BULLARD and COLLINS: IMPROVED METHOD TO ANALYZE TRIMETHYLAMINE 



a contribution by DMA of 1.69 mg N/100 g flesh to 

 the TMA value at 22° C. In frozen flesh of gadoid 

 fish, the DMA content might be high relative to 

 TMA and would result in a substantial error in the 

 TMA value unless determined at low tempera- 

 tures with KOH where DMA would contribute 0.1 

 mg N or less. 



We next studied the effects of the three bases 

 and the presence or absence of FA on the total 

 extractability of DMA. Similar extractions were 

 performed by Castell et al. (1974) but these au- 

 thors only considered the carbonate system with 

 FA present. The same DMA solutions (15.9 /xg 

 DMA-N/ml) with or without FA were extracted as 

 before but at room temperature for 15 min, i.e., 4 

 ml DMA solution, 10 ml toluene, 1 ml FA (or not), 

 and 3 ml base. After removing about 7 ml toluene 

 for drying and reacting with picric acid, the 

 remainder of the toluene layer was carefully 

 aspirated off, 10 ml toluene added, and reextract- 

 ed. This process was repeated for a total of six 

 extractions. In the absence of FA, DMA was 

 released rapidly from the salt by 45% KOH, about 

 half as fast by 25% KOH, and slowly by 50% 

 K2CO3 (Table 3). If FA were present however, 

 many extractions would be required to extract all 

 of the DMA which, in agreement with Sprung 

 (1940), showed that FA reacted with DMA to give 

 TMMD. The data further showed that TMMD was 

 relatively soluble in toluene but the extractabili- 

 ties were different because each base had a differ- 

 ent rate of reaction with TMMD, i.e., a rapid 

 release of TMMD with K2CO3 and slow with 25% 

 KOH. The possibility of each base having a 

 different salting-out effect was eliminated when 

 equal absorbancies were obtained if the same 

 extractions were made with the addition of 0.5 g 

 KCl (data not given). 



The data also showed that in the carbonate 

 method TMMD was released to the toluene phase 



more rapidly than DMA and explains the known 

 interference of DMA in the presence of FA by the 

 Dyer (1945) method. Formaldehyde might best be 

 left out in the 50% K2CO3 method. The lower 

 picrate color absorbancies in the KOH systems 

 with FA present might also be explained by the 

 law of mass action (equilibrium) as was done in 

 the section on NH3. In the equilibrium (FA + DMA 

 ^TMMD) the concentrations of FA, DMA, and 

 TMMD in the aqueous phase are dependent on the 

 type and concentration of base. The products (FA 

 and DMA) of the hydrolysis of TMMD would be 

 formed at a rate dependent upon these same 

 variables and DMA would be rapidly removed 

 from the aqueous phase in the KOH systems 

 because of the rapid extraction of DMA by toluene. 

 Since the absorbancies in the KOH systems were 

 relatively low in the presence of FA, the concen- 

 tration of DMA from the hydrolysis of TMMD 

 must have been low. In the carbonate system 

 however, DMA from TMMD was slowly released 

 from the aqueous phase into the toluene layer. 

 Apparently a low concentration of DMA existed in 

 the equilibrium formed in the carbonate sy.stem 

 and favored the extraction of TMMD by toluene. It 

 is likely that both TMMD and DMA were extract- 

 ed by toluene at rates that depend on the base and 

 temperature used. 



To further study the extraction of TMMD, the 

 same multiple extractions described for DMA 

 were made using purified TMMD in 5% TCA but 

 at a slightly lower concentration 115.0 /xg TMMD- 

 N/ml). The absorbancies of the TMMD-picrates 

 (Table 4) were nearly the same as the absorban- 

 cies of the DMA-picrates (Table 3). The similarity 

 of data between DMA and TMMD inferred again 

 that FA and DMA react to give TMMD. The 

 addition of FA forced the reaction toward TMMD 

 where the type and concentration of base con- 



TaBLE 3. — Dimethylamine hydrochloride: the absorbancies of 

 picrates in multiple extractions in the trimethylamine test as 

 affected by the three bases used. Samples were extracted for 15 

 min at room temperature using 4 ml 15.9 ng DMA-N/ml, with 

 formaldehyde ( + ) and without formaldehyde (0). 



Table 4.— N N N'N'-tetramethylmethanediamine: the ab- 

 sorbancies of picrates in multiple extractions in the trimethyl- 

 amine test as affected by the three bases used. Samples were 

 extracted for 15 min at room temperature using 4 ml 15.0 /ig 

 TMMD-N/ml, with formaldehyde 1+) and without formalde- 

 hyde (0). 



469 



