Putrescine, cadaverine, and indole production by bacteria isolated from wild and aquacultured penaeid shrimp stored at 0, 12, 24, and 36 degrees C.

Putrescine, cadaverine, and indole production capabilities of bacteria isolated from wild domestic and aquacultured Ni-caraguan penaeid shrimp in progressive decomposition states were evaluated. The numbers and types of microorganisms responsible for the production of putrescine, cadaverine, and indole in wild and aquacultured shrimp increased with increasing decomposition temperature and time. Throughout the storage experiments, mean aerobic plate counts (log/g) ranged from 4.5 to 9.7 and 4.5 to 9.0 for domestic and Nicaraguan shrimp, respectively. Vibrio spp. were more prominent in Nicaraguan shrimp (Litopenaeus vannamei) than in domestic shrimp (Litopenaeus setiferus and Litopenaeus brasiliensis). The only amine-producing (putrescine) microorganism isolated from wild and aquacultured shrimp at all temperatures of decomposition (0, 12, 24, and 36 degrees C) was Shewanella putrefaciens. On the basis of putrescine production by S. putrefaciens at 0 and 12 degrees C and putrescine production by S. putrefaciens, Vibrio spp., and Morganella morganii at 24 and 36 degrees C, putrescine should be considered a potential chemical indicator of decomposition in shrimp.

Gas chromatographic determination of cadaverine, putrescine, and histamine in foods.

A GLC procedure for the quantitative determination of the diamines putrescine and cadaverine has been developed, using their perfluoropropionyl derivatives. The amines were extracted from foods with methanol; an interval standard, hexanediamine, was added and a dry residue of their hydrochloride salts was prepared. The salts were derivatized with perfluoropropionic anhydride by heating for 30 min at 50 degrees C. The reaction mixture was separated on an alumina column to remove excess reagent, and the derivatives were eluted with a solution of 30% ethyl acetate in toluene. GLC separations were performed on a 3% OV-225 column held at 180 degrees C. The retention times were 4.3, 5.7, and 7.0 min for the derivatives for putrescine, cadaverine, and the internal standard, respectively. Less than 1 microgram diamine/g tissue could be quantitated, using either an electron capture detector or a nitrogen-specific detector. The procedure was applied to cheese and a variety of fishery products. An increase in the diamines correlated with the presence of decomposition in some of the products. A collaborative study of the method is planned.

High pressure liquid chromatographic method for indole in shrimp: development of method and collaborative study.

A collaborative study on the determination of indole in shrimp was conducted in which a high pressure liquid chromatographic (HPLC) method and a spectrofluorometric method were compared with the AOAC gas-liquid chromatographic (GLC) method (18.075-18.078, 13th ed.). In the HPLC method, 10 g shrimp was blended with methanol, an internal standard was added, and the extract was filtered. Indole was separated on an octadecylsilane reverse phase column, using 60% MeOH-H2O, and quantitated with a fluorescence detector (excitation 280 nm, emission 330 nm) by comparing the indole peak height with that of an internal standard, 2-methyl-indole. Recoveries at a 25 micrograms/100 g level averaged 104% with a range of 90-127%, and at a level of 35 micrograms/100 g averaged 102% with a range of 93-112%. In the spectrofluorometric method, 25 g shrimp was extracted with 2% EtOAc-hexane. After several washes, indole was partitioned into a saturated NaCl-MeOH solution and its fluorescence was measured (excitation 280 nm, emission 332 nm). Recoveries at a 25 micrograms/100 g level averaged 93% with a range of 0-255% and at a level of 35 micrograms/100 g averaged 64% with a range of 0-107%. Recoveries obtained by the AOAC-GLC method at a level of 25 micrograms/100 g averaged 96% with a range of 81-116% and at a level of 35 micrograms/100 g averaged 101% with a range of 81-119%. The coefficients of variation were 20, 10, and 64% at a 25 micrograms/100 g level for the GLC method, the HPLC method, and the spectrofluorometric method, respectively. The HPLC method was adopted as official first action for indole levels in shrimp exceeding 1 microgram/100 g.

Ultraviolet detection procedure for liquid chromatographic determination of indole in shrimp.

In an ultraviolet detection procedure, indole was extracted from shrimp by blending with carbonate buffer and then partitioned into ethyl acetate. Before quantitation by high pressure liquid chromatography (HPLC), interferences were removed by the use of a polyamide cleanup column. Concentrated column effluent was injected into the chromatograph. Indole was detected by monitoring absorbance at 217 nm and quantitated by comparison of indole peak height to that of an internal standard, 2-methylindole. Recoveries of indole ranged from 72 to 117%. Results agreed well with those by the official AOAC gas-liquid chromatographic (GLC) method.

Fluorometric determination of histamine in cheese.

Thirty-one samples of cheese obtained from retail outlets were analyzed for histamine, using an official AOAC fluorometric method. The types of cheese analyzed and the ranges of histamine found were: colby, 0.3--2.8; camembert, 0.4--4.2; cheddar, 1.2--5.8; gouda, 1.3--2.4; provolone, 2.0--23.5; roquefort, 1.0--16.8; mozzarella 1.6--5.0; and swiss, 0.4--250 mg histamine/100 g. Ten of the 12 samples of swiss cheese contained less than 16 mg histamine/100 g. The remaining 2 samples which contained 116 and 250 mg histamine/100 g were judged organoleptically to be of poor quality. An investigation of one processing facility showed that the production of histamine in swiss cheese may have been a result of a hydrogen peroxide/low temperature treatment of the milk supply. Recovery of histamine added to methanol extracts of cheese ranged from 93 to 105%. Histamine content was confirmed by high pressure liquid chromatographic analysis of the methanol extracts.

Fluorometric determination of histamine in tuna: development of method.

Tuna extracts are treated with an anion exchange resin to remove interfering materials, histamine is derivatized with o-phthaladehyde, and the fluorescence of the resulting compound is measured fluorometrically. Replicate analyses of acceptable and decomposed tuna packed in oil or water agreed within 1 mg at a level of 10 mg/100 g and within 12 mg at a level of 100 mg/100 g. Recoveries of histamine added to fish were greater than 90 and greater than 83% at levels of 10 and 100 mg/100 g, respectively. The new method is more rapid and specific and is simpler than previous methods because no liquid-liquid extractions or chromatographic separations of histamine are required. The sensitivity of the method allows quantitation of less than 10 mg histamine/100 g sample. The accuracy and precision of the fluorometric method are comparable to those of the official AOAC colorimetric method.

Fluorometric determination of histamine in tuna: collaborative study.

Six samples of canned tuna, albacore, yellowfin, and skipjack, in water or oil pack were analyzed in duplicate by a fluorometric method and the AOAC colorimetric method. For the fluorometric method, recoveries of histamine added to acceptable tuna averaged 99% with a range of 91 to 107%. Agreement between laboratories for the analyses of decomposed tuna containing 20-200 mg histamine/100 g sample was excellent. Results from the fluorometric method are comparable with those from the AOAC colorimetric method; the fluorometric method has been adopted as official first action.