine samples, the urine was mixed. The samples were stored at -80°C pending analyses.
Sample Type:	Urine
Storage Conditions:	-80℃

Treatment:

Treatment ID:	TR001814
Treatment Summary:	At an age of 14 weeks, 10 LBW-LEBV (BiW: 1.07 + 0.09 kg; EBV: -2.52 + 3.97 g/d, compared to an average crossbred pig with a protein deposition of 165 g/d), 10 LBW-HEBV (BiW: 1.02 + 0.13 kg; EBV: 10.47 + 4.26 g/d), 10 HBW-LEBV (BiW: 1.80 + 0.13 kg; EBV: - 2.15 + 2.28 g/d), and 10 HBW-HEBV (BiW: 1.80 + 0.15 kg; EBV: 11.18 + 3.68 g/d), male growing pigs (Synthetic boar x (Dutch Landrace x Large White)) were allotted to the experiment. The pigs were individually housed in metabolism cages (1.80 x 0.80 m) at a room temperature of 22oC. They were subjected to N- balance measurements in two sequential periods of 5 d using a restricted feeding regime. After a 6-d adaptation period to the metabolism cages, pigs were adapted for 5 days to the experimental diets before the start of the first 5-d balance period. Pigs were assigned to a protein adequate (A) or protein restricted (R, 70% of A) regime in a change-over design. LBW: Low birth weight; HBW: High birth weight; LEBV: Low estimated breeding value for protein deposition; HEBV: High estimated breeding value for protein deposition.

Treatment ID:

TR001814

Treatment Summary:

At an age of 14 weeks, 10 LBW-LEBV (BiW: 1.07 + 0.09 kg; EBV: -2.52 + 3.97 g/d, compared to an average crossbred pig with a protein deposition of 165 g/d), 10 LBW-HEBV (BiW: 1.02 + 0.13 kg; EBV: 10.47 + 4.26 g/d), 10 HBW-LEBV (BiW: 1.80 + 0.13 kg; EBV: - 2.15 + 2.28 g/d), and 10 HBW-HEBV (BiW: 1.80 + 0.15 kg; EBV: 11.18 + 3.68 g/d), male growing pigs (Synthetic boar x (Dutch Landrace x Large White)) were allotted to the experiment. The pigs were individually housed in metabolism cages (1.80 x 0.80 m) at a room temperature of 22oC. They were subjected to N- balance measurements in two sequential periods of 5 d using a restricted feeding regime. After a 6-d adaptation period to the metabolism cages, pigs were adapted for 5 days to the experimental diets before the start of the first 5-d balance period. Pigs were assigned to a protein adequate (A) or protein restricted (R, 70% of A) regime in a change-over design. LBW: Low birth weight; HBW: High birth weight; LEBV: Low estimated breeding value for protein deposition; HEBV: High estimated breeding value for protein deposition.

Sample Preparation:

Sampleprep ID:	SP001807
Sampleprep Summary:	In a 96-well plate, the urine samples (112.5 µL) were diluted with H2O (112.5 µL) and 25 µL acetonitrile (100%, ACN) containing an internal standard mix of glycocholic acid (glycine-1-13C) and p-chlorophenylalanine to a final concentration of 0.01 mg/mL was added. The samples were incubated for 20 min at 4°C and subsequently centrifuged (3700 rpm, 25 min, 4°C). The supernatant, 100 µL per well, was transferred to a 200 µL 96-well plate and a protective film was welded on the plate using a heat sealer, and the plate was centrifuged at 3700 rpm, 4°C for 25 min prior to the LC-MS analysis. Samples were kept in the autosampler at 10°C, and the injection volume was 3 µl.

Sampleprep ID:

SP001807

Sampleprep Summary:

In a 96-well plate, the urine samples (112.5 µL) were diluted with H2O (112.5 µL) and 25 µL acetonitrile (100%, ACN) containing an internal standard mix of glycocholic acid (glycine-1-13C) and p-chlorophenylalanine to a final concentration of 0.01 mg/mL was added. The samples were incubated for 20 min at 4°C and subsequently centrifuged (3700 rpm, 25 min, 4°C). The supernatant, 100 µL per well, was transferred to a 200 µL 96-well plate and a protective film was welded on the plate using a heat sealer, and the plate was centrifuged at 3700 rpm, 4°C for 25 min prior to the LC-MS analysis. Samples were kept in the autosampler at 10°C, and the injection volume was 3 µl.

Combined analysis:

Analysis ID	AN002808	AN002809
Analysis type	MS	MS
Chromatography type	Reversed phase	Reversed phase
Chromatography system	Thermo Dionex Ultimate 3000 RS	Thermo Dionex Ultimate 3000 RS
Column	Waters Acquity BEH HSS T3 (100 x 2.1mm,1.8um)	Waters Acquity BEH HSS T3 (100 x 2.1mm,1.8um)
MS Type	ESI	ESI
MS instrument type	QTOF	QTOF
MS instrument name	Bruker Impact HD	Bruker Impact HD
Ion Mode	NEGATIVE	POSITIVE
Units	peak area	peak area

Chromatography:

Chromatography ID:	CH002075
Chromatography Summary:	Chromatographic separation was performed on a Dionex UltiMate 3000RSL Binary UHPLC System (Thermo Scientific Dionex, Sunnyvale, CA) equipped with a HSS T3 C18 UHPLC column, 1.8 µm, 100x2.1 mm (Waters Corporation, Milford, MA). The column was maintained at 30°C. Samples were kept in the autosampler at 10°C, and the injection volume was 3 µl. The mobile phases were 0.1% formic acid in Milli-Q water (A) and 0.1% formic acid in acetonitrile (B). The gradient program for the urine samples was as follows: 0-8 min, linear gradient from 5-70% B; 8-8.5 min, linear gradient from 70-100% B; 8.5-9.5 min, 100% B and return to initial conditions in 0.2 min. The column was re-equilibrated at 5% B for 2 min at the beginning of each run and the flow rate was set to 400 µL/min.
Instrument Name:	Thermo Dionex Ultimate 3000 RS
Column Name:	Waters Acquity BEH HSS T3 (100 x 2.1mm,1.8um)
Column Temperature:	30
Flow Rate:	0.4 ml/min
Solvent A:	100% water; 0.1% formic acid
Solvent B:	100% acetonitrile; 0.1% formic acid
Chromatography Type:	Reversed phase

MS:

MS ID:	MS002603
Analysis ID:	AN002808
Instrument Name:	Bruker Impact HD
Instrument Type:	QTOF
MS Type:	ESI
MS Comments:	The eluent was introduced into the mass spectrometer by electrospray ionization, with capillary set in the positive and negative mode to 4500 and 3600 V, respectively. End-plate offset voltage was set to 500 V. Nitrogen was used as both nebulizer and drying gas with a gas pressure of 1.8 bar. Drying gas temperature and flow were 200 °C and 8.0 L/min, respectively. Spectra were acquired over the scan range of 50−1000 m/z. A solution of lithium formate clusters (5 mM) (water/isopropanol/formic acid in a 50:50:0.2 v/v/v ratio) was injected prior to each chromatographic run as an external calibrant.
Ion Mode:	NEGATIVE

MS ID:	MS002604
Analysis ID:	AN002809
Instrument Name:	Bruker Impact HD
Instrument Type:	QTOF
MS Type:	ESI
MS Comments:	The eluent was introduced into the mass spectrometer by electrospray ionization, with capillary set in the positive and negative mode to 4500 and 3600 V, respectively. End-plate offset voltage was set to 500 V. Nitrogen was used as both nebulizer and drying gas with a gas pressure of 1.8 bar. Drying gas temperature and flow were 200 °C and 8.0 L/min, respectively. Spectra were acquired over the scan range of 50−1000 m/z. A solution of lithium formate clusters (5 mM) (water/isopropanol/formic acid in a 50:50:0.2 v/v/v ratio) was injected prior to each chromatographic run as an external calibrant.
Ion Mode:	POSITIVE