MBTH method

The MBTH method (3-methyl-2-benzothiazolinone-hydrazone) is a nonselective colorimetric method for the determination of aliphatic aldehydes of low molecular weight (VDI 3862, Part 1, 1990). MBTH reacts on aldehydes to give an azide. In parallel, a reactive cation is formed by oxidation of MBTH with Fe(III). In a further step a blue ionic dye is formed (see Eq. 3). The absorbance is monitored at 628 nm. 

This reaction is less sensitive than that of the pararosaniline method. Other aldehydes undergo an analogous reaction, but the yield is generally lower. The MBTH method quantifies total aldehydes in ambient air in terms of their formaldehyde equivalents. Strong reducing agents can interfere with the determination of aldehydes. 

VDI 3862, Part 1 (1990) Gaseous emission measurement-measurement of aliphatic aldehydes (C, to Cj) MBTH method. Beuth, Berlin. 

Number of controls Carbazole method Orcinol method (years/months) MBTH method ... 

Carbazole method Orcinol method MBTH method ... 

Where applicable, the MBTH-method is of value for determining total sialic acid concentration without prior hydrolysis or sialidase treatment. However, the problems arising with 0-acyl-sialic acids and periodate oxidation also apply here, and saponification may be necessary for the complete assay of sialic acids. 

The level of 0-acylation of sialic acids can also be quantitated by difference measurements with the periodic acid/thiobarbituric acid method (Neuberger and Ratcliffe 1972, Skoza and Mohos 1976, Sarris and Palade 1979, see section II. 2), with the periodate/resorcinol (II. 1. b)p)) and the MBTH method (II. 3), and a fluorimetric method (Shukla and Schauer 1981, II. 4). Using such assays care should be taken to run suitable controls for the saponification step and to consider... 

Miyazawa et al,143 determined the Amadori compounds from dioleylphos-phatidylethanolamine by normal-phase HPLC as the 3-methyl-2-benzothiazolinone hydrazones (MBTH) through their UV absorption (318 nm) with detection limits of 4.5 and 5.3 ng for the fructosyl (F) and the lactulosyl (L) compound, respectively, and linear ranges of up to about 2 ng. Using their method, they obtained the following results infant formula, F 32-112, L 49-88 ng g mayonnaise, F 12.2 ng g-1 chocolate, F 3.9, L 1.5 ng g-1 cow s milk, L 0.079 /xg mL-1 soybean milk, F 0.24, L 0.13 /tg mL-1 and rat plasma, F 0.23 ng mL-1. Significant amounts were not detected in human milk. 

In some methods, the final mixture is acidified slightly to stop the reaction, and the intensity of the yellow chro-mophore is measured at 400 nm. In stronger acid solution, the color becomes pink, with maximum absorbance at 540 nm, and both sensitivity and stability are improved. Other approaches to measurement of the H2O2 produced include the peroxide-mediated oxidative coupMng of 3-methyl 2-benzothia2olinone hydrazone (MBTH) with N,N-dimethyl-aniline (DMA) catalyzed by peroxidase or the oxidative coupHng of p-aminophenazone (PAP) to phenoi. Both procedures have been automated. The... 

An alternative method for POase detection using modified MBTH has been reported (Capaldi and Taylor, 1983) but not yet applied in EIA. 

Conyers and Kidwell (1991) proposed a modified CN/ enhancer amine method of POase detection (Table 7.10E) that increases its detectability to that of APase using BCIP/NBT (Fig. 7.9.III). Among the three amines proposed, the concentration of MBTH was most critical for optimal detection. It offers the possibility, due to the red precipitation, of distinguishing two probe systems on a single membrane or in situ preparations (e.g., with APase yielding a blue precipitate from BCIP/NBT). POase, oxidized by H2O2, will oxidize substituted phenylenediamines or MBTH (DH2) to cationic elec-... 

The European Pharmacopoeia has two different colorimetric methods for limit testing of free formaldehydes based on a condensation of the formaldehyde being determined with acetylacetone and MBTH respectively. 

Spectrophotometry utilizing the reagents 4-aminoantipyridine and MBTH is the classical technique for the nonspecific determination of phenolic compoimds. It is the basis for several official methods and was discussed earlier. 

A new colorimetric method for the measurement of free and glycosidically bound sialic acid was introduced by Durand et al. (1974) and Massamiri et al. (1978, 1979), based on the reaction of methyl-3-benzothiazolinone-2-hydrazone (MBTH) with the formaldehyde liberated on periodate oxidation of the sialic acid side chain. The molar extinction coefficient for Neu5Ac is 67,000 at 625 nm, and the method is thus similar in sensitivity to the periodic acid/thiobarbituric acid methods. 

Determination of phenolic groups was achieved by Besthorn s Hydrazone Method [33] which includes spectrophotometric measurements. In this method 3-methyl-2-benzothiozolinone (MBTH) interacts with the quinones produced by the enzyme to yield red products instead of brown colored pigments in the absence of the color reagent [34]. The pathway proposed by Rodriguez et al is shown in Scheme 3 [35]. 

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