Characterization of Whey Cheese Packaged Under Vacuum

216

Journal of Food Protection, Vol. 63, No. 2, 2000, Pages 216±221 Copyright ᮊ, International Association for Food Protection

Characterization of Whey Cheese Packaged under Vacuum

MANUELA E. PINTADO AND F. XAVIER MALCATA*

Escola Superior de Biotecnologia, Universidade CatoÂlica Portuguesa, Rua Dr. AntoÂnio Bernardino de Almeida, P-4200-072 Porto, Portugal

MS 99-24: Received 12 February 1999/Accepted 28 June 1999

ABSTRACT Њ

Vacuum packaging was assayed at 4 C and was tested in comparison to unpackaged counterparts, in both microbiological Downloaded from http://meridian.allenpress.com/jfp/article-pdf/63/2/216/1673220/0362-028x-63_2_216.pdf by guest on 24 September 2021 and physicochemical terms, in studies pertaining to the preservation of RequeijaÄo, a traditional Portuguese whey cheese. Bacteria were absent (i.e., Ͻ10 CFU/g) in whey cheeses on the day of manufacture as a result of thermal processing. After storage, both unpackaged and packaged cheeses exhibited high viable counts of Bacillus, Pseudomonas, Enterobacteriaceae, and lactic acid bacteria (especially lactococci). Yeasts, staphylococci, enterococci, and spore-forming clostridia were severely inhibited by the package vacuum combined with the increasing acidi®cation developed therein. Whey cheeses packaged under vacuum underwent substantial acidi®cation, slight depletion of lactose, and no signi®cant variation in moisture content or texture; conversely, unpackaged whey cheeses exhibited substantial loss of water and a concomitant increase in rigidity. Vacuum packaging strongly inhibited lipolysis (even if viable counts of some microbial groups were high); saturated fatty acids (mainly C16:0 and C14:0) accounted for ca. 73% of the total free±fatty acid content, whereas the most concentrated unsaturated fatty acids were C18:1 and C18:2 (ca. 14% each). The conclusions generated in our study are, in general, useful for a wide range of whey cheeses worldwide: i.e., RequeÂson (Spain), Ricotta (Italy), Broccio (France), and Anthotyro (Greece). In addition, our conclusions are particularly helpful in terms of improving the safety of RequeijaÄo, a widely acclaimed dairy specialty.

Large amounts of whey are produced worldwide as a (after it is seasoned with salt, sugar, or honey) and is sold by-product of cheese manufacture. A fraction of this whey in its mold at virtually every supermarket nationwide (27). is still used in some Mediterranean regions to produce whey The lack of ef®cient packages greatly affects the safety as- cheeses following traditional protocols (9). Whey cheeses sociated with consumption of this cheese and also severely fall under several designations, e.g., RequeijaÄo (Portugal), constrains its shelf life, major concerns arise from environ- Broccio (France), RequeÂson (Spain), Ricotta (Italy), and ment-borne microorganismsÐnamely psychrotrophs, Anthotyro (Greece). Coagulation of proteins in cheese yeasts, and molds (even if good sanitary handling was en- whey via heating has been proven to be an interesting and sured along the trade chain)Ðso development of tailor- easy means to recover whey proteins (17, 22); on the other made packages is in order. Studies of cheese packaged un- hand, although denaturation of whey proteins does not af- der vacuum have been reported by Fedio et al. (8); studies fect signi®cantly their nutritional content, a posteriori ap- of cheese packaged under modi®ed atmosphere have been plication by the food industry is limited because of the in- reported by Rosenthal et al. (26) and by Fedio et al. (7). solubility, gritty aspect, and poor functional properties of The objective of this study was to characterize the whey proteins (31). Hence, whey cheese is one of the best physicochemical and microbiological evolution of Requei- vehicles for integral consumption of said whey proteins. jaÄo packaged under vacuum and stored at 4ЊC, and to com- Ricotta is the best-known whey cheese, and several pare this evolution with that of its unpackaged counterpart. studies have been performed in an attempt to improve the MATERIALS AND METHODS ®nal nutritional and textural properties of this cheese (12, 15, 16, 23, 29, 30); Anthotyro has also received some at- Preparation and packaging of whey cheese. Experimental tention (11), as has RequeijaÄo (24, 25). The richest (organ- whey cheeses were produced following the traditional manufac- oleptically) RequeijaÄo is produced on the farm level from turing protocol used for RequeijaÄo, a protocol that consisted of ovine whey, and such practice has surely derived from the heating the starting material (i.e., ovine whey, from Bordaleira great importance of dairy sheep, which has been observed sheep milk), which was mixed with 10% (vol/vol) caprine milk (from Serrana goats), at 95ЊC for ca. 15 min under gentle stirring. for ages in the Mediterranean Basin (14). The traditional The curd ¯oating on the surface was scooped out into plastic protocol for the manufacture of RequeijaÄo consists of heat- molds and was allowed to drain and cool down to room temper- ing whey (often ovine whey with ca. 10% (vol/vol) added ature for a few minutes; whey cheeses were then immediately caprine milk) at 95ЊC for at least 15 min under gentle stir- transported under sterile and refrigerated conditions to our labo- ring; the coagulum thus formed is then transferred with a ratory. Whey cheeses were then kept in their molds in the open scoop into perforated plastic or wooden molds and is left air at 4ЊC, so as to mimic the conditions prevailing in the exhi- to drain for a few hours. This cheese is usually eaten fresh bition refrigerated stand at the supermarket, or were vacuum packaged in Cryovac BB4L bags (Grace, Barcelona, Spain), using a * Author for correspondence. Multivac A300/42 machine (Wolfertschwenden, Germany) and a J. Food Prot., Vol. 63, No. 2 VACUUM-PACKAGED WHEY CHEESE 217 vacuum of 100 mbar, for 2 s. An additional experiment employing protein interference; 10 g of sample was homogenized with 30 ml packaging with air led to such poor results that it was dropped of a 0.5 M solution of perchloric acid (Merck) for 3 min in a from further consideration; such results included signi®cantly Stomacher Lab Blender 400, was allowed to stand for 2 h in a worse performance compared with the results of the open-air ex- closed vessel at refrigeration temperature, and was ®nally ®ltered periment. The thickness of the (multilayer) package ®lm was 9 Ϯ through a 0.22-␮m Syr®l ®lter membrane (Nucleopore, Cam- 1 ␮m for the inner layer, 18 Ϯ 1 ␮m for the middle layer, and bridge, Mass.). 37 Ϯ 1 ␮m for the outer layer. The package ®lm could be con- Complete extraction of free fatty acids (FFAs) was achieved sidered to be a high barrier, because the transmission rate of steam with diethylether, according to Nunez et al. (19), using6gof through this ®lm was 1.08 Ϯ 0.17 g´mϪ2´dϪ1, the transmission cheese in each assay. Quanti®cation was performed according to rate of oxygen was 0.701 ml´mϪ2´dϪ1´bar, the transmission rate of the method initially described by Garcia et al. (10) and later mod- carbon dioxide was 0.340 ml´mϪ2´dϪ1´bar, and the transmission i®ed by BalcaÄo and Malcata (1), using p-bromophenacylbromide rate of nitrogen was 0.328 ml´mϪ2´dϪ1´bar. All whey cheeses were (Sigma), 18-crown-6-ether (Sigma), and potassium carbonate stored at 4ЊC. Experiments run at all storage times (0, 2, 6, 10, (Merck) for derivatization, and formic acid (Romil, Leicester, UK) and 15 days) were replicated and duly analyzed as described be- for quenching. Nonanoic and heptadecanoic acids (in the concen- Downloaded from http://meridian.allenpress.com/jfp/article-pdf/63/2/216/1673220/0362-028x-63_2_216.pdf by guest on 24 September 2021 low. trations of 6.41 and 11.13 mM, respectively) were used as internal standards. To 1.5 ml of FFA extracts, selected volumes of solution Microbiological analyses. A sample of whey cheese (10 g), of internal standards (ranging from 0.1 to 0.6 ml, as appropriate) taken randomly and aseptically, was homogenized with 90 ml of were added prior to derivatization. Separation was effected in a sterile 2% (wt/vol) sodium citrate (Merck, Frankfurt, Germany) C-18 reverse phase column (Beckman, San Ramon, Calif.) at 33ЊC for 3 min at room temperature in a Stomacher Lab Blender 400 using 5% (vol/vol) acetonitrile in methanol (Romil) and 5% (vol/ (Seward Medical, London, UK). Serial decimal dilutions were vol) acetonitrile in water as solvents and 20 ␮l as sample injection prepared with 0.1% (wt/vol) peptone water (Sigma Chemical Co., volume; detection was by absorbance at 254 nm. St. Louis, Mo.) and were plated in duplicate on a variety of media, Rheological measurements. Force-displacement curves of as follows: total aerobic count of mesophilic and psychrophilic cheeses were recorded using an Instron puncture tester (model bacteria on plate count agar (LAB M, Bury, UK), incubated for 4501, High Wycombe, UK). Puncture was effected with a 5-mm 2 days at 30ЊC and for 7 days at 6ЊC, respectively; lactococci on plunger at a constant penetration rate of 20 mm/min for a ®xed M17 agar (Lab M), supplemented with cycloheximide (100 mg/ height (25 mm). Six determinations per cheese were made at room liter) (Sigma) to inhibit yeast growth, incubated at 30ЊC for 2 days temperature (23ЊC), and only consistent data were taken for fur- under anaerobic conditions; lactobacilli on Rogosa agar (Oxoid, ther consideration. Basingstoke, UK), supplemented with cycloheximide (100 mg/ liter), incubated at 30ЊC for 5 days under anaerobic conditions; Statistical analysis. The whole data were statistically pro- staphylococci on Baird Parker medium (Lab M), supplemented cessed using analysis of variance to assess the effects of the type with egg yolk tellurite (Lab M), incubated at 37ЊC for 2 days; of package and storage time. Fisher's Protected least signi®cant Enterobacteriaceae on violet red bile glucose agar (Lab M), with difference t test was applied to all experimental results in order overlay, incubated at 37ЊC for 1 day; yeasts and molds on potato to assess intrapair signi®cant differences at the 5% signi®cance dextrose agar (Lab M), supplemented with sterile 10% (vol/vol) level. All statistical analyses were performed with the Statview lactic acid, incubated at 25ЊC for 5 days; Bacillus spp. on Bacillus package (Abacus Concepts, Berkeley, Calif.). cereus agar (Lab M), incubated at 30ЊC for 2 days; enterococci on KF agar (Lab M), incubated at 37ЊC for 1 day; Pseudomonas RESULTS AND DISCUSSION on Pseudomonas agar (Lab M), incubated at 30ЊC for 2 days; and Microbial growth. The (speci®c) growth rates were spore-forming clostridia, preheated at 80ЊC for 10 min and plated calculated as the difference between the log viable numbers on reinforced clostridium medium (Lab M), with overlay, supple- at each of two consecutive times (see Table 1) divided by mented with 4% (wt/vol) sodium sul®te (Merck) and 7% iron the corresponding time interval. Viable counts of microbial citrate (Merck), incubated anaerobically in an atmosphere of N 2 groups throughout storage time are summarized in Table 1. at 30ЊC for 3 days (28). All counts were performed according to the surface viable- On the day of manufacture, all cheeses contained less than count technique by Miles and Misra (13), except for the violet red 10 CFU/g. The total aerobic counts exhibited similar evo- bile glucose agar and the reinforced clostridium medium, which lution for vacuum-packaged (VP) and unpackaged (UNP) were determined according to Busta et al. (4). whey cheeses (i.e., a gradual increase throughout storage); however, the growth by 10 days was slower, especially in Chemical analyses. Determinations of pH were made with the VP whey cheese. In the VP cheeses, the counts of the measurements of the surface and bulk of cheese using a potenti- viable psychrotrophs decreased by ca. 0.5 log cycles by 15 ometer Crison Microph 2001 (Barcelona, Spain). The dry weight days. Viable counts of the same order of magnitude (109 was determined according to the International Dairy Federation CFU/g) in fresh Ricotta were also reported by Fadda et al. method (standard 4: 1958). (6). Lactose and lactic acid were quanti®ed in the same runÐ Bacillus spp. were present at very high counts but de- from calibration curves previously prepared with chromatographic creased between 10 and 15 days in the VP whey cheese. standardsÐusing a high-performance liquid chromatography sys- tem (Beckman Instruments, Fullerton, Calif.) with an Aminex Pseudomonas, lactococci, and Enterobacteriaceae under- HPX-87X cation exchange column (Bio-Rad, Richmond, Calif.), went similar evolution throughout storage. These major groups of bacteria in Ricotta cheese were generally consid- a ¯ow rate of 0.5 ml/min of 0.005 N H2SO4 (Merck) as eluant (20), and detection at 30ЊC by measurement of the refractive index ered to be important air-borne and utensil-borne contami- for lactose and measurement of UV absorbance for lactic acid. nants in dairy industries (2, 6, 11, 21). For these four mi- Prior to analysis, all samples were pretreated in order to eliminate crobial groups, vacuum packaging did not impart a signif- 218 PINTADO AND MALCATA J. Food Prot., Vol. 63, No. 2

TABLE 1. Experimental data obtained for viable numbers of various microbial groups in whey cheeses throughout storage at 4ЊC Experimental data (log [CFU/g])

Spore- Psychro- Entero- Cheese Storage Entero- Staphylo- forming Mesophilic philic Pseu- bacteri- Lactoba- Yeasts and package time (d) cocci Lactococci Bacillus cocci clostridia bacteria bacteria domonas aceae cilli molds

Without 0 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 2 0.00 4.13 4.03 3.86 0.00 4.57 4.19 3.63 0.00 3.24 0.00 6 3.94 4.60 5.30 4.43 0.00 6.19 6.46 5.45 3.94 4.57 4.56 10 5.98 7.93 7.22 5.58 2.69 8.78 8.54 8.41 5.89 5.92 6.04 15 5.20 9.01 8.66 5.36 5.15 9.35 9.45 9.49 7.85 7.05 6.15 Under 0 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 vacuum 2 0.00 4.53 3.90 0.00 0.00 4.93 4.70 4.92 0.00 0.00 0.00 Downloaded from http://meridian.allenpress.com/jfp/article-pdf/63/2/216/1673220/0362-028x-63_2_216.pdf by guest on 24 September 2021 6 0.00 5.28 4.98 0.00 0.00 7.01 6.97 6.89 4.32 4.77 0.00 10 0.00 7.42 8.49 3.98 0.00 8.47 8.15 7.57 6.69 5.22 3.35 15 4.81 8.54 7.14 5.39 3.59 8.65 7.66 8.60 7.13 5.41 3.86 icant advantage relative to the UNP whey cheese, because The analysis of variance indicated that the nature of no reductions were observed until 15 days. the package has statistically signi®cant effects (P Ͻ 0.05) The effect of vacuum upon lactobacilli became appar- for the microbiological groups measured; the Fisher's Pro- ent as soon as 2 days into storage, and a difference of ca. tected least signi®cant difference t tests were signi®cant for 1.5 log cycles was already observed by 15 days between all microbiological groups except pseudomonads (P ϭ the VP and UNP whey cheeses. Important differences were 0.1831) and Enterobacteriaceae (P ϭ 0.0853). The analysis also observed in the case of enterococci (see Table 1) and of variance and Fisher's Protected least signi®cant differ- staphylococci, as these groups were detected only between ence t test also indicated that storage time had signi®cant 10 and 15 days for the VP whey cheeses; nevertheless, the effects for all microbial groups considered, except Bacillus counts in VP whey cheeses became equivalent to those of spp. (P ϭ 0.1385) between 10 and 15 days. the UNP cheeses. Yeasts, molds, and spore-forming clos- Physicochemical characteristics. The values of the in- tridia exhibited increases in viable counts later during stor- ner and outer pH were similar for VP and UNP whey chees- age, especially in VP cheeses; such viable counts in VP es by 15 days of storage, and these values ranged from 6.7 cheeses were clearly lower than those for UNP cheeses, so to 6.0 and from 6.2 to 4.8, respectively (see Fig. 1). How- these observations unfold a notable in¯uence of vacuum ever, the VP whey cheese showed a higher degree of acid- upon the viability of these microorganisms. Yeasts, an im- i®cation on the surface in spite of its lower viable counts; portant family of microbial contaminants in the dairy in- this is probably a result of the production of organic acids, dustry, were not present in major numbers even in UNP as fermentative metabolism is favored over oxidative me- whey cheeses. Spore-forming clostridia, often described as tabolism (that would eventually lead to CO ) in the anaer- contaminants contributed by the rennet (5), displayed the 2 obic environment prevailing in the VP cheeses. The slight lowest counts in the whey cheeses. decrease in the outer pH suggests that most of the microbial population actually lays on the surface, hence con®rming the idea that environmental contamination of cheese is mainly responsible for such high microbial counts. The dry weight and the rupture slope (a rheological measurement) are depicted in Figure 2. Penetration tests were preferred to compression tests because they improved the reproducibility. The penetration tests have shown a good correlation with sensory analyses pertaining to the ®rmness and rigidity of the sample (3). From the curve, the rupture load, which is generally associated with ®rmness, did not exhibit a good reproducibility, unlike the rupture slope, which is usually associated with rigidity (33); there- fore, only the latter parameter was used to differentiate between the VP and the UNP whey cheeses. The value of the rupture slope for the VP whey cheeses did not show an important variation throughout storage, yet the rupture FIGURE 1. Evolution of inner pH (Ⅲ, ▫) (SEM ϭ 2.83 ϫ 10Ϫ3) slope of the UNP whey cheeses increased slightly by 2 days and outer pH (●, ⅙) (SEM ϭ 3.58 ϫ 10Ϫ3) throughout storage at and increased strongly between 10 and 15 days, as observed 4ЊC for unpackaged (solid symbols) whey cheese and whey cheese in Figure 2. Moisture content showed a good negative cor- packaged under vacuum (hollow symbols). relation with rupture slope (␳ϭ0.961) in the case of UNP J. Food Prot., Vol. 63, No. 2 VACUUM-PACKAGED WHEY CHEESE 219 Downloaded from http://meridian.allenpress.com/jfp/article-pdf/63/2/216/1673220/0362-028x-63_2_216.pdf by guest on 24 September 2021

FIGURE 2. Evolution of rupture slope (Ⅲ, ▫) (SEM ϭ 1.04 ϫ FIGURE 3. Evolution of lactose content (Ⅲ, ▫) (SEM ϭ 6.72 ϫ 10Ϫ3) and moisture content (●, ⅙) (SEM ϭ 2.34 ϫ 10Ϫ1) through- 10Ϫ1) and lactic acid content (●, ⅙) (SEM ϭ 1.55 ϫ 10Ϫ2) out storage at 4ЊC for unpackaged (solid symbols) whey cheese throughout storage at 4ЊC for unpackaged (solid symbols) whey and whey cheese packaged under vacuum (hollow symbols). cheese and whey cheese packaged under vacuum (hollow symbols). whey cheeses, an observation that agrees with the decrease crease was only observed for C in UNP and VP whey in rigidity of cheese that corresponds to a decrease in mois- 10:0 cheeses; for the long-chain fatty acid fraction (ϾC ), no ture content. The lactose content (see Fig. 3) did not un- 14:0 variation was observed that was signi®cant at the 5% level. dergo signi®cant variation throughout storage for the VP Despite the overall increase observed, especially in UNP whey cheeses, which indicates that a small fraction of mi- whey cheeses, the fraction of each individual FFA did not croorganisms can metabolize this sugar. Unexpectedly, a change signi®cantly throughout storage, except in the cases very important increase was observed in the case of UNP of C ,C ,C , and unsaturated FFAs in the UNP whey cheeses, which probably results from the concomitant 10:0 14:0 18:0 whey cheeses, and in the cases of C ,C , and unsat- decrease in moisture content. Lactic acid concentration (see 16:0 18:0 urated FFAs in the VP whey cheeses. Vacuum apparently Fig. 3) showed a slight increase between 2 and 6 days in inhibits lipolysis in RequeijaÄo, despite the high microbial VP whey cheeses, probably owing to microbial activity, but counts detected. a strong decrease was observed by 6 days, probably the Results obtained regarding fresh RequeijaÄo are similar result of catabolism of this acid, as observed elsewhere by to those obtained with Italian Ricotta (32) in terms of mois- Fedio et al. (8) in VP cheeses. This decrease led to an ture content, lactose concentration, and concentrations of increase of concentrations of shorter organic acids and of total saturated and unsaturated FFAs; however, while C CO , which will likely cause decreases in pH; this obser- 16:0 2 was the predominant unsaturated FFA (accounting for 20 vation is further con®rmed by the good correlation between to 25% [wt/wt]), our results indicated that the predominant outer pH and lactose content (␳ϭ0.979) and, to a lesser extent, with inner pH (␳ϭ0.942). The values for the total FFA content at various stages of storage for VP whey cheeses (see Fig. 4) displayed a slight increase between 6 (0.498 mg/g) and 15 days (0.734 mg/g), whereas UNP whey cheeses exhibited a very signi®cant increase between 6 (0.498 mg/g) and 15 days (6.332 mg/g). Several groups of microorganisms can play a role in lipolysis, viz., molds and bacteria (e.g., Pseudo- monads are proli®c lipase producers); however, molds are more affected by anaerobiosis than are bacteria, so the se- vere reduction in the concentration of FFAs when going from the UNP to the VP cheeses might be a clue regarding the dominant role of molds in lipolysis. The overall pro®le of FFAs in RequeijaÄo (see Table 2) was quite different from that seen in ovine cheeses (18): RequeijaÄo was richer in small chain FFAs (viz., C4:0,C6:0, and C8:0) and C12:0 and was poorer in C16:0 and C18:0. The small-chain fatty acid FIGURE 4. Evolution of total free fatty acid content (●, ⅙) (SEM Ͻ Ϫ fraction ( C10) did not undergo a signi®cant variation ϭ 1.78 ϫ 10 5) throughout storage at 4ЊC for unpackaged (solid throughout the 15-day storage period. Within the medium- symbols) whey cheese and whey cheese packaged under vacuum chain fatty acid fraction (C10:0 to C14:0), an important in- (hollow symbols). 220 PINTADO AND MALCATA J. Food Prot., Vol. 63, No. 2 7 Ϫ saturated FFAs are C16:0 and C14:0 (at ca. 11 to 13% [wt/ % 9.4 7.5 9.5 0.0 0.0 0.0 10 10.7 11.1 (w/w) wt] each), whereas C18:1 and C18:2 were the most abundant ϫ 18:3

C unsaturated FFAs (at ca. 13 to 14% [wt/wt] each).

mg/g The results of the analysis of variance have indicated 0.412 0.063 0.476 0.069 0.000 0.000 0.061 0.000 2.379 that the nature of the package accounted for statistically 6 Ϫ signi®cant effects (P Ͻ 0.05) regarding all chemical param- % 9.5 10 11.0 13.3 13.2 11.5 13.4 10.7 11.6 (w/w) eters assayed. Storage time also played a signi®cant role: ϫ 18:2

C between 0 and 2 days, no statistically signi®cant effects mg/g 0.598 0.081 0.066 0.066 0.063 0.067 0.469 0.068 were observed, whereas between 0 and 6 days and between 3.202

6 2 and 6 days, only outer pH, moisture, and concentration Ϫ %

10 of C18:3 changed signi®cantly. These observations con®rm 18.4 16.0 14.5 13.9 13.4 14.2 16.6 13.6 (w/w) ϫ 18:1 the hypothesis that chemical modi®cations are only appar- C ent when microorganisms reach high viable numbers. For Downloaded from http://meridian.allenpress.com/jfp/article-pdf/63/2/216/1673220/0362-028x-63_2_216.pdf by guest on 24 September 2021 mg/g 1.163 0.118 0.072 0.070 0.074 0.071 0.726 0.080

2.570 the other combinations of storage times, all chemical pa- 6

Ϫ rameters were signi®cantly affected, except C14:0 content % 8.9 10 10.4 13.3 14.0 12.1 13.4 10.4 11.6 throughout the whole storage period, rupture slope between (w/w) ϫ 18:0 0 and 10 days, between 2 and 10 days, and between 6 and C 10 days, outer pH and moisture between 6 and 10 days, mg/g 0.566 0.076 0.066 0.070 0.066 0.067 0.455 0.069 1.245 and lactic acid content between 10 and 15 days. 6 Ϫ C Њ %

10 CONCLUSIONS 12.1 10.5 13.0 12.6 11.1 12.6 10.8 11.2 (w/w) ϫ 16:0

C The major microbial groups present throughout storage mg/g 0.765 0.077 0.064 0.063 0.061 0.064 0.471 0.066 of whey cheese are Pseudomonas, Bacillus, Enterobacte- 1.367 riaceae, and lactic acid bacteria, which appear as a conse- 5 Ϫ quence of environmental contamination. Packaging under % 8.9 9.4 9.5 9.0 9.7 10 11.7 11.4 11.7 (w/w) vacuum inhibits growth of yeasts, staphylococci, entero- ϫ 14:0

C cocci, and spore-forming clostridia. RequeijaÄo packaged

mg/g under vacuum undergoes relevant acidi®cation, likely de- 0.566 0.069 0.058 0.057 0.052 0.059 0.396 0.057 5.848 rived from lactose metabolism, which may create a positive 6 Ϫ organoleptic effect, whereas unpackaged RequeijaÄo dis- % 8.8 7.7 9.6 9.3 8.4 9.3 8.7 8.3 10 (w/w) plays a signi®cant loss of moisture and a concomitant in- ϫ 12:0

C crease in rigidity. Unpackaged RequeijaÄo develops high

mg/g levels of FFAs after several days of refrigerated storage; 0.556 0.057 0.047 0.046 0.046 0.047 0.382 0.049 5.893 vacuum inhibits lipolysis in RequeijaÄo, despite the high mi- 5 Ϫ crobial counts observed. This is an advantage in terms of % 7.8 8.5 8.2 7.7 8.5 9.5 8.2 10 10.2 (w/w) good sensory quality (obtained by avoiding excessively ϫ 10:0

C rancid and soapy ¯avors). mg/g 0.648 0.057 0.042 0.041 0.042 0.043 0.415 0.049 1.200 ACKNOWLEDGMENTS 5 Ϫ Funding for author M. E. Pintado was provided by a Ph.D. fellow- % 6.3 5.9 6.6 6.5 6.0 6.5 6.3 6.2 10 (w/w) ship issued within the framework of CIENCIA (BD-2526/93-IF), admin- 8:0 ϫ

C istered by Junta Nacional de InvestigacËaÄo CientõÂ®ca e TecnoloÂgica, Por- tugal. Partial funding for this project was obtained from a research grant, mg/g 0.400 0.043 0.033 0.032 0.033 0.033 0.274 0.037 1.302 issued within the framework of PRAXIS, entitled RecuperacËaÄo de pro- teõÂnas e producËaÄo de goma alimentar a partir de soro laÂcteo (PBICT/C/ 5 Ϫ BIO991/95), administered by FundacËaÄo para a CieÃncia e a Tecnologia, % 5.1 4.4 5.2 5.1 5.4 5.1 4.9 4.8 10

(w/w) Portugal, and coordinated by author F. X. Malcata. The authors thank Ms. 6:0 ϫ

C FaÂtima PocËas and Ms. Rita Quadrado for technical support. mg/g 0.325 0.032 0.026 0.026 0.030 0.026 0.214 0.028 2.776 REFERENCES 5

Ϫ 1. BalcaÄo, V. M., and F. X. Malcata. 1998. Intersteri®cation and aci- % 10 4.2 7.4 4.3 5.7 3.8 5.3 3.7 4.2 dolysis of butterfat with oleic acid by Mucor javanicus lipase: chang- (w/w) 4:0 ϫ es in the pool of fatty acids residues. Enzyme Microb. Technol. 22: C 511±519. mg/g 0.268 0.055 0.021 0.029 0.021 0.027 0.163 0.025 2. Bissonnette, N., R. A. Lachance, J. Goulet, M. Landgraf, and C. E. Park. 1980. Evidence of thermonuclease production by Bacillus spp. and enterococci in naturally contaminated cheese. Can. J. Microbiol. Experimental data obtained for concentration of various free fatty acids in whey cheeses throughout storage at 4 26:722±725. Vacuum Vacuum Vacuum Vacuum 3. Brennan, J. C., R. Jowitt, and A. Williams. 1975. An analysis of the action of the general foods texturometer. J. Texture Stud. 6:83±100. 02 Without 0.024 Without 4.9 0.026 5.1 0.033 6.6 0.042 8.4 0.043 8.7 0.056 11.3 0.065 13.0 0.070 14.0 0.070 14.1 0.066 13.3 0.000 0.0 6 Without 12 Without 10 Without 4. Busta, F. F., E. H. Peterson, D. M. Adams, and M. G. Johnson. 1984. of the mean 2.149 Storage Standard error TABLE 2. time (d) Package Colony count methods, p. 62. In M. L. Speck (ed.), Compendium J. Food Prot., Vol. 63, No. 2 VACUUM-PACKAGED WHEY CHEESE 221

of methods for the microbiological examination of foods. American and P. Gaya. 1986. The effect of ripening and cooking temperature Public Health Association, Washington, D.C. on proteolysis and lipolysis in Manchego cheese. Food Chem. 21: 5. Cosentino, S., O. Matza, M. E. Fadda, and F. Palmas. 1989. QualitaÂ 115±123. microbiologica di cagli e innesti utilizzati nella produzione di for- 20. Oliemann, C. 1988. Bepaling van links en rechtsdraaiend melkzuur maggio di pecora. Ann. Igiene 1:1521±1528. in gefermentede zuivel-produkten met behulp van HPLC. Voedings- 6. Fadda, M. E., F. Palmas, S. Cosentino, and R. Cagiano. 1989. Eval- middelentechnologie 21:18±19. uation of microbial contamination of dairy products. Hyg. Moderna 21. Ottogalli, G., G. Rondinini, and C. Cappelleti. 1981. Some micro- 92:408±417. biological and hygienic aspects of Ricotta cheese. Ann. Microbiol. 7. Fedio, W. M., A. Macleod, and L. Ozimek. 1994. The effect of Enzymol. 31:77±78. modi®ed atmosphere packaging on the growth of microorganisms in 22. Panzer, P. C., E. F. Shoppet, H. I. Sinnamon, and N. C. Aceto. 1976. cottage cheese. Milchwissenschaft 49:622±628. Continuous coagulation of cottage cheese whey protein. J. Food Sci. 8. Fedio, W. M., L. Ozimek, and F. H. Wolfe. 1994. Gas production 41:1293±1296. during storage of Swiss cheese. Milchwissenschaft 49:3±7. 23. Pereira, A. J. G., M. E. B. PoÂvoa, and G. R. Cruz. 1982. In¯ueÃncia 9. Ferraz, A. 1998. AdaptacËaÄo ambiental no sector dos lacticõÂnios. AIP da velocidade de aquecimento sobre a qualidade do Ricotta. VII Ambiente 22:22±29. Congresso Nacional de LacticõÂnios, Juiz de Fora, Brazil.

10. Garcia, H. S., H. R. Reyes, F. X. Malcata, C. G. Hill, and C. H. 24. Pintado, M. E., J. A. Lopes da Silva, and F. X. Malcata. 1996. Char- Downloaded from http://meridian.allenpress.com/jfp/article-pdf/63/2/216/1673220/0362-028x-63_2_216.pdf by guest on 24 September 2021 Amundson. 1990. Determination of the major free fatty acids in acterization of RequeijaÄo and technological optimization of its man- milkfat using a three component mobile phase for HPLC analysis. ufacturing process. J. Food Eng. 30:363±376. Milchwissenschaft 45:757±759. 25. Roseiro, M. L. B., and R. A. Wilbey. 1991. The effect of whey 11. Kalogridou-Vassiliadou, D., N. Tzanetakis, and E. Litopoulou-Tza- storage on the production of whey cheese. Sheep Dairy News 3:25± netaki. 1994. Microbiological and physicochemical characteristics of 44. ``Anthotyro,'' a Greek traditional whey cheese. Food Microbiol. 11: 26. Rosenthal, I., B. Rosen, S. Bernstein, and G. Popel. 1991. Preser- 15±19. vation of fresh cheeses in a CO2-enriched atmosphere. Milchwissen- 12. Mathur, B. N., and K. M. Shahani. 1981. Ricotta cheese could be schaft 46:706±708. your best vehicle for whey. Dairy Field 164:110±114. 27. Santiago, L. 1993. O requeijaÄo em Portugal. Via LaÂctea 2:71±73. 13. Miles, A. A., and S. S. Misra. 1938. The estimation of the bacteri- 28. Shapton, D. A., and R. G. Board. 1971. Isolation of anaerobes. Ac- cidal power of the blood. J. Hyg. 45:41±45. ademic Press, New York. 14. Mills, O. 1986. Sheep dairying in BritainÐa future industry. J. Soc. 29. Vodret, A. 1970. La Ricotta pecorina sarda. Sci. Technol. Latt.-Cas. Dairy Technol. 39:88±90. 21:310±313. 15. Modler, H. W. 1988. Development of a continuous process for the 30. Weatherup, W. 1986. The effect of processing variables on the yield production of Ricotta cheese. J. Dairy Sci. 71:2003±2009. and quality of Ricotta cheese. Dairy Ind. Int. 51:41±45. 16. Modler, H. W., and D. B. Emmons. 1989. Production and yield of 31. Wingerd, W. H., S. Saperstein, and L. Lutwak. 1970. Bland, soluble, whole-milk Ricotta manufacture by a continuous process. II. Results whey protein concentrate has excellent nutritional properties. Food and discussion. Milchwissenschaft 44:753±756. Technol. 24:758, 760±761, 764. 17. Modler, H. W., and D. B. Emmons. 1989. Properties of whey protein 32. Ziino, M., F. Salvo, I. Stagno-d'Alcontres, and B. Chiofalo. 1993. concentrate prepared by heating under acidic conditions. J. Dairy Composition of Ricotta cheese from Pinzirita sheep bred in the areas Sci. 60:177±184. surrounding Mt. Etna volcano (Catania-Italy). Sci. Technol. Latt.- 18. NaÂgera, A. I., L. J. R. BarroÂn, and Y. Barcina. 1993. Lipid fraction Cas. 44:217±231. composition of cow's, sheep's, and goat's cheese, and the in¯uence 33. Zoon, P. 1991. The relation between instrumental and sensory eval- on its quality. Rev. Esp. Sci. Technol. Aliment. 33:345±363. uation of the rheological and fracture properties of cheese. Bull. Int. 19. Nunez, M., C. Garcia-Aser, M. A. Rodriguez-Martin, M. Medina, Dairy Fed. 268:30±35.