Worm lipid extraction, separation, and analysis were performed as previously reported (53 (link)). In general, about 200,000 L4 worms were collected, and the protein was quantified with a Pierce BCA Protein Assay Kit. About 15 mg protein was used for total lipid extraction. For lipid separation, 40 μl of total lipid was loaded in triplicate on TLC silica plates (Merck) to separate TAG, PE, PI, PS, and PC. The TLC plates were developed in the solvent system chloroform:ethanol:water:triethylamine (30:35:7:35) (54 (link)) and dried in fume hood. About 0.005% primuline was sprayed evenly on the plate. The spots corresponding to TAG, PE, PI, PS, and PC were visualized under UV light and scraped from TLC plates and transesterified (2.5% H2SO4 in methanol). Next, esterified C15:0 was added as an internal standard, and lipids were extracted for GC analysis to determine the relative levels of TAG and PL fractions as previously described (45 (link)). Fatty acids were determined with an Agilent 7890 series gas chromatographer equipped with a 30 m × 0.25 mm × 0.25 μm DBWAX column (Agilent), with nitrogen as the carrier gas at 1.4 ml/min, and a flame ionization detector (53 (link)).
Agilent 7890 series
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The Agilent 7890 series is a line of gas chromatography systems designed for analytical laboratories. The core function of these systems is to separate and analyze complex mixtures of volatile compounds. The 7890 series provides precise control of operating parameters, such as temperature and gas flow, to ensure reliable and consistent results.
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5 protocols using agilent 7890 series
1
Worm Lipid Extraction and Analysis
2
Fatty Acid Composition of Black Goat Meat
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The fatty acid composition of black goat meat was estimated using a procedure
reported byO’Fallon et al.
(2007) . For the separation of fatty acid methyl esters, 0.7 mL of 10 N
potassium hydroxide (KOH) and 6.3 mL of methanol was mixed with each sample (1
g). This mixture was then added to a water bath that was kept at a constant
temperature of 55°C. The samples were heated for 1.5 h, with vigorous
shaking every 30 min. Subsequently, 0.58 mL of 24 N H2SO4was added to the compound after being cooled for 1–2 min in ice-cool
water. The mixture was then heated again at the same constant temperature using
the same process. Centrifugation was carried out at 1,100×g for 5 min
(HANL Combi-514R, Hanil Scientific, Inchon, Korea) after adding 3 mL of hexane
and placing it in a vial using a Pasteur pipette. This assay was conducted under
the following conditions using a gas chromatography-flame ionization detector
(Agilent 7890 series, Agilent, Wilmington, DE, USA): The injector was in split
mode with a split ratio of 25:1 at a temperature of 250°C, and the
detector was flame ionization. High-purity air, H2, and He were used
as the carrier gas, and the flow rate was 40 mL/min for H2 and 400
mL/min for air. The column for analysis was HP-88 (60 m×250
μm×0.2 mm). A relative percentage of all evaluated fatty acids was
used to represent each fatty acid.
reported by
(2007)
potassium hydroxide (KOH) and 6.3 mL of methanol was mixed with each sample (1
g). This mixture was then added to a water bath that was kept at a constant
temperature of 55°C. The samples were heated for 1.5 h, with vigorous
shaking every 30 min. Subsequently, 0.58 mL of 24 N H2SO4was added to the compound after being cooled for 1–2 min in ice-cool
water. The mixture was then heated again at the same constant temperature using
the same process. Centrifugation was carried out at 1,100×g for 5 min
(HANL Combi-514R, Hanil Scientific, Inchon, Korea) after adding 3 mL of hexane
and placing it in a vial using a Pasteur pipette. This assay was conducted under
the following conditions using a gas chromatography-flame ionization detector
(Agilent 7890 series, Agilent, Wilmington, DE, USA): The injector was in split
mode with a split ratio of 25:1 at a temperature of 250°C, and the
detector was flame ionization. High-purity air, H2, and He were used
as the carrier gas, and the flow rate was 40 mL/min for H2 and 400
mL/min for air. The column for analysis was HP-88 (60 m×250
μm×0.2 mm). A relative percentage of all evaluated fatty acids was
used to represent each fatty acid.
3
GC-MS Analysis of Volatile Compounds
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After extraction, GC–MS analysis was carried out through a GC (Agilent 7890 Series, Agilent Technologies) coupled with a triple‐axis detector (Agilent 5975 Inert XL MSD, Agilent Technologies) and a DB‐5 column (30 m × 0.25 mm i.d. × 0.25 μm; Agilent Technologies). The SPME fiber head was inserted into the sample hole at 250°C with desorpting for 1 min, running time for 30 min. And fiber head was maintained 5 min to remove impurity in the sample hole, and this hole employed no crack (hole) mode. The flow rate of the carrier gas (99.9995% He) was 40 cm/s. The temperature program of the GC oven was kept at 50°C for 1 min, then was raised to 100°C at 5°C/min, and increased to 250°C at the rate 10°C/min, and it was kept for 9 min. Helium at 1.2 ml/min acted as the carrier gas, and splitless injection mode was adopted. The quadrupole mass filter and the ion source were at 106 and 200°C, respectively. The mass spectrometer was used at 70 eV in the electron ionization mode, and the total ion current in the scanning range of 33–350 m/z was monitored to record the chromatograms. The NIST library was used for preliminary identification and combined with the retention time, the actual components, and the retention index to further determine most of the components.
4
Fatty Acid Composition of Marinated Frozen Meat
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The fatty acids composition of 7 d marinated frozen/thawed meat (1 d of thawing)
was determined by using a slightly modified method described by O'fallon et al. (2007) (link). After the
separation of fatty acid methyl esters, the fatty acid analysis was performed
using the Gas Chromatograph-Flame Ionization Detector (FID; Agilent 7890 series,
Agilent, Santa Clara, CA, USA) under the following conditions. The injector was
split mode with a split ratio of 25:1, the temperature was 250°C, and the
detector was FID. High purity air, high purity H2, and helium was
used as the carrier gas. The flow rate was 40 mL/min for H2 and 400
mL/min for air. HP-88 column (60 m×250 μm×0.2 mm) was used
for the analysis. Fatty acids composition is expressed as a percent of meat.
was determined by using a slightly modified method described by O'fallon et al. (2007) (link). After the
separation of fatty acid methyl esters, the fatty acid analysis was performed
using the Gas Chromatograph-Flame Ionization Detector (FID; Agilent 7890 series,
Agilent, Santa Clara, CA, USA) under the following conditions. The injector was
split mode with a split ratio of 25:1, the temperature was 250°C, and the
detector was FID. High purity air, high purity H2, and helium was
used as the carrier gas. The flow rate was 40 mL/min for H2 and 400
mL/min for air. HP-88 column (60 m×250 μm×0.2 mm) was used
for the analysis. Fatty acids composition is expressed as a percent of meat.
5
Fatty Acid Profiling of Meat Samples
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Fatty acid composition of the meat sample was measured according to
O’Fallon et al. and Uyen et al. with some modification [22 (link),23 (link)]. The sample (1 g) was mixed with 0.7 mL of 10 N KOH and 6.3 mL
of methanol and placed in a constant-temperature water bath at 55°C. The
mixture was vigorously shaken every 30 min for 1 h and 30 min. After cooling in
ice water for 1–2 min, 0.58 mL of 24 N H2SO4 was
added to the samples and the mixture was shaken again every 30 min for 1 h and
30 min while being heated in the constant-temperature water bath at 55°C.
After cooling in ice water, 3 mL of hexane was added, and the mixture was
centrifuged at 3,000×g for 5 min using a Combi-514R centrifuge (Hanil) to
separate the fatty acid methyl esters. The upper layer was transferred to a vial
using a Pasteur pipette, injected into the gas chromatograph equipped with the
HP-88 column (60 m × 0.2 mm × 250 μm) and a flame
ionization detector (Agilent 7890 series, Agilent Technologies, Wilmington, DE,
USA). The injector was set to a split ratio of 25:1 with a temperature of
250°C, and the temperature of the detector was set to 250°C. A
high-purity mixture of air, H2, and He was used as a carrier gas with
a flow rate of 400 mL/min for air and 40 mL/min for H2.
O’Fallon et al. and Uyen et al. with some modification [22 (link),23 (link)]. The sample (1 g) was mixed with 0.7 mL of 10 N KOH and 6.3 mL
of methanol and placed in a constant-temperature water bath at 55°C. The
mixture was vigorously shaken every 30 min for 1 h and 30 min. After cooling in
ice water for 1–2 min, 0.58 mL of 24 N H2SO4 was
added to the samples and the mixture was shaken again every 30 min for 1 h and
30 min while being heated in the constant-temperature water bath at 55°C.
After cooling in ice water, 3 mL of hexane was added, and the mixture was
centrifuged at 3,000×g for 5 min using a Combi-514R centrifuge (Hanil) to
separate the fatty acid methyl esters. The upper layer was transferred to a vial
using a Pasteur pipette, injected into the gas chromatograph equipped with the
HP-88 column (60 m × 0.2 mm × 250 μm) and a flame
ionization detector (Agilent 7890 series, Agilent Technologies, Wilmington, DE,
USA). The injector was set to a split ratio of 25:1 with a temperature of
250°C, and the temperature of the detector was set to 250°C. A
high-purity mixture of air, H2, and He was used as a carrier gas with
a flow rate of 400 mL/min for air and 40 mL/min for H2.
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