Acid-Catalyzed Oligomerization of Glycerol Investigated by Electrospray Ionization Mass Spectrometry

A oligomerização do glicerol catalisada por ácido foi investigada por espectrometria de massas com ionização electrospray, em modo de íon positivo, i.e. ESI(+)-MS. Os dados de ESI(+)-MS revelaram que a 280 °C e em meio ácido (pH 2), glicerol sofre sucessivas condensações intermoleculares para produzir espécieis oligoméricas, i.e. [(glicerol) n – (n – 1) H 2 O] (n = 2-5). Esses resultados de ESI(+)-MS, em associação com as informações obtidas por análises de infravermelho e RMN de H e C, indicaram que esses oligômeros sofrem desidratações para formar uma mistura de produtos, composta principalmente por compostos cíclicos, assim como alquenos e derivados carbonílicos (aldeídos e cetonas).

In this work, we have used direct infusion electrospray ionization mass (and tandem mass) spectrometry, in conjunction with IR and NMR spectroscopy, to accomplish an overview of the acid-catalyzed oligomerization of glycerol.The results revealed that the reaction mechanism is complex with the formation of an unexpectedly large variety of products.

Experimental
Ultrapure glycerol (272 mmol), H 2 SO 4 (2.72 mmol), and distilled water (3 mL) were mixed in a 200 mL glass reactor connected to a condenser and a collector flask.The system was placed inside a ceramic furnace and heated with a slope of 30 °C min -1 up to 280 °C.Aliquots were taken at intervals of 30 min and diluted 1000 times with MeOH/H 2 O 1:1.The pH of the diluted samples were adjusted to 2 by dropping H 2 SO 4 0.1 mol L -1 .
ESI-MS analyses were conducted in an MS instrument (Agilent Trap XCT, Santa Clara, CA) equipped with an electrospray source operating in the positive ion mode.The diluted aliquots were directly infused into the ESI source at a flow rate of 5 mL min -1 by a micro syringe.ESI source conditions were as follows: heated capillary temperature 150 °C; sheath gas (N 2 ) flow rate 5 L min -1 ; spray voltage 4 kV; tube lens offset voltage 25 V, capillary voltage 25 V.The mass spectra were acquired in a 50-500 m/z range and were obtained as an average of 50 scans, each one requiring 0.2 s.For the MS/MS experiments, the precursor ions were isolated with an isolation width of 1 m/z unit and fragmented via collision-induced dissociation (CID) with the target gas (helium).The relative collision energy was adjusted to yield product ions in quantifiable abundance.
The aliquots were also analyzed by FTIR (Fourier Transform Infrared Spectrometry), as liquid films using a Perkin Elmer model GX FT-IR instrument (Norwalk, CT, USA), and 1 H and 13 C NMR (Nuclear Magnetic Resonance), in D 2 O solution and recorded on a Bruker CXP-200 spectrometer (Billerica, MA, USA).

Results and Discussion
Typical ESI(+)-MS of the reaction aliquots taken at different times are shown in Figure 1.The presence of an intense ion of m/z 93 (protonated glycerol = [glycerol + H] + ) is clearly noticeable indicating the subsistence of glycerol in the reaction medium even after 2 h reaction.Furthermore, other ions of m/z 75 ([glycerol -H 2 O + H] + ) and 57 ([glycerol -2 H 2 O + H] + ) are also detectable.These ions probably arises from in-source dehydrations of [glycerol + H] + as they are also distinguishable in the ESI(+)-MS (not shown) of an aqueous solution of pure glycerol at pH 2.
A remarkable presence of an ion of m/z 167 is also noticed in Figure 1.This corresponds to the protonated form of diglycerol, i.e. [(glycerol) 2 -H 2 O], formed under these reaction conditions via the condensation of two molecules of glycerol and loss of water.This condensation can occur via the primary or secondary hydroxyl groups at the glycerol molecule to yield linear (a,a-diglycerol) and branched (a,b-diglycerol; b,b-diglycerol) isomers, as displayed in Scheme 1.In the ESI(+)-MS (Figure 1), the presence of ions of m/z 149 and m/z 131 is also promptly recognized.These ions are ascribed to be the protonated forms of products formed by successive dehydrations of diglycerol, namely [diglycerol -H 2 O] and [diglycerol -2 H 2 O], respectively (Table 1).These findings thus indicate that under these experimental conditions (acidic medium and heating), diglycerol can easily lose one or two molecules of water to form a myriad of isomeric products.Scheme 3 shows, for instance, prototype products possibly formed as a result of the mono-dehydration of diglycerol, such as the cyclic species 1a-c (their formation have been reported by Barrault and coworkers [18][19][20][21][22][23][24] that submitted glycerol to similar reaction conditions than those employed herein) besides the acyclic carbonyl compounds 2a-b and the alkene 2c.The presence of the last products in the reaction medium was evidenced by the data from IR and NMR spectra (not shown), which were acquired from an aliquot taken after a reaction time of 2 h.Hence, an absorption band at 1725 cm -1 , typically related to the C=O stretching of aldehydes and ketones, was observed in the IR spectrum.The NMR records also revealed the presence of aldehydes (as evidenced by the signals at 9.6 ppm in the 1 H NMR as well as 177 and 178 ppm in the 13 C NMR) in addition to alkenes (owing to the existence of signals at 7.1 ppm in the 1 H NMR as well as 100 and 166 ppm in the 13 C NMR).
Note that all the products resulting from the monodehydration of diglycerol, including the ones shown in Scheme 3 (1a-c and 2a-c), possess the same chemical formula (C 6 H 12 O 4 ) and bear similar functional groups (especially hydroxyl substituents).Hence, as these protonated molecules are simultaneously selected inside the ion trap and dissociate in a quite similar way (yielding a number of fragments arising mainly from successive losses of water and other small molecules), the unambiguous characterization of a particular product based exclusively on such fragmentation profile is certainly unfeasible.
As expected, the mass-selection and fragmentation of such protonated oligomers, of m/z 241, 315, and 389, respectively, yielded product ions arising mainly from successive eliminations of water (Table 2).As previously stated, these products are likely comprised by a mixture Table 1.Primary products (diglycerol, triglycerol, tetraglycerol, and pentaglycerol) and their dehydration products formed upon acid-catalyzed oligomerization of glycerol at 280 o C. All these products were observed as their protonated forms in the ESI(+)-MS (Figure 1 of different isomers (linear and branched) that are formed depending on the nature of the OH group (primary or secondary) active in the condensation step.
Similarly to that verified for diglycerol, the heavier oligomers, i.e. triglycerol, tetraglycerol, and pentaglycerol, promptly generate a number of dehydration products (possibly comprised by a mixture of cyclic and unsaturated acyclic compounds: alkenes, aldehydes, and ketones) under these experimental conditions.The protonated forms of such compounds are detected in the ESI(+)-MS (Figure 1) and a general overview of their chemical compositions is presented in Table 1.
In Figure 3, the fraction of the ions of m/z 93, 167, 241, 315, and 389 (protonated glycerol, diglycerol, triglycerol, tetraglycerol, and pentaglycerol, respectively), given as a quotient ratio between the absolute intensity of one of such ions and the sum of the absolute intensities of the whole set of ions, are plotted against the reaction time.This is certainly a convenient approach to monitor the changes in the relative concentrations of such species during the reaction course.Hence, the results show that after 2 h reaction more than 90% of glycerol is consumed.Furthermore, during the first 30 min a relatively high concentration of diglycerol is formed.At longer reaction times, however, its concentration decreases whereas the amount of the heavier oligomers (triglycerol, tetraglycerol, and pentaglycerol) concomitantly increases.These results, in association with the observed gradual rise in the medium viscosity (from 1000 cp for pure glycerol to 5978 cp for the raw material obtained after 2 h reaction), strongly support the assumption that glycerol is continuously converted into the heavier and more viscous oligomeric compounds.
In Figure 4 the fraction of protonated diglycerol (of m/z 167) and its dehydration products (of m/z 149 and 131) are plotted against the reaction time.Curiously, the results show that diglycerol only becomes predominant after a reaction time of 30 minutes.Before this time, the fully dehydrated compound, [diglycerol -2 H 2 O], is found to be the major product.Similar trend was observed for triglycerol and its dehydration products (not shown).These findings probably indicate that at the beginning of the reaction process, diglycerol and triglycerol are quickly converted into their dehydration products.At longer reaction times, however, the increasingly amount of water in the reaction medium allows the reconstitution of the initial condensation adducts.

Conclusions
The results described herein demonstrated the usefulness of ESI-MS in revealing new and relevant aspects on the acid-catalyzed oligomerization of glycerol.Hence, under the drastic conditions employed herein (acidic medium and heating) a number of oligomers (diglycerol, triglycerol, tetraglycerol, and pentaglycerol) could be detected.In addition, ESI(+)-MS, in conjunction with the data provided by IR and NMR techniques, clearly revealed the presence of secondary products formed by the dehydration of such primary oligomers.These dehydration products are possibly comprised by a mixture of cyclic and unsaturated acyclic compounds (alkenes, aldehydes, and ketones).

Figure 1 .
Figure 1.ESI(+)-MS of the acid-catalyzed oligomerization of glycerol conducted in aqueous medium at 280 °C.The aliquots were collected after reaction times of: (a) 30 min; (b) 60 min; (c) 120 min.The ions marked with an asterisk (*) refer to dehydration products (see also Table1for a general overview of the products formed under these conditions).

Figure 2 .Scheme 2 .
Figure 2. ESI(+)-MS/MS experiment with the ion of m/z 167.For a complete interpretation of the fragmentation pathways of protonated a,a-diglycerol, see Scheme 2.

Scheme 4 .
Scheme 4. Oligomeric products formed by the self-condensation of glycerol in aqueous acidic medium at 280 o C. Note that only the linear isomers, formed via condensations of the primary hydroxyl groups, are displayed herein.

Figure 3 .
Figure 3.Fractions of the ions of m/z 167 (protonated diglycerol), 241 (protonated triglycerol), 315 (protonated tetraglycerol), and 389 (protonated pentaglycerol) as a function of reaction time.Each fraction was calculated as the quotient ratio between the absolute intensity of one of such ions and the sum of the absolute intensities of the whole set of ions.

Figure 4 .
Figure 4. (a) Fractions of the ions of m/z 167 ([diglycerol + H] + ), 149 ([diglycerol -H 2 O + H] + ), and 131 ([diglycerol -2 H 2 O + H] + ) as a function of reaction time.Each fraction was calculated as the quotient ratio between the absolute intensity of one of such ions and the sum of the absolute intensities of the whole set of ions.
Table 1 for a general overview of the products formed under these conditions).Vol. 20, No. 9, 2009 As expected, the mass-selection and fragmentation of the ion of m/z 167 (Figure 2) yielded mainly product ions arising from losses of one or two molecules of water (m/z 149 and 131, respectively) besides to other product ions, such as [glycerol + H] + (m/z 93), [glycerol -H 2 O + H] + (m/z 75), and [glycerol -2 H 2 O + H] + (m/z 57).To illustrate the formation of such fragments, the dissociation pathways for protonated a,a-diglycerol are shown in Scheme 2. ) Scheme 3. Examples of products possibly formed as a result of the mono-dehydration of diglycerol.