A related method involves treating phthalimide with sodium hypobromite in aqueous sodium hydroxide, followed by neutralization.[11] In the era when indigo dye was obtained from plants, it was degraded to give anthranilic acid.
Anthranilic acid is biosynthesized from chorismic acid by the action of anthranilate synthase. In organisms capable of tryptophan synthesis, anthranilate is a precursor to the amino acid tryptophan via the attachment of phosphoribosyl pyrophosphate to the amine group. After then, cyclization occurs to produce indole.
Industrially, anthranilic acid is an intermediate in the production of azo dyes and saccharin. It and its esters are used in preparing perfumes to mimic jasmine and orange, pharmaceuticals (loop diuretics, such as furosemide) and UV-absorber as well as corrosion inhibitors for metals and mold inhibitors in soy sauce.
N-hydroxyphthalimide (NHPI), which is best known as an organocatalyst for efficient C-H activation, has been found to be oxidized by quinoid compounds to its corresponding catalytically active nitroxide-radical. Here, we found that NHPI can be isomerized into isatoic anhydride by an unusually facile two-step method using tetrachloro-1,4-benzoquinone (TCBQ, p-chloranil), accompanied by a two-step hydrolytic dechlorination of highly toxic TCBQ into the much less toxic dihydroxylation product, 2,5-dichloro-3,6-dihydroxy-1,4-benzoquinone (chloranilic acid). Interestingly, through the complementary application of oxygen-18 isotope-labeling, HPLC combined with electrospray ionization quadrupole time-of-flight and high resolution Fourier transform ion cyclotron resonance mass spectrometric studies, we determined that water was the source and origin of oxygen for isatoic anhydride. Based on these data, we proposed that nucleophilic attack with a subsequent water-assisted Lossen rearrangement coupled with rapid intramolecular addition and cyclization in two consecutive steps was responsible for this unusual structural isomerization of NHPI and concurrent hydroxylation/detoxication of TCBQ. This is the first report of an exceptionally facile double-isomerization of NHPI via an unprecedented water-assisted double-Lossen rearrangement under normal physiological conditions. Our findings may have broad implications for future research on hydroxamic acids and polyhalogenated quinoid carcinogens, two important classes of compounds of major chemical and biological interest.
N-hydroxyphthalimide (NHPI) is a very unique hydroxamic acid, with two carbonyl groups linked to the nitrogen atom. NHPI is known to be used with certain co-catalysts to generate phthalimide N-oxyl radical (PINO), the key active organo-catalytic species for efficient C-H activation and subsequent oxygenation of hydrocarbons with dioxygen12,13,14,15. Therefore, we expected that a similar pathway may apply to the reaction between NHPI and TCBQ to produce the radical intermediate PINO in our system. According to our previous study16, we proposed an alternative pathway in which NHPI may attack TCBQ via nucleophilic substitution to initially form a transient intermediate NHPI-NO-TrCBQ (here, we use NHPI-NOH to refer to NHPI), followed by homolysis of the N-O bond, forming N- and O-centered radicals. However, to our surprise, neither the redox nor the nucleophilic substitution/homolysis pathway was observed during the reaction between NHPI and TCBQ. Through complementary applications of oxygen-18 isotope-labeling, high-performance liquid chromatography combined with electrospray ionization quadrupole time-of-flight mass spectrometry (HPLC-ESI-Q-TOF-MS) and high resolution Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR-MS) studies, we found that TCBQ induced an unusually facile two-step isomerization of NHPI to isatoic anhydride (IA) via a water-assisted double Lossen-type rearrangement coupled with rapid intramolecular nucleophilic addition under normal physiological conditions.
The loss of a proton from the nitrogen atom to form the anionic intermediate under alkaline conditions is considered to be essential for the classic Lossen rearrangement. Indeed, we recently found that benzohydroxamic acid can be activated by halogenated quinones (XBQs) to produce phenyl isocyanate, which requires the formation of an anionic N intermediate17,23. However, in the present study, the anionic N intermediate cannot be formed by losing a proton because the N atom is linked to two carbonyl groups. It has been reported that O-activated N-hydroxyphthalimide could also undergo Lossen rearrangement24,25,26, but various bases were required to trigger the ring-opening of the adduct that typically occurs in organic solutions. Evidence from the analysis described above and the results from oxygen-18 isotope-labeling for direct water involvement suggest that the reaction between NHPI and TCBQ may proceed through a previously unknown Lossen-type rearrangement pathway.
While studying the time course of IA generation, we noticed that IA was produced quickly and then slowly degraded (Fig. 5C). Because it has been shown that IA hydrolyzes into anthranilic acid in dilute alkaline solutions27, we speculated that a similar hydrolysis may occur under our conditions. HPLC-ESI-MS analysis using authentic anthranilic acid as the standard confirmed that this was indeed the case (SI Figure S4).
As shown above, TCBQ was first hydrolyzed to TrCBQ-OH and then to DDBQ. The yield of DDBQ was quite different when using either TCBQ (65%) or TrCBQ-OH (nearly 100%) (Fig. 2A,B) as the starting chemical, indicating that a side reaction may occur in the NHPI/TCBQ system. After carefully screening all possible species involved in this reaction, anthranilic acid was considered to be the most likely to react with TrCBQ-OH to produce the mysterious Product II via nucleophilic substitution28,29. Fortunately, we found that this is true. Using MS, we determined that Product II was 2,5-dichloro-3-(N-2-carboxyl phenyl)-6-hydroxy-1,4-benzoquinone (SI Figure S5).
Based on our previous work17,23, we expected that o-carboxy benzohydroxamic acid could be readily activated by TCBQ to generate o-carboxy phenyl isocyanate via Lossen rearrangement. If the mechanism proposed above for the formation of IA via the o-carboxy phenyl isocyanate intermediate were correct, then IA should also be produced from o-carboxy benzohydroxamic acid activated by TCBQ. We found that this is true (Fig. 7C,D). These results strongly support that o-carboxy phenyl isocyanate is the intermediate in the formation of IA from the reaction between NHPI and TCBQ.
IA, known for over a century, is an extremely versatile compound that easily reacts with both electrophiles and nucleophiles, lending itself to a wide range of applications in the manufacturing of agricultural chemicals, dyes, fragrances, pharmaceuticals and miscellaneous industrial chemicals28. Recently, IA derivatives, mainly N-methyl-IA and 1-methyl-7-nitro-IA, were also used to alter the ribose moiety of tRNA and mRNA for further structural and functional studies35,36. Three types of reactions have been commonly used to prepare IA: (1) cyclization of anthranilic acid with carbonic acid derivatives, (2) oxidation of isatin in glacial acetic acid and (3) rearrangement of phthalic acid derivatives (SI Scheme S1)37.
Some of these methods have been successfully applied in industrial production. However, they usually work under alkaline or acidic conditions and/or through heating. Furthermore, hypertoxic chemicals such as phosgene, chromium trioxide and chloroformate were also involved in these methods. Here, we developed a new method for the synthesis of IA from NHPI. Compared to traditional methods, this reaction could occur in water at room temperature and under neutral or even weakly acidic pH. The activating reagent TCBQ (also called p-chloranil) is readily available commercially, and its main final product is the non-toxic dihydroxylation product DDBQ. These features make this method more environmentally friendly.
In our previous studies, we found that TCBQ and H2O2 can produce highly reactive hydroxyl radicals via a metal-independent mechanism5,6,40,41,42,43, which can cause oxidative damage to DNA and other macromolecules44,45,46,47. Based on our finding that NHPI can effectively detoxify TCBQ, we expect that NHPI should also effectively protect plasmid pBR322 DNA from single-strand and double-strand breakage induced by TCBQ/H2O2. We found that this is indeed the case (SI Figure S6). Our present and previous studies demonstrated that11,17,48,49 NHPI and other hydroxamic acids may also be especially suited for detoxifying halogenated quinones. Further research is needed to investigate whether NHPI and other hydroxamic acids could be used safely and effectively as prophylactics for the prevention or treatment of human diseases, such as liver and bladder cancer associated with carcinogenic halogenated quinoid compounds.
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