Test for Detecting Spoilage in Beef

Transcription

1 APPLIED MICROBIOLOGY Vol. 12, No. 4, p July, 1964 Copyright 1964 American Society for Microbiology Printed in U.S.A. Evaluation of the Extract-Release Volume Phenomenon as a Rapid Test for Detecting Spoilage in Beef JAMES M. JAY AND KALLIOPI S. KONTOU Department of Biology, Wayne State University, Detroit, Michigan ABSTRACT JAY, JAMES M. (Wayne State University, Detroit, Mich.), AND KALLIOPI S. KONTOU. Evaluation of the extract-release volume phenomenon as a rapid test for determining spoilage in beef. Appl. Microbiol. 12: Ground beef of several grades, obtained fresh and held refrigerated until spoiled, was presented to a test panel for scoring on color, odor, and tactile response (tackiness) as to degree of acceptance. Panel scores were correlated with total bacterial counts, ninhydrin-reactive substances, and ERV (extract-release volume) on the same meat. ERV correlated highest with bacterial counts the largest number of times; tackiness, odor, ninhydrin, and color followed in that order. Correlation between bacterial numbers and organoleptic qualities was best, with tackiness followed closely by odor, and then by color. Correlation between tackiness and odor was high. The degree of correlation between bacterial numbers, tackiness, and ERV was high enough to warrant the use of the ERV phenomenon as a rapid test of microbial quality of beef. An ERV value of 25 under the conditions of the test was supported as a divider between acceptable and unacceptable ground beef. Received for publication 31 March 1964 In a previous communication from this laboratory, the extract-release volume (ERV) phenomenon of beef was shown to respond to spoilage in beef by a linear reduction in value with the intensification of the spoilage process (Jay, Food Technol., in press). This procedure is based on the amount of aqueous extract released from a slurry of meat when allowed to pass through filter paper for a given period of time. Fresh beef of good organoleptic quality with a relatively low bacterial number releases large volumes of extract (high ERV), whereas beef in the process of microbial spoilage with a higher bacterial number releases less (low ERV). In the previous report, highly significant differences were shown between ERV values and bacterial numbers from beef of good microbial quality with no undesirable organoleptic qualities as compared with beef held until definite spoilage had set in. To be meaningful, a test for the detection of spoilage in meats, among other things, should respond to the desirable and undesirable organoleptic qualities that one generally associates with it. None of the common indices of microbial quality in use today meet this criterion. It seemed desirable to compare ERV results with those of a test panel in an effort (i) to determine whether the two sets of data correlated and, if so, to what degree; (ii) to relate test panel 378 findings to ERV results, bacterial numbers, and a-anmino acid values; and (iii) to determine a more meaningful and precise ERV cutoff value between beef of good and pooi microbial quality. Previous work indicated that this value lay between 20 and 30 ml under the conditions of the test. In this report, we present results of a test panel evaluation of ground beef of different grades and age with respect to odor, color, and tactile response, which is compared with ERV data, bacterial numbers, and ninhydrin-positive substances from the same meat. A statistical treatment of the data is given. MATERIALS AND METHODS The ERV test, bacterial numbers, and ninhydrin-positive substances (a-amino acids, hereinafter referred to as amino acids) were determined as previously described (Jay, in press). The panel consisted of four to six persons (average number, five) and was composed of graduate students and faculty in Biology and Home Economics. The test panel was asked to score raw ground beef on a 10-point scale by comparing the test meat with a reference sample which was provided fresh-ground daily at the same time and treated in every other way as the test samples. The 10-point scale was divided as follows: 1 to 5, undesirable, and 5 to 10, desirable, with a score of 10 being highly desirable. The test meat was coarse-ground (0.64-cm plate) and held under refrigeration in large wax-paper covered pans. On test days, coarse-ground meat was mixed in the pan, and portions were removed and fine-ground (0.32-cm plate) for the panel. A giant hamburger-type pattie was prepared, and each panelist was presented a slice and asked to score it on color, odor, and tactile response (tackiness). The reference samples consisted of either frozen U.S. Choice or Prime round which was removed from the freezer and ground just before the others. Portions of each were removed for analysis prior to preparation for panel. The meat employed consisted of approximately equal portions of the various hindquarter muscles. RESULTS Table 1 presents the averaged values for all reference samples employed in this study. The ERV and panel values were high, and the bacterial numbers were relatively low. The percentage of fat in the reference samples ranged from 10 to 12. The statistical treatment of these meat data

2 VOL. 12) 1964 RAPID TEST FOR DETECTING SPOILAGE IN BEEF 379 (mean, SD, and correlation or r) are presented in Tables 2 and 3, where it can be seen that r was low and organoleptic values were high along with ERV. The mean bacterial number per gram was log Figures 1 and 2 present the ERV values, bacterial numbers, and panel data from 1-day-old cutter-grade beef without and with added fat, respectively. The ERV decreased steadily during the test period as the bacterial number increased, in both series. The panel rated both sets of meat poor on color between the first and fifth days. Color was rated an average of 6 on the seventh and eighth days in the TABLE 1. Average of values for all reference samples employed Days ERV Log bac- Amino Odor Tactile Color terial no. acids _ X* SD * X = mean. Panel sample all-lean series, and it was rated an average of 8 in the sample with 12 % fat (Fig. 2). This property of fresh beef appearing light in color at the onset of incipient spoilage has been observed repeatedly, and it appears to coincide with the appearance of a fairly large bacterial population. Odor and tactile response were rated almost the same in both series, and both characteristics are seen to have been scored rather erratically, with the fat-containing samples showing much the same pattern as for color. It is interesting to note that the panel rejected the fat-containing series on the third day but accepted portions of the same meat 1 and 2 days later. These two series were the first presented to the panel, and this might explain the erratic pattern of the scored qualities. The mean, SD, and r of the data illustrated are presented in Tables 2 and 3. The r value between ERV and bacterial numbers was significant at the 5 % level on the all-lean series, whereas no statistically significant r values existed between the others. Data on ERV and bacterial numbers, and scores on tackiness and odor in the fat-containing series, showed significance at the 5 % level. On the whole, the data of the fat-containing series showed higher degrees of correlation than those of the all-lean meat. The low rating on tackiness given the all-lean meat on the first day was probably due to the fact that this meat was only 1 day postmortem, and such beef tends to be tacky until it reaches its full state of rigor. This no doubt explains the low color rating as well. The TABLE 2. Mean and standard deviation of test panel data presented in Fig. 1, 2, 3, 5, 6, and 7 and Table 1 ERV Bacterial no. Tactile Odor Color a-amino acids SD X SD X SD X SD X SD X SD Fig Fig Fig Fig Fig Fig Table * X = mean. TABLE 3. Statistical correlation (r) between ERV, bacterial numbers, a-amino acids, and test panel scores on beef color, odor, and tactile response illustrated in Fig. 1, 2, 8, 5, 6, and 7, and Table 1 Panel sample ERV Bacterial no. Tactile Bacterial no. a-amino acids Tactile Odor Color Tactile Odor Color Odor Fig * a Fig * * Fig * Fig * * * Fig Fig * Table ** 0.772* 0.888** 0.904** * Significant at the 5% level. ** Significant at the 1% level. A dash indicates no data.

3 380 JAY AND KONTOU APPL. NIICROBIOL. lack of fat in the case of the meat rated in Fig. 1 would tend to make the color less desirable to a panel. The meat employed in the evaluations of Fig. 3 was the same as that of Fig. 1 and 2 after aging 15 days in a cold room on the hindquarter before grinding. Again, ERV dropped between the third and fifth days as the bacterial numbers increased. The panel rejected this meat the fifth day on both odor and tackiness, whereas it rated color good throughout. Just what accounted for the rise in tackiness and odor values on the day after rejection is not clear. The amino acid values decreased rather steadily throughout the holding period. This trend has been observed repeatedly. The r value between ERV and bacterial numbers is seen again to be rather high in Table 3, even though not significant at the 5 % level. The correlation between tackiness and odor was, however, significant at the 5 % level. A sample of U.S. Good beef was employed in the panel series shown in Fig. 4. This meat was ground 7 days after slaughter with approximately 10 % fat added. It can be seen that the ERV again fell sharply after the fourth day, TIME () FIG. 1. Test panel scores on color, odor, and tackiness along with ERV values and bacterial numbers from a sample series consisting of all-lean cutter-grade beef starting 1 day after slaughter. See Tables 2 and S for statistical data. 8 60r c -P and the bacterial numbers increased steadily from log 7.20 to 9.45 in the 5-day period. The panel rejected this meat the third day on odor and tackiness, and it became spoiled between the fourth and fifth days by ERV. The low score for color on the first day was probably due to the fresh ground state of this meat at the time it was presented to the panel. It can be seen again that amino acid values decreased steadily as the meat continued to spoil. Figure 5 presents the data on a portion of the meat employed in Fig. 4 after being frozen for 21 days. This meat was allowed to thaw at room temperature prior to being coarse-ground and placed in panis for panel use. ERV and bacterial numbers were essentially the same after freezing as before. The panel rejected the frozen on the third day, as it did the fresh. The most significant difference between the two series in question was the higher initial level of amino acids in the frozen. The r between ERV and log numbers was significant at the 5 % level, as was the case with log numbers and tackiness, and tackiness and odor. Figures 6 and 7 show data from a standard-grade cow approximately 5 days after slaughter, with the series of 9 35.\.*10 a 30,- _XERV /" OR_ CL- 8.5 o 6 >20t -0 X4 > K I 0 --zw z 3 5 / 4.1 FIG. 3. Panel scores on organoleptic qualities along with ERV values, bacterial numbers, and ninhydrin-positive substances (amino acids) from a series of samples of cutter-grade beef held 15 days after slaughter containing approximately 10% fat. <,>6 4 w 0 m > cn5 x a -i w z 4 3 FIG. 2. Test panel scores on organoleptic qualities along with ERV values and bacterial numbers from cutter-grade beef starting 1 day after slaughter containing 12% fat. See Tables 2 and S for statistical data. 0 '' -- FIG. 4. ERV values, bacterial counts, amino acid levels, and test panel scores on a series of samples from U.S. Good grade beef starting 7 days postmortem.

4 VOL RAPID TEST FOR DETECTING SPOILAGE IN BEEF 381 the former having no fat whereas the latter contained approxinmately 10 %. ERV, bacterial numbers, amino acids, and panel data were essentially the same in both cases. The panel rejected the all-lean meat on the second day and the fat-containing meat 1 day later. In the case of the latter series, ERV values correlated quite well with all others, with each following essentially the same downward course with the intensification of the spoilage process. While amino acid and ERV values behaved the same way with the all-lean meat, panel data were more erratic due possibly to a lack of fat in the meat. With respect to r between ERV and bacterial numbers, significance at the 5 % level is seen in four of the six cases presented (Table 3). It should be noted that while ERV did not correlate significantly with the panel's determination of odor and tackiness, r values are nevertheless rather high, with those for tackiness being higher than those for odor, except in the case of the first sample (Fig. 1). It should be further noted that r between ERV and color was the lowest. 9 (0 10 6( U 0 > -J 71L-. z r (04 0~ z ;o FIG. 5. Test panel scores and other data from a series of samples of U.S. Good grade beef held frozenfor 21 days. See Tables 2 and 3 for statistical treatment of these data l\\- _ C9 Ia J2 4 l0ot.~ 6 FIG. 6. All-lean standard-grade beef samples scored on color, odor, and tackiness, along with ERV, amino acid, and bacterial count data..2 Table 3 further shows that r between odor and tackiness was highly significant (1 % level) in one of the panel series subjected to this analysis, significant (5 %) in three others, and rather low in the first panel series. The r value between ERV and amino acids was not statistically significant in the four samples analyzed, but was rather high in two. Data are presented in Table 4 from ten beef samples which the panel accepted on all three criteria (color, odor, and tackiness above 5), and a similar number which were rejected on all three criteria (scores below 5), with the exception of four that were scored acceptable on color alone. Some of these data were taken from samples prepared by mixing various amounts of putrid beef with fresh beef prior to fine-grinding, and others were taken at random from some of the foregoing panel test series. The object of the data presented in Table 4 is to show the degree of difference between panel-accepted and panel-rejected meat and the extent to which the various indices responded to this difference. Panel scores on accepted meat were high for all three qualities with mean values of 7.99, 8.25, and 8 for tackiness, odor, and color, respectively (average, 8.07). Bacterial numbers per gram were generally low, with a range of log 6.3 to 8.85 (mean, 7.03). ERV values ranged from 25.5 to 51.5 (mean, 37.6) with a SD of On the other hand, the rejected beef had low ERV values, with a range of 3.5 to 36.5 (mean, 17.5) with a SD of Bacterial numbers on rejected samples averaged log 9.13, with the averages for tackiness, odor, and color being 3.59, 3.46, and 4.97, respectively (average, 3.98). The greatest per cent difference based on mean values between accepted and rejected meat was found with odor scores (58), followed by tackiness (55), ERV (53), color (38), 8 7 c 6 w J 5 -J w z4a Co 0 z cd.3a 30 0 FIG. 7. Panel scores, ERV, bacterial numbers, and amino acid values from portions of the same beef of Fig. 6 with 10 to 12% added fat..2.1

5 382 JAY AND KONTOU APPL. MICROBIOL. TABLE 4. Sample no. Bacterial numbers, ERV values, and panel evaluations on ten panel-accepted and ten panel-rejected samples of raw ground beef ERV Bacterial no. Tackiness Odor Color Panel avgb ACCa REJ ACC REJ ACC REJ ACC REJ ACC REJ ACC REJ Mean SD Per cent differencec P 7.05**** 6.43**** 9.82**** 8.87**** 5.12**** 8.51**** a ACC, accepted by panel; REJ, rejected by panel. b Average of tackiness, odor, and color values. c Based on mean values. **** Significant above the 0.1% level. and bacterial numbers (23); the average of organoleptic qualities was 51. The statistical significance between the differences in all cases is seen to be extremely high (P < 0.001), with the difference between tackiness being highest, followed by odor, ERV, bacterial numbers, and color. With respect to the ERV value that corresponded with panel rejection, the data in Table 5 taken from Fig. 1 to 5 show a mean value of 30.4 (range 21 to 35 ml) and a mean log bacterial number of 7.85 (ca. 70,000,000). The panel scores indicated in Fig. 6 and 7 were not included, since the panel had only one opportunity to score this meat before offness developed. DISCUSSION The degree of correlation between ERV and amino acid levels was not very significant, although high r values were obtained in two cases. The apparent slow decrease in TABLE 5. ERV and bacterial numbers of six series of samples at the earliest time of rejection by averages of all panel scores Panel sample ERV Log bacterial no. No. of days held Fig Fig Fig Fig Fig X* SD * X = mean. quantity of amino acids with an increase in bacterial numbers was observed consistently. This phenomenon was observed also by Saffle et al. (1961), and, as was found by these authors, this index appears not to be reliable as an index of microbial quality in beef. The highest degree of r between any two panel indices in this study was found between scores for tackiness and odor where three of the six experimental series samples were statistically significant at the 5 % level. A high degree of r between these two organoleptic qualities was to be expected, and this, therefore, might be taken to indicate the degree of competence of the panel. It should be noted that the panel data presented represent the scores of all panel participants at each scoring. When scores of one or more members who deviated widely from the others were removed, the same general pattern as shown was maintained, but plots of the scores produced less erratic curves and r values tended to be slightly higher when related to ERV and bacterial numbers (Kontou, 1964). Kraft and Ayres (1952) and Ayres (1960), along with other investigators, have shown the relationship between odor and slime development on refrigerated meats. These authors found that off odor development preceded sliminess and that sliminess was directly related to bacterial numbers. Therefore, a high degree of correlation between tackiness (sliminess) and odor should indicate a similar correlation between either of these values and bacterial numbers. In the present study, r between bacterial numbers and tackiness was somewhat higher than odor. Of the two, odor would be less reliable than tackiness, since it is more subjective. Too, off odors are not always associated

6 VOL. 12) 1964 RAPID TEST FOR DETECTING SPOILAGE IN BEEF 383 with incipient microbial spoilage, as Shank, Silliker, and Goeser (1962) demonstrated in beef an off odor condition not directly related to microbial spoilage. Although the panel did not highly relate ERV to the organoleptic qualities, this test correlated four of six times to bacterial numbers at the 5 % level of significance. Since large numbers of bacteria are generally associated with spoiled beef, ERV determinations would be a much faster way of indicating the approximate microbial quality of beef than are bacterial plate counts; the former require only about 20 min, and the latter require several days for results. The mean ERV value of 30.4 at the earliest time of rejection by the average values of all panel scores (Table 5) is higher than a value obtained previously when one of us (J.M.J.) scored a larger number of samples of ground beef under similar conditions in the laboratory. The mean ERV at the time of offness from 26 different samples was 24.6 (SD, 5.7) with an average log bacterial number per gram of 8.48 (Jay, in press). Of the 26, 5 had ERV values above 30, and 6 were below 20. When all panel scores (Fig. 1 to 7) were averaged, the number of days from freshness to rejection was, respectively, 3.0, 3.6, and 4.3, on color, odor, and tackiness; corresponding log bacterial numbers per gram were 6.6, 7.3, and 8.2, and the corresponding ERV was 43, 35, and 25. In other words, if tackiness scores are taken to reflect more accurately the desirable microbial condition of ground beef, the panel confirmed the previous finding of 25 as a probable cut off value for ERV. This value is equivalent to an approximate log bacterial number of 150,000,000 per g. Since ERV is affected somewhat by grade of beef and added proteases (Jay, in press), it seems desirable to conduct more determinations before a precise value is settled upon that would separate desirable and undesirable beef from the microbial standpoint. Almost any value, of course, would be somewhat subjective, and ERV offers a type of flexibility that can be utilized to compensate for this. ACKNOWLEDGMENTS Appreciation is expressed to Jayne C. Bates and Gwendolyn D. Wright for technical assistance and to Marqueta C. Huyck of the Home Economics Department for assistance on the panel study. This investigation was supported by Public Health Service research grant EF from the National Institutes of Health. Contribution no. 115 from the Department of Biology, College of Liberal Arts. LITERATURE CITED AYRES, J. C Temperature relationships and some other characteristics of the microbial flora developing on refrigerated beef. Food Res. 25:1-18. KONTOU, K. S An organoleptic evaluation from freshness to frank spoilage of paired raw and cooked ground beef. M.S. Thesis, Wayne State University, Detroit, Mich. KRAFT, A. A., AND J. C. AYRES Post-mortem changes in stored meats. IV. Effect of packing materials on keeping quality of self-service meats. Food Technol. 6:8-12. SAFFLE, R. L., K. N. MAY, H. A. HAMID, AND J. D. IRBY Comparing three rapid methods of detecting spoilage in meat. Food Technol. 15: SHANK, J. L., J. H. SILLIKER, AND P. A. GOESER The development of a nonmicrobial off-condition in fresh meat. Appl. Microbiol. 10: