The relationship between the molecular sizes of acids and alcohols and their elution volumes in gel permeation chromatography

SHORT COMMUNICATIONS

5x9

The relationship between the molecular sizes of acids and alcohols and their elution volumes in gel permeation chromatography Separation by gel permeation chromatography is based on the ability of porous cross-linked polymer to distmgursh between molecules of different sizes, hence the relationship of elution volume with molecular weight or molecular size IS important However, there IS confusion about which is the best method. While some authcrs plot the clutlon volume directly against the molecular weighti, others either relate the cube root of the distribution cocfficicnt to the square root of the molecular weights or the distribution coefficient to the reciprocal of the logarithm of the molecular wcghta All claim to obtain a satisfactory linear relationship Yet, the most frequently used method IS to plot the clutron volume against the logarithm of the molecular weight; this method is widely employed by polymer chemists for the dctermmatron of molecular weight distribution For low-molecular-weight substances, a well-cstabhshed relationship between clution volume and molecular weight cnahles one not only to make some judgcment on the unknown substance but also roughly to predict the separability HENDRICKSOS AND MOORER studied various low-molccularweight compounds and mtroduccd the idea of cffectlve chain length in terms of carbon number. It was felt a more concentrated study should bc made in this dircctron. Straight-chain carboxyhc acids and alcohols were therefore studied

A fifiaratrrs A smglc gel pcrmcatlon column, I 2 cm x 1x5 cm, was used, the column substrate was I3io-bead S-X8 and the eluant tctrahydrofuran. A Chromo Syringe Pump (Waters Associates Inc ) was used at a flow rate of 0.730 ml/min A differential refractometer (Waters Associates Inc.) was employed as the detector

Chentacals All the chemicals were obtained from Eastman Kodak Inc

Results aftd dzscmsiorts Since the major mterest of this study was to establish an acceptable relationship between molecular +nze and elution volume, only straight-chain saturated compounds were employed to avoid possible complications, wluch might arise from structural effects of side chains The elution volumes of 7 hydrocarbons, ro acids and 12 alcohols were measured; duphcate measurements showed excellent reproducibility The data were first plotted against the logarithm of the molecular weight as shown in Fig I A fairly smooth linearity was obtained for each group. It was observed that the elution volumes of acids and alcohols always were slightly smaller than those of the analogous hydrocarbons, z.e. acids and alcohols eluted out as a larger molecule Since hydrocarbons are rather stable compounds, it is obvious that some corrections should be made for the elution volumes of aci& and alcohols. A similar fmdrng was also reported by HENDRICKSON AND MOOR@ and was attributed to hydrogen bondmg between the hydroxyl group of the solute and the elution solvent tetrahydrofuranG It is also Interesting to note that actds and the an,aIogous alcohols had almost identical elution volumes, which indicates that acids do not form duners m tctrahyANRL Chitrr Ada. 39 (lgG7)

5x9-521

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COMMUKICATIONS

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39 (1967)

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(rog so), or I 25 A, was assIgned as tic basic unit. The lcngthr of other functional groups wcrc calculated and converted mto effective carbon cham unit4 Thus method was found very convcnlent to cvpl c’ss the 51ze of low-molecular-wclght compounds and is used below Table I lists the clutlon volume, calculated carbon cham number and observed caibon chain number of each solute The carbon chain numbers of hydrocarbons wcrc frrst plotted against their elutlon volumes The cstabllshcd linear relatlonslup was then used a5 the standard curve to obtain the observed carbon chain number for ac& and alcohols The effecttvc chain length for oxygen IS o 67 carbon unit4 The last figure 15 mcanmgless m the logarlthnuc scale, therefore o 7 was allowed for oxygen m calculating the carbon chain number for acids and alcohols ‘I’hc cllffcrcnces between the calculated and observed values arc listed as D1 m Table I It was found that IA ~a.5 conslstcntly around 3 o carbon units ‘J‘hc average of 1A for 20 acids and alcohols cxcludmg methanol and ethanol was also 3 o carbon units. ‘I’ll15 number allows for the corrcctlon for the tetrahydrofuran mcJkCUk attached to acids and alcohols by hydrogcbn bondmg The S~LC:of tetlahydrofuran 1%about 3 2 11, w111ch corresponds to 2 0 carbon units, but, considering the weak hydrogen bondmg, a correctton of 3 o carbon units should bc adcquatc The average devldtlon hetwccn the calculated value and the corrected observed value, which 1s listed as D2 m Table I, cd>(JX~‘hc 15 only 0 35 carbon units This means that an unknown straiglit-chain aclcl or alcohol can be prcdlcted with a fair accurac> The elutlon volumes were again plotted VCYWS the logarithm of cham length m the correctecl value (Fig 2) The curve obtained was 5ufflclcntly linear to pcrmlt derivation of the following equation Elution volurnc: (ml) = 121 9-41 YkJg #c,

Amerlcalr Cyttmcnttd Co ) Stamford, Cow ~3904 (U S.A ) I Xl

AND A CUHDLL. Corupf Rrud , 25G (1963) 5224 Appl C/Iei~J , 6 (rgfij) 233 3 1’ &I ~hNIWI.lPPO AUD J (; SUHAK, / C/WOt?ZafO~, 13 (X961) 1.18. 4 J C HI~NDHICKSONANDJ C.MOORE,J PolyntcvSrr ,4 (I~GG) 107. lDr~~~:wr~~ AND h L MCCLXLLAN, Tire Ffydrogor Uomz, tV 5 c; C 2

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Secondary standards. Part 1. PI ickel ammine

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39 (1967)

5X9-521

complexes

A major difficulty m assessing new methods and in applying methods which require cahbratlon curves, 15 the provlslon of rcllablc standard materials that can bc used without further standardlsatlon. Comparatively few reagents arc pure enough or close enough to the theoretical composltlon to be used as primary or even secondary standards It would be desirable to have a list of reagents that could reasonably be expected to bc of at least secondary standard quality, wluch may be defined for Aural

Chant

Acfa,

39

(1967)

521-524