1 Introduction

In last decades, natural science has developed remarkably, which has accompanied the change of content of education in high schools and universities drastically. Indeed, high school biology textbooks are completely different in comparison with those of 30 years ago. On the other hand, chemistry, despite its great development, has become a systematized discipline, and the content of high school chemistry textbooks has not changed significantly from that of 30 years ago. As the contents of textbooks are fixed and become common knowledge, it is natural that no one tries to revise them. However, the contents of textbooks are not always correct, and it is necessary to revise them when different findings are obtained.

The iodoform test is one of the important qualitative analyses to know the presence of acetyl group in the substrate, which is commonly taught in high school or introductory chemistry. When a substrate possessing an acetyl group is reacted with iodine under alkaline conditions, iodoform is observed as yellow precipitates [1], 2]. A compound possessing an α-hydroxyethyl group is also positive to this test because it is oxidized to an acetyl group by iodine in situ (Scheme 1). This reaction is also useful in organic synthesis because oxidation easily occurs accompanying fission of strong C–C bond [1, 3].

Scheme 1
scheme 1

Substrates positive to the iodoform test

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Descriptions about iodoform test in Japanese chemistry textbooks are summarized in Table 1. While the iodoform test using aldehydes and ketones is shown, no description is found with regard to the test using ethyl acetate as a substrate [4, 56, 7, 8, 9, 10]. Only single textbook [11] mentions that ethyl acetate is negative to this test because base-catalyzed hydrolysis proceeds to afford acetate anion that cannot generate dianion (Scheme 2). Similar descriptions are found in other books. When ethyl acetate is used as a substrate, yellow precipitates may occur, which is explained as being caused by ethanol produced by hydrolysis.

Table 1 Reaction conditions for the iodoform test in high school textbooks in Japan
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Scheme 2
scheme 2

Base-catalyzed hydrolysis of an ester functionality

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The reaction mechanism of the iodoform reaction is illustrated in Scheme 3. This reaction is initiated by deprotonation of the acetyl group to form the enolate ion, which immediately reacts with iodine leading to α-iodized ketone. Repeating this process twice forms triiodoacetyl group. The three electron-withdrawing iodo groups enhance the electrophilicity of the adjacent carbonyl carbon to facilitate the attack of a hydroxide. Since the iodo groups stabilize the α-anionic species, the triiodomethilide anion serves as a good leaving group to undergo the nucleophilic substitution to furnish the corresponding carboxylic acid accompanied by formation of iodoform as yellow precipitates. As far as this mechanism is concerned, it would not be surprising if similar reaction occurs with alkyl acetate (R = alkoxy). Indeed, yellow precipitates are observed when sec-butyl acetate and sec-amyl acetate are heated for a long time [12], 1638),however, it is not sure whether acetate moiety showed positive result or not because alcohols formed by hydrolysis are positive to the iodoform test. Therefore, we considered the iodoform reaction may proceed similarly even when acetic acid derivatives, alkyl acetate 1 and acetamides 2 (Table 2), are used as a substrate.

Scheme 3
scheme 3

Reaction mechanism for the iodoform reaction

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Table 2 Alkyl acetates 1 and acetamides 2 used for the iodoform test
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2 Results and discussion

At the beginning of the study, we verified whether the method described in the textbook was appropriate using acetone, positive substrate for the iodoform test. In a test tube, to a solution of acetone (1.0 mL, 18 mmol), aqueous solution of iodine–potassium iodide (1/2) (1.0 mL containing 0.49 mmol of iodine) was added dropwise using a syringe. Then, 2.2 M aqueous solution of sodium hydroxide was added using syringe until brown color of iodine disappeared. During this process, yellow precipitates were observed, which indicates the subsequent heating is not always necessary. After heating the resultant mixture at 60 °C for 1 min., yellow precipitates of iodoform were observed.

Five alkyl acetates 1a–e were employed as an ester substrate (Table 1). When hydrolysis of these esters occurs, esters 1b and 1d afford ethanol and 2-propanol (isopropyl alcohol), which show positive to the iodoform test. Hence, methyl ester 1a and propyl ester 1c were employed for comparison with 1b. Isopropyl ester 1d and tert-butyl ester 1e were expected to tolerate against the hydrolysis because of their bulkiness. When esters 1a–e were subjected to the iodoform test according to the above-mentioned method, yellow precipitates was not observed. However, separation to two layers was observed in cases of 1b–e, because alkyl acetates are immiscible with water (Fig. 1). These results indicated that the hydrolysis insufficiently occur under the employed conditions. When ethyl acetate 1b is hydrolyzed, acetic acid and ethanol are formed, both of which are miscible with water. Hence, the reason for the negativity of alkyl acetates 1b–e to the iodoform test was considered to be due to the heterogeneous system, which provides little opportunity for contact between the substrate and hydroxide ion.

Fig. 1
figure 1

Reaction mixture after the iodoform test using 1b, 1c and 1e

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THF was employed as an amphiphilic solvent to dissolve both a substrate and aqueous solution of sodium hydroxide homogeneously. Ethyl and propyl esters 1b and 1c were used as substrates. When sodium hydroxide solution was added, the brown color of iodine was disappeared; however, yellow precipitates were not observed in both cases. Two plausible reasons were considered for the negative results; (1) iodoform reaction did not proceed, (2) even though iodoform was formed, it could not be detected as yellow precipitates because of high solubility in THF. Hence, the reaction mixture was extracted with diethyl ether and evaporated to afford yellow solid as a residue. The solid was confirmed to be iodoform by comparison of 1H and 13C NMR spectra with those of authentic sample. It is noteworthy that the iodoform reaction proceeded for the alkyl acetates by conducting the reaction in homogeneous solution. When 1-propanol was used as a solvent, the iodoform reaction similarly proceeded, but yellow precipitates were not detected because this solvent also dissolve iodoform easily. Thus, THF and 1-propanol cannot be used as a solvent for the qualitative analysis.

Based on results obtained so far, three criteria are necessary for the solvent in iodoform test; the solvent (1) dissolves the substrate, (2) is miscible with aqueous solution of sodium hydroxide, (3) hardly dissolves iodoform. From these viewpoints, methanol was considered to be most appropriate because of the similar physical and chemical properties to those of water. When esters 1a–e were subjected to iodoform test in methanol, yellow precipitates were observed in cases of 1a–d (Fig. 2). Regarding tert-butyl ester 1e, separation to two layers was observed during addition of aqueous sodium hydroxide solution due to low polarity of 1e. The ester functionality undergoes ester exchange upon heating in alcohol, which accelerated by base catalyst (Scheme 4). If such reaction occurs in methanol under the employed conditions, ethyl and isopropyl esters 1b and 1d produce ethanol and 2-propanol, respectively, which are positive to the iodoform test. However, this possibility can be excluded because methyl and propyl esters 1a and 1c are also positive to this test.

Fig. 2
figure 2

Reaction mixtures after the iodoform test using 1a–d conducted in methanol

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Scheme 4
scheme 4

Base-catalyzed ester exchange

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This protocol was applied to other type of acid derivative, acid amide 2 (Table 2), which are less reactive for the hydrolysis than esters 1. When acetamide 2a and N-methylacetamide 2b were subjected to the iodoform test in methanol, no precipitates were observed because of the acidic N-hydrogen, which prevents the formation of the corresponding dianionic enolate ions (Scheme 5). To the contrary, N,N-dimethylacetamide 2c underwent the iodoform reaction to form yellow precipitates.

Scheme 5
scheme 5

Reaction of acetamides 2a and 2b with hydroxide ion

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3 Conclusion

We focused on description in the high school textbook that alkyl acetate 1 are negative to the iodoform test because of the hydrolysis of the ester functionality. When the test was conducted in aqueous media, alkyl acetates 1 were certainly negative, but is due to the immiscible property of the substrate. Indeed, using amphiphilic solvents such as THF and 1-propanol was effective to undergo the iodoform reaction. However, these solvents cannot be used for the qualitative test because yellow precipitates were not observed because of high solubility of iodoform into these solvents. This disadvantage was overcome by using methanol as a solvent. Not only alkyl acetates 1a–d but also N,N-dimethylacetamide 2c were positive to the iodoform test. These results will have a great impact to education in high school/inductor chemistry because they overturn what has been as common sense.