AMI. chsm. 1081, 5.9.565-566
Flwa 1. Ropaaonatlow three-way rnlcromaahg valve.
fittedwith geared micrometer heads. The trap, which contains a Teflon-coated magnetic stirring bar used to prevent concentration gradients, is necked down to 3/a in. glass tubing sealed in place hy means of a Swagelock nut which compa Teflon ferrule. The Swagelock nut attachea to a custom-made fiafter removing one port of a S w a g e l d SS2oo.I union cross,it was silver soldered to the closed end of a Swagelock SsGoO-C cap. A in. hole, drilled all the way through the cross using the port opposite the cap as a guide, made possible insertion of in. SS tubing to the desired depth in the trap. The valves are mounted on I/, in. Plexiglas a t a distance which permits proper meshing of identical gears which are glued on the two heads with quick-setting epoxy glue. Thus,
5e!l
one valve closes if the other is opened. Carrier gas entering a t lower left has two paths to follow before leaving at middle right: over and around the acid. The proportion of each detenninea the degree of sa!mation. Ten percent of the carrier gas flow via the formic acid trap gave the required GLC characteristics. S i m i i l y plumbed valvea,whoae outputs before pflow through a drying tower and water huhbler, reapeaively, made possible control of the relative humidity (RH) of an air stream. The same approach, hut on a larger d e , was used to control the humidity of a 40 L/min flow of air supplied to a bank of eight olfactometers (2). Low flows (less than 1 mL/min) of compressed carbon dioxide were easily set by using two geared valvea in parallel: the output of one valve was used in bioassays of mosquito response to human emanations; the output of the other valve was dumped. Smooth control of the flow rate was possible because of the invariant hack-pressure seen by the tank reducing valve. Finally, connection of the valvea with two different inputs which join downstream would appear useful in the generation of h ear gradienta as used in liquid chromatography or in the preparation of a twegan mixture. Such a twegas mixture is required in chemical ionization mass spectrometry. The proportion of reagent gas could be varied while keeping relatively constant the high pressure of carrier gas in the ion source.
LITERATURE CITED (1) Adcmn. R. 01: Bvga. R. D. Anal. CXm. 100% 95.647449. (2) Rka, Q. D.: Honk. R. n.: Smrm. J. . I .Msd. €nfmcf. 1078. 1 4 715-719.
RP~IVEDfor review July 28,1980. Accepted December 1, 1980. Mention of a commercial product doea not constitute a reeommendation or an endorsement of this product by the University of Florida or the U.S.Department of Agriculture. The research reported in this manuscript was conducted in part with contract funds transferred from the Medical Research and Development Command, Office of the Surgeon General, US.Army.
Analysis of Methanol for Reversed-Phase Gradient Elution Liquid Chromatography Net1 E. Splngnrn." Clare 1.GarvbGould. and Lorato L. Vuolo Amwlcan Hsallh Fwndefbn. NaW Dnna InsMute fa- Lhaw Fmenfbn, VahUa, New
The existence of organic impurities in highquality methanol can cause serious problems when employing the common technique of reversed-phase high-performance liquid chromatopraphy, using either isocratic or gradient elution techniques. The availability of many different grades of solvents from a large number of suppliers only makes it more difficult for the chromatographer to select the product most suited to his purposes. This paper presents a simple method for evaluation of organic solvents and demonstrates the variability between lots of methanol from different suppliers. A promdure of this type has been shown to be useful in evaluating water (I) and was suggested to be useful for organic solvents. ' P m n t address VA Wadworth Cancer Center, Box 691/11N. Wilshire and Sawtelle Boulevards. L a Angel-, CA 90073.
rork
10595
EXPERIMENTAL SM;"ITON
An Altex (Berkeley, CA) liquid chromatograph,consisting of two Model llOA pump was controlled by a Model 420 p m gramerlcontroller. The column was a L i C h o r b (E.Merck Products, Darmstadt, Germany) C18 column (4.6 X 250 mm),and the detector waa an Altex Model 153 fitted with an BrL, 1.0 em pathlength flow cell and monitored at 254 om. Water used waa Baker (Phillipshurg,NJ) HPLC grade. The methanol8 evaluated were Burdick and Jackson (Muskegon,MI) Distilled in Glass,Baker HPLC, MCB (Norwood. OH) OmnisOlv, E. Merck LiChmsolv, Mallickodt (SL Louis, MO) ChromAR and Fisher (Fair Lawn, NJ) HPLC. All solvents were degassed under vacuum with magnetic stirring or ultrasonic cavitation. For reduction of the chance of contamination of the solvent, previously unopened bottles were used directlyaolventa were not transferred to other mntainera. The solvent lines were flushed
566
ANALYTICAL CHEMISTRY, VOL. 53, NO. 3, MARCH 1981 Table I. Analysis of Impurities in Methanol re1 brand no. absorbancea contaminantsb 0.01 A
0.014 0.008 0.010 0.011 0.018 0.016 0.021 0.029
1 1C
I
I:
2 3 4 5 5c
6
1 1.1 1.5 1.6 3.9 3.8 7.6 > 27
a Absorbance at 254 nm of 100% methanol relative to water. Sum of integrations of supurious peaks relative to the lowest obtained. Different lots of the same product.
Table 11. Analysis of Eluted Contaminants from Methanol
Flgure 1. Chromatograms of identically run gradients from 0% to 100% methanol of brands 1 and 4.
before the column was switched in. After the base line stabilized at 1.0 mL/min of methanol, the program was started. The program consisted of holding 20 min successively at 100% methanol and then 100% water, followed by a 30-min linear gradient back to 100% methanol. This procedure was repeated two or three times for each bottle of methanol evaluated. Output chart records of repeats were identical at the limit of visual discrimination. The spurious peaks from brand 1and brand 6 methanols were collected into carefully washed glassware. The eluate. was reduced to dryness under a stream of nitrogen, taken up in a small volume of brand 1 methanol, and reinjected into the HPLC set for a 30-min gradient from water to 100% methanol.
RESULTS AND DISCUSSION Figure 1 shows the results of gradient elution from 100% water to 100% methanol on a C18 reversed-phase column. Brand 1 showed very little overshoot of the absorbance of methanol and no later spurious peaks. In contrast, brand 4 showed substantial overshoot and several undesirable peaks. This, along with the results from the other brands, is quantitated in Table I. The overshoot and spurious peaks (contaminants) were integrated and summed to determine the quality of the methanol. The range of contaminants present is quite broad. Brand 6 had more than 27 times the contaminants of brand 1. While the difference between lots of brand 1was small, the two lots of brand 5 were not similar in quality. The information given on the solvent bottles reported as actual lot analyses was found to differ from measured values considerably. Our measured values were in some cases 100% above that reported, while in other cases as much as 50% below the reported absorbance.
injection none brand 1 none brand 6 See Table I.
r el absorbancea contaminantsa 0.025 0.025 0.021 0.022
1
1.4 1.4 11.2
The eluate from brands 1 and 6 were collected, dried, resuspended in brand 1methanol, and reinjected into the HPLC with the same gradient to ensure that the spurious peaks were due to isolatable contaminants. Table I1 verifies that the contaminants in methanol can be isolated, eliminating several possible explanations for their appearance (Le., solvent degassing, other low molecular weight alcohols). While the overshoot seen in all the brands of methanol could be due to impurities in the high-quality water used, the major contaminants seen in some brands on methanol are unlikely to come from this source since the water was identical for all brands studied. We conclude that the different absorbance changes and spurious peaks are due to impurities in the methanol. It is recommended that the chromatographer evaluate each lot of methanol to be used by this simple procedure to determine the suitability for his specific purposes. The lobto-lot and brand-to-brand differences are larger than the manufacturers would lead one to believe.
LITERATURE CITED (1) Bristol, Douglas W. J. Chfomtogr. 1980, 188, 193-204
RECEIVED for review August 25,1980. Accepted November 24, 1980. This work was supported in part by NCI grant CA06357 and by CA24217 through the National Large Bowel Cancer Project.
