Solid Sample Injector for Gas liquid Chromatography Robert R. Lowry, Department of Agricultural Chemistry, Oregon State University, Cotwallis, Ore.
0
UR PROJECT requires the
analysis of many lipid samples for their fatty acid content. I n carrying out the gas liquid chromatography of the methyl esters we were dissatisfied with the amount of tailing which occurred throughout the chromatogram and, in particular, the resultant poor resolution between the palmitate and palmitoleate and between the stearate and oleate peaks. Various trials involving the usual parameters available were tried with little success. I n addition, we were having a difficult time with the septums used in syringe injeciion as they frequently fragmented 2nd the smaller particles clogged the 1,ubing below the septum. This necessitated frequent, time-consuming cleaning of the injection port. Recently, two papers (1, 2 ) appeared describing devices for solid sample injection. The particular problems we were having would not be completely resolved by the. use of either device, because of the physical design of our instrument, a Beckmm Model GC-2. As a result, we designed and built the device described herein. The easiest approach from the front of the instrument was to remove the gas sampling valve and make a 3/ls~nchnotch in the oven lid 3l/8 inches from the inside right front corner. The carrier gas tubing from the regulator was replaced with a longer length of tubing that permitted it to be attached to the solid sample injector or the regular syringe injection port. The:,e are the only modifications required to permit use of our solid sample injector in the instrument. Since the changes to the instrument are minimal, reversion to the regular syringe and septum injection is quick and easy. If it is necessary to retain the gas sampling valve, our device can be located to the left of it by drilling a suitable hole In the front panel of the instrument. Figure 1 shows the basic components of the injector. The 3/lrinch hole in the inside brass block is located at the same level as the front wall of the oven. Experience has shown that this dimension varies from instrument to instrument, hencp no dimension can be given. ThP thermocouple well is in the same horizontal plane. The S/16-inch hole running the length of block is for a porcelain heater cartridge, G E No. XP10x15, used in coffee percolators. This permits rapid, controlied volatilization of the sample. Since these heaters also vary slightly in size, it is preferable to check their diameter bsfore drilling the
and attaching it using four 6-32 machine screws. This forms a n effective glass-to-metal seal that is gastight a t 25 p s i . and requires only occasional renewal of the O-ring. Figure 2 shows the assembled device and its location in the instrument. The heater leads extend to a variable transformer which permits control of the block temperature. The carrier gas enters into the device at the outside block. Since the spoon comes within '/4 inch of the column, sample introduction is considerably improved in this respect also. In normal syringe injection, a distance of approximately 8 inches exists between the point of injection and the column.
hole. The brass tubing used to connect the block and to make the spoons is available in hobby shops, as is the piano wire for the spoon assembly. The assembly is soldered together using a high melting eutectic such as Beckman No. 155. This eutectic was also used on the spoon assembly. Control of the stopcock which is located inside the front panel is accomplished by using a lever and rod assembly as shown. While we made the spoons detachable from the remainder of the spoon assembly, this is unnecessary unless the spoon and sample are to be weighed. The injector is assembled by placing an O-ring, the glass assembly, and the clamp on the end of the outside block
F N T WALL Of OVEN
TOP VIEW
'
I
DRILL HOLE FOR THLRYOCOUPLC
iI
OUT
IO€ BLOCK ORILL
I I
g0.D.BRASS TUBINO 9
a TAP FOR
*6-32
MACH SCREWS
I
-
SWAGELOK+ P o 0 R -3 If'TO RLOUCERSI
&
A -BLOCK
ASSEMBLY
0' RlNO JOINT REX t e 7 9 0
@- QLASS ASSEMBLY
-3-48tHD.
BRASS
STANDARD SLPTUM COR O C - 2
SCRLW wc I CULTURE T U
,24er C o m . t . o w
9Figure 1.
PIANO WIRE
SPOONASEYBLY
Components of solid sample injector VOL. 36, NO. 7 , JUNE 1964
1407
Table I. Reproducibility of Injections with NIH Standard "C" (C, t o C, fatty acid methyl esters, &9 Yocomposition, 8- to IO-Mg. injections)
Mean
std. dev.
a.Gt"d,
Figure 2. Photograph showing location of iniector in the instrument Sample injection is accomplished by placing a measured volume of the dissolved sample into the spoon and allowing the solvent to evaporate. A gentle stream of gas can be used to hasten evaporation. The spoon assembly is then placed so the spoon is in the outer half of the glass assembly. With the screw cap tightened, the stopcock is then opened and the spoon is rapidly pushed into the heated block. No interruption of the gas flow occurs a t any time. Escessive evaporation time
Figure 3. injection Sample used: rated
1408
Comparison of meihods of NIH standard
"C" ( C d m
ANANTICAL CHEMISTRY
satu-
+
Yo
C*
C,,
0.29 0.89 0.41 0.65 0.49 0.60 0.23 1.55
2.43 2.99 2.63 2.65 2.72 2.68 0.20 2.99
C. 5.31 6.18 6.03 5.85 5.78 5.83 0.33 5.96
(with or without a gas stream) can result in loss of the more easily vaporized sample components. Bleeding carrier gas into the out.er area of the glass assembly has not been necessary, nor has removal pf the spoon been necessary until the chromatogram is finished. The spoons do need frequent cleaning, however, even when analyzing samples such as purified fatty acid methyl esters. Figure 3 shows a comparison of standard syringe injection and use of the injector described using 8 to 10 fig. of methyl esters. The peaks of the short-chain fatty acid methyl esters are no longer on the slope created by the solvent peak and all the peaks are improved by the absence of tailing and improved symmetry. Figure 4 is a similar comparison of methods of injection showing the affect on the separation of a sample of fatty acid methyl esters obtained from rat liver mitochondria. Approximately 11 fig. were used for each chromatogram. The effects of tailing made the estimation of adjacent peaks difficult if not impossible when using syringe injection. Improvement of peak symmetry permits measurement of peak width a t half-height, and also. permits accurate peak height measurements. For comparative work, the loss of low boiling constituents is quite marked, as shown in Table I. All runs shown were made with a '/,inch X 6-foot column of 15y0ethyleneglycol succinate on Chromosorb P (35 to 80 mesh) at 190" C. in a Beckman GC-2 with a flame attachment using helium as a carrier . .gas.. . The . only variable was the :tion.
C, 11.99 12.23 12.20 12.56 12.33 12.26 0.21 11.96
CIS
Ca
20.86 20.25 20.54 20.23 20.07 20.39 0.28 19.41
25.*58 24 90 25.39 25.42 25.44 25 3.5 0 26 24.01
Figure 4. injection
C*O 33.54 32.55 32.81 32.65 33.16 3%.94 0.41
33.19
Comparison of methods of
Sample used: fony acid methyl esters of id h e r mitochondria
ACKNOWLEDGMENT
The author expresses appreciation to S. T. Likens for many helpful suggestions during the construction and use of the described device, and to the National .Institutes of .Health, Bethesda, ~~. .. . . .. 1
(1) McComas, D. B., Goldfien, A , ANAL.
CHEM.35,263 (1963). (2) Renshsw, A., Biran, L. A., J . Chrmnatog. 8,343 (1962).
Presented at the 17th Northwest Regional Meeting of the American Chemical Society, Bellingham, Wash., June 1963. Research undertaken in cooperation nith the Officeof the Surgeon General, UeparG m a t of the A m y , itnder Contract DA-
