Abstracts. H. Gaudreau 02.

Scientific Publications - Work Done by Microbiology Reader Bioscreen C

Can. J. Microbiol./Rev. Can. Microbiol. 45(11): 891-897 (1999)

Effect of ultrafiltration of yeast extracts on their ability to promote lactic acid bacteria growth

H. Gaudreau, C.P. Champagne, J. Conway, and R. Degré

ABSTRACT

Five yeast extracts (YE) were fractionated by ultrafiltration (UF) with 1, 3, and 10 kDa molecular weight cutoff membranes, concentrated by freeze-drying, and the resulting powders of yeast extract filtrates (YEF) were evaluated for their growth-promoting properties on nine cultures of lactic acid bacteria (LAB). There was an increase in a-amino nitrogen content of the YEF powders as the pore size of the UF membranes used to filter the YE solutions decreased. The source of YE had a much greater effect than UF on the growth of LAB. This was also the case for the YEF contents in total and a-amino nitrogen. Growth curves of the LAB showed that maximum growth rate (m_max) data were on average 30% higher with bakers' YE than with brewers' YE, while maximum optical density (OD_max) values were on average 16% higher with bakers' YE. This could be related to the higher nitrogen content of the bakers' YE used in this study. Modification by UF of the YE had no significant influence on the growth of 4 of the 9 LAB strains. The three strains of Lactobacillus casei were negatively influenced by UF, as they did not grow as well in the media containing the YEF obtained after filtering with 1 and 3 kDa membranes. On a total solids basis, the 2.5× retentates from the 10 kDa membrane gave, on average, 4% lower m_max and 5% lower OD_max values as compared to cultures where the corresponding YEF was used as medium supplement. This could also be partially related to the different nitrogen contents of the filtrates and retentates.

Key words: Lactococcus, Pediococcus, Lactobacillus, amino acids.

Introduction

Yeast extracts (YE) are the water soluble portion of autolysed yeast cells. The beneficial effect of YE on the growth of lactic acid bacteria (LAB) is well established (Smith et al. 1975; Aeschlimann and von Stockar 1990). YE are an excellent source of the B-complex vitamins and are often used to supply these factors in bacteriological culture media (Difco 1984). They are also a reliable source of peptides and amino acids (Peppler 1982).

Amino acids and peptides stimulate the growth of LAB (van Boven and Konings 1986; Hemme et al. 1981; Juillard et al. 1995) and they have been shown to be partially responsible for YE stimulation of the growth of lactic cultures (Benthin and Villadsen 1996). Peptides can be superior to free amino acids as a nitrogen sources for LAB because they

can provide the cell with amino acids in a form that can be utilized more efficiently (van Boven and Konings 1986).

Some commercial applications, microbiological media for example, require that YE produce clear solutions. One way of obtaining clear solutions is by filtering the YE. There is little information of the effect of filtration on the growthpromoting abilities of YE. Desmazeaud and Hermier (1972) separated peptide fractions by gel filtration of digested casein and found that peptides with molecular weights in the range 1–2.5 kDa stimulated growth of Streptococcus thermophilus. Fractions of casein hydrolysates obtained by ultrafiltration (UF) have been used to supplement culture medium and to standardize milk cultures (St-Gelais et al. 1993). These results suggest that UF fractions of YE may have different growth-promoting abilities, but no information is available on the effect of UF on the biological value of YE with respect to LAB.

The aim of this study was to examine if UF of YE influences their growth-promoting properties in various lactic acid cultures, and to provide a comprehensive view of the effect of ultrafiltration on the biological value of YE on lactic cultures. However, using only one strain and one YE product would not have enabled this. The literature shows that there is variability in both YE (Potvin et al. 1997) and lactic culture responses to growth-promoting compounds (St-Gelais et al. 1993; Valdez et al. 1985a). Therefore, in order to provide a comprehensive picture, five YE were studied with nine strains of lactic cultures. Furthermore, two lots of each YE were used in order to reduce the effect of the YE lot.

Materials and methods

Commercial yeast extracts

Five commercial YE were obtained, and their suppliers are listed in Table 1. Since variability between lots has been reported (Potvin et al. 1997), two lots of each source were used. With each lot, two assays were carried out, for a total of 4 separate assays. Three YE were from bakers’ yeasts (A, B, E) while two were from brewers’ yeasts (C, D).

Ultrafiltration of yeast extracts

YE were suspended in deionized water to obtain a solution containing 10% (w/v) solids. This solution was pre-filtered on an 8 µm Whatman (No. 2 filter paper) membrane. The filtrate was further processed by UF, using a tangential filtration system (Minitan Filter plates, Millipore, Bedford, U.S.A.), with membranes having molecular cutoffs of 10, 3, or 1 kDa. The system enabled the use of four 30 cm² membranes for a total filtration surface of 120 cm². A 500 mL solution of 10% YE was used, and the UF process was stopped after the recovery of 300 mL of filtrate. The yeast extracts filtrates (YEF) were concentrated by lyophilization in a LyoTech (Lyo San Inc., Lachute, Canada) at 24°C for 72 h and stored at – 20°C until used. Total and a-amino nitrogen content, total solids, turbidity, and growth-promoting properties of each YE and YEF were evaluated. All chemical analysis was done in duplicate for each lot tested and the results presented are the means of the two values obtained.

Chemical analyses of YE and YEF

a -Amino nitrogen

The a-amino nitrogen of the fractions was determined by titration following reaction with formaldehyde (USP 1985). The 5% YEF solutions were adjusted at pH 7.0 with 0.1 M NaOH or 0.1 M HCl.

Total nitrogen determination

The total nitrogen determination was done using a FP-428 LECO apparatus (LECO Corporation, Saint Joseph, Mich.), operated under the following conditions: sample size, 150 mg; oxidation furnace temperature, 900°C; oxidation standby temperature, 650°C; purge cycles, 3; minimum timeout, 30 s; comparator level, 1.00; loop select low range, flow constants at high; gases, oxygen 99.99% and helium 99.99%. The calibration standard was composed of 150 mg EDTA (No. 502–092, 9.56 ± 0.03% nitrogen, LECO Corporation, Saint Joseph, Mich.)

Turbidity

Turbidity of YE and YEF solutions were determined with a Orbico-Hellige turbidimeter (Model 965; Farmingdale USA). Hydrazine sulfate standards (VWR; West Chester Pa., USA) were used to calibrate the turbidimeter.

Water content

Water content of the YE, filtrates, and YEF powders were obtained by dry weights after an incubation at 105°C for 16 h.

Organisms and growth conditions

Lactobacillus plantarum EQ27, Lactobacillus casei EQ28, EQ85, EQ70, Lactobacillus acidophilus EQ57, Pediococcus acidilactici MA1815-M, and Pediococcus pentosaceus EQ44 were kindly provided by Lallemand-Equipharm SA (Aurillac, France). Lactococcus lactis subsp. cremoris Wg2L, a proteinase negative (Prt^-) culture was graciously provided by D. St-Gelais (Food Research and Development Center (FRDC), St-Hyacinthe, Canada) while Lactococcus lactis subsp. cremoris R2 Prt⁺ culture, was from our own FRDC collection.

The Lactobacillus and Pediococcus strains were grown at 37°C in MRS broth (BDH, Darmstadt, Germany), until their pH reached 4.6, which required approximately 8 h. The cultures were put on ice for 30 min to stop the acidification process. The lactococci strains were obtained following a 16h/22°C incubation in reconstituted skim milk (11% solids). The milk was sterilized at 110°C for 10 min.

Mother cultures were prepared by mixing 20 mL of cell suspensions, freshly grown on MRS or milk, with 50 mL of 20% skim milk and 50 mL of a 20% glycerol solution (glycerol and milk were sterilized separately). The milk/glycerol cell suspensions were divided into 1 mL fractions which were added to sterile cryovials (Nalgene, Rochester, N.Y.) and stored at –70°C until used.

Media

For the testing of the various YE and YEF in spectrophotometric assays, the base medium used for the growth of the lactobacilli and pediococci had the following composition, per L of medium: Glucose, 20 g; K2HPO4, 3 g; KH2PO4, 3 g; sodium citrate, 3 g; Tween 80, 1 g; MgSO₄·7H₂O, 0.2 g; MnSO₄·2H₂O, 0.05 g; and YEF at the set concentrations for the different strains. A glucose and MgSO4·2H2O solution was first prepared with compounds having twice the concentration (40 g/L and 0.4 g/L respectively) and autoclaved 15 min at 121°C. The rest of the medium was also prepared at a double concentration and sterilized the same way. Both solutions were mixed following sterilization.

The media used for the assays with lactococci was the same as for lactobacilli and pediococci, except for glucose which was substituted by lactose. In these assays, it was not required to prepare separate 2× solutions prior to sterilization. The same media were used to test various >10 kDa retentates.

Previous studies have shown that the appropriate concentration of YE in the basal medium, used to evaluate the biological value of various YE, must be defined for each strain (Potvin et al. 1997; Champagne et al. 1999). Preliminary studies were thus carried out using the method of Champagne et al. (1999) to determine the appropriate concentrations for the determination of maximum growth ( max) or maximum optical density _(ODmax).Thus, YE or YEF were added at 5 g/L for lactococci, at 1 g/L for the three L. casei strains and at 2 g/L for the other cultures.

Growth-promoting properties evaluation of YE and YEF

An automated turbidimetric instrument (Bioscreen C, Labsystems, Corp., Helsinki, Finland) was used to assess the effect of YE or YEF added to the base medium on the increase in turbidity of the cultures. The method described by Champagne et al. (1999) was used.

Sterile media having various YE and YEF were prepared in test tubes, inoculated at 1% (v/v) from a fresh culture, and 350- L samples were added aseptically to each microwell. Microplates were incubated for 24 h at 37°C for the lactobacilli as well as the pediococci, and at 22°C for the lactococci. The absorbency at 600 nm of each microwell was automatically recorded every 15 min. Prior to reading, the plates were shaken at low intensity for 5 s. Two replicates for each fractions were evaluated on Bioscreen for both YE lots tested. Thus, four separate assays were carried out.

Statistical analyses

Statistical analyses were performed using SAS Institute, Inc. (SAS 1989, Cary, N.C.) software. Analysis of variance was used with significance defined at P < 0.05. The biomass values in the spectrophotometric assay system were transformed into Ln values of biomass and plotted against time for the exponential growth part of the curve (Sigma Plot software, Jandel). The slope of the curve, giving the _µmax,was obtained by simple linear regression. The term fermentation time is employed to represent the time corresponding to the end of the exponential phase of the growth curve of the strains.

Results and discussion

Nitrogen content of the YE or YEF

The total nitrogen (total N) and a-amino nitrogen ( - amino N) contents of the original and filtered YE powders are shown in Table 1. The variance analysis suggests that the effect of filtration was not significant with respect to nitrogen contents. Paired T tests show, however, that the total N contents of the retentates and the YEF 10 kDa products were indeed significantly (P < 0.05) different from the original YE. As well, regression analyses show that the systematic increases in a-amino N in the YEF powders as filter poresize decreased were real. The data shows nevertheless that the effect of the YE source was much greater than that of UF on total and a-amino N contents.

Brewers’ YE had sightly lower total N content than bakers’ YE. In the original YE, the proportion of a-amino N in total N was approximately 50%, which suggests that there was an important hydrolysis of protein and peptides during the autolysis process. The source of YE affected the content of a-amino N of YE (Table 1).

The a-amino N content of the YEF powders increased as the molecular weight cutoffs of the UF membranes, used to filter the YE solutions, decreased (Table 1). On average,

YEF powders obtained from the 1 kDa unit contained 30% more a-amino N than in the original YE. This was expected, as filtration removed the high molecular weight compounds and increased the proportional content of the remaining peptides and amino acids. The proportion of a-amino N to total N becomes more important as the pore size of the filtration membranes are reduced, which suggests that most of the nitrogen is probably composed of peptides and amino acids. These results are in accordance with technical data given by Ohly (1998), which cites the protein-related fraction of standard salt-free YE powder to contain 35–40% amino acids, 40–60% short peptides and 2–5% oligopeptides/proteins having MW greater than 3 kDa.

Table 1. Nitrogen composition of yeast extracts and yeast extracts fractions.


		Total nitrogen^b	a-amino nitrogen^b
Yeast^a	Filtration	(g/100 g)	(g/100 g)
A	None	10.9 cdef	5.3 cde
(Bakers’)	Retentate^c	11.0 cde	4.8 ef
	10 kDa	11.6 bc	5.9 cde
	3 kDa	11.0 cde	6.7 abcd
B	1 kDa None	10.9 cdef 11.9 bc	7.1 abcd 6.7 abcd
(Bakers’)	Retentate	12.0 bc	6.2 cde
	10 kDa	13.4 a	7.4 abc
	3 kDa	12.9 ab	8.4 ab
	1 kDa	12.7 ab	8.8 a
C	None	7.7 gh	4.0 ef
(Brewers’)	Retentate	7.1 h	3.4 f
	10 kDa	9.5 f	5.1 de
	3 kDa	9.7 ef	5.7 cde
	1 kDa	9.8 ef	6.4 bcd
D	None	9.8 def	6.4 bcd
(Brewers’)	Retentate	9.6 ef	5.9 cde
	10 kDa	11.0 cde	7.1 abcd
E	3 kDa 1 kDa None	10.9 cdef 10.9 cdef 10.8 cdef	7.2 abcd 7.5 abc 6.1 cde
(Bakers’)	Retentate	10.3 cdef	5.2 de
	10 kDa	11.6 bc	6.0 cde
	3 kDa	11.3 cd	6.7 abcd
	1 kDa	11.1 cde	7.2 abcd

^aSupplied by Bio Springer (Maisons-Alfort, France); Difco (Detroit, Mich., U.S.A.); Lallemand (Montreal, Que., Canada); and Red Star (Juneau, U.S.A.). The products were coded A to E so as to prevent any prejudice to the companies.

^bFor a given column, means that are followed by the same letter (a, b, c, d, e , f) are not significantly different (P > 0.05). ^cRetentate: UF retentate (2.5×) of the 10 kDa filter.

Overall effects of YE source and UF on the growth of LAB

Three characteristics of the LAB spectrophotometry growth curves were examined: _µmax,OD_max, and the fermentation time. Jensen and Hammer (1993) have shown that a µ of 0.3 h^-1is obtained with Lc. lactis when 8 amino acids are added in the growth medium, but increases to 0.7 h^-1when 19 amino acids are included. This suggests that the _µmaxdata provide an indication of the number of essential and nonessential growth factors present in the YE; OD_maxwould in-

In all but two instances, statistical analyses showed that at least one experimental factor had a significant influence (Table 2). In all cases where a global effect was detected, the source of the YE had a significant impact. Thus, the source of YE had a much greater effect than UF on the growth of LAB. This was also the case with respect to contents in total and a-amino N.

It is noteworthy that there was no interaction between the effects of UF and YE source (Table 2). Therefore, the effect of fractionation of YE on the growth of LAB was deemed constant and did not vary significantly from one YE to another.

Although it was assumed that high correlations would be obtained between _µmaxand fermentation time, regression analyses did not show this to be systematic. With some strains, it was very high (R² > 0.95 for L. cremoris R2 and L. casei EQ28), while for others it was very poor (R² < 0.08 for L. casei EQ85 and EQ70). The same can be said for the relationships between _µmaxand OD_max, as well as for ^ODmax and fermentation time (data not shown).

Table 2. Statistical analyses results of ^ODmax, µmax and fermentation time data obtained by automated spectrophotometry for various lactic acid bacteria.

					Strains
	Wg2L	R2	EQ28	EQ85	EQ70	EQ27	EQ57	EQ44	Ma 18/5M
^ODmax data Global effect	0.0001**	0.0001**	0.0001**	0.0001**	0.0001**	0.0001**	0.0001**	0.06	0.0001**
YE effect	0.0001**	0.0002**	0.0001**	0.0001**	0.0001**	0.0001**	0.0001**	ND	0.0001**
Filtration effect	0.87	0.489	0.0001**	0.0001**	0.49	0.812	0.137	ND	0.0001**
YE * Filtration interaction	0.933	0.973	0.791	0.705	0.248	0.261	0.726	ND	0.015*
µmax data Global effect	0.0001**	0.0001**	0.0010**	0.0007**	0.0005**	0.0001**	0.0001**	0.0001**	0.0001**
YE effect	0.0001**	0.0001**	0.0004**	0.0001**	0.0001**	0.0001**	0.0001**	0.0001**	0.0001**
Filtration effect	0.447	0.0006**	0.0001**	0.0101*	0.0001**	0.107	0.103	0.12	0.0003**
YE * Filtration interaction	0.164	0.112	0.13	0.508	0.254	0.151	0.455	0.0017**	0.12
Fermentation time data Global effect	0.0001**	0.0001**	0.0002**	0.0001**	0.0001**	0.0001**	0.0001**	0.0001**	0.611
YE effect	0.0001**	0.0001**	0.0001**	0.0001**	0.0001**	0.0001**	0.0001**	0.0001**	ND
Filtration effect	0.312	0.0257*	0.0171*	0.519	0.963	0.14	0.0014**	0.0248	ND
YE * Filtration interaction	0.15	0.0327*	0.491	0.144	0.105	0.617	0.701	0.0012**	ND

Note: **, Highly significant effect (P < 0.01); * Significant effect (P < 0.05). ND = not determined.

Effect of YE source on the growth of LAB

There were wide variations in the responses of the individual strain to the native YE (Table 3). This is undoubtedly related to the fact that LAB have variable requirements in growth factors (Desmazeaud and de Roissard 1994; Hugenholtz et al. 1987). Lactococci can stop growing after depletion of essential free amino acids and peptides (Thomas and Mills 1981; Thomas and Pritchard 1987). The LAB are notorious for their variable responses to growth and drying conditions (Valdez et al. 1983, 1985a, and 1985b), and this study adds to the literature in this respect. Nevertheless, the use of 9 strains and 5 YE in the experimental plan has the advantage of enabling the detection of trends. One noticeable trend was that the LAB used in this study had a

slight preference for bakers’ YE, as _gmaxdata were on average 30% higher with bakers’ YE than with brewers’ YE, while _ODmaxvalues were on average 16% higher with bakers’ YE. It is noteworthy that the brewers’ YE used in this study contained lower nitrogen levels. This suggests another trend, in that growth responses of LAB might be partially related to nitrogen contents of the YE. These data would thus appear to be in line with the results of Smith et al. (1975), who showed that amino acid material was the best overall constituent inYE for stimulating bacterial growth.

The growth rates obtained for Lactobacillus acidophilus (0.13 to 0.25 h^-1) agree with results of Liu and Moon (1982), but they are inferior to those reported in the literature with the lactococci (Bibal et al. 1988; Jensen and Hammer 1993). This could be explained by the YE concentration employed in this experimental procedure, which was low. It must be emphasized that the methodology used in this study was designed to put in evidence differences in the growthpromoting properties of YE (Champagne et al. 1999), and not to obtain the highest possible growth rates.

Effect of YE UF on the growth of LAB

Variance analyses reveal that YE UF had no significant (P > 0.05) influence on the growth of Lc. cremoris Wg2L, Lb. plantarum EQ27, Lb. acidophilus EQ57, and P. pentosaceus EQ44. For these strains, either UF does not generate differences in YEF composition important enough to significantly influence growth, or the cells can adapt to the varying proportions of peptide size in the YEF. Indeed, the lactococci have multiple permease systems designed for the specific assimilation of amino acids, dipeptides/tripeptides, and oligopeptides (Juillard et al. 1996). Thus, they could easily adapt to the varying content of the YEF and not be influenced by the limited modifications in peptide lengths found in the YEF products.

On the other hand, growth properties of Lb. casei and P. acidilactici Ma 18/5M, were influenced by UF (Table 2). In addition to P. acidilactici Ma 18/5M, it is noteworthy that the growth of all three strains of Lactobacillus casei was

significantly lower in the YEF obtained following UF with 1 and 3 kDa membranes. Only one strain, Lc. cremoris R2, had growth rates significantly higher (P < 0.01) in media enriched with YEF in comparison with the entire YE. A certain number of conclusions can be reached with these results. First, this shows again the strain/species variability in responses of LAB to media composition. Secondly, the increase in low molecular weight nitrogen compounds, obtained in YEF with decreasing pore size membranes (Table 1), is not a critical factor in YE growth stimulation of Lb. casei. These results, and the reports of numerous peptidase activities in Lb. casei (Arora and Lee 1994; Habibi-Najafi and Lee 1994), suggest that these lactobacilli have a preference for peptides as a nitrogen source in their growth medium. Van Boven and Konings (1988) have demonstrated that some bacteria have preferences for peptides over free amino acids, since direct transport of peptides into the cell prior to hydrolysis can reduce the amount of metabolic energy used for amino acid uptake.

Table 3. Effect of YE sources on the growth of various lactic acid bacteria.

				Yeast extract
Strain	Parameters	A	B	C	D	E
Lactococcus lactis subsp. cremoris Wg2L	µmax	0.123 a	0.108 a	0.024 b	0.010 b	0.112 a
	^ODmax	0.857 a	0.949 a	0.588 b	0.347 b	0.910 a
	Fermentation time	18.0 b	22.5 a	23.5 a	N.D.	19.3 b
Lactococcus lactis subsp. cremoris R2	µmax	0.137 d	0.210 a	0.181 c	0.200 b	0.133 d
	^ODmax	0.991 b	1.094 a	1.063 a	1.100 a	0.987 b
	Fermentation time	17.4 a	14.0 b	14.5 b	13.3 c	16.8 a
Lactobacillus casei EQ28	µmax	0.217 a	0.240 a	0.233 a	0.238 a	0.225 a
	^ODmax	1.022 a	0.636 b	0.720 ab	0.700 b	0.932 ab
	Fermentation time	12.6 a	6.1 d	8.6 c	7.1 cd	10.4 b
Lactobacillus casei EQ85	µmax	0.264 b	0.321 a	0.274 b	0.288 b	0.277 b
	^ODmax	1.164 a	1.203 a	0.868 b	0.990 b	1.147 a
	Fermentation time	11.1 a	9.8 b	8.8 c	9.5 c	10.5 ab
Lactobacillus casei EQ70	µmax	0.171 b	0.194 a	0.171 b	0.183 ab	0.175 b
	^ODmax	1.050 b	1.222 a	0.898 c	1.162 a	1.145 a
	Fermentation time	18.0 b	17.0 b	15.8 c	18.5 b	19.3 a
Lactobacillus plantarum EQ27	µmax	0.305 b	0.390 a	0.297 b	0.358 a	0.292 b
	^ODmax	1.109 ab	1.233 a	0.845 d	1.048 c	1.077 bc
	Fermentation time	10.5 a	9.0 b	8.8 c	7.8 d	9.8 a
Lactobacillus acidophilus EQ57	µmax	0.234 a	0.129 b	0.250 a	0.153 b	0.184 ab
	^ODmax	0.670 ab	0.327 c	0.649 ab	0.468 c	0.582 bc
	Fermentation time	8.3 a	5.5 c	7.5 a	7.0 b	7.8 a
Pediococcus acidilactici MA 18/5M	µmax	0.080 bc	0.069 bc	0.054 c	0.128 ab	0.098 bc
	^ODmax	0.248 bc	0.237 c	0.245 bc	0.294 ab	0.304 a
	Fermentation time	6.56 a	5.4 b	5.3 b	5.3 b	6.4 a
Pediococcus pentosaceus EQ44	µmax	0.259 c	0.390 ab	0.342 b	0.380 ab	0.253 c
	^ODmax	0.717 c	1.046 a	0.849 b	1.048 a	0.650 c
	Fermentation time	10.0 a	10.0 a	9.3 b	10.0 a	9.5 b

Note: For each row, means followed by the same letter (a, b, c, d) are not significantly different (P > 0.05).

Growth of LAB in media supplemented with retentates and filtrates

In most cases, there was no advantage to carrying out YE UF with membranes of 1 and 3 kDa. Therefore, industrial UF of YE to obtain products that give clear solutions would preferably be produced with 10 kDa membranes. From this filtration process, a by-product (a concentration of compounds with MW over 10 kDa) is produced. It was examined whether the retentates obtained could still be used for industrial microbiological media that do not require clear solutions.

On a total solids basis, the 2.5× retentates from the 10 kDa membrane gave, on average, total N contents 12% lower than in the filtrate of the five YE tested (Table 1). There was a systematic decrease in a-amino N, which was on average 20%. This resulted in slightly lower _µmaxvalues in cultures grown with the retentates as medium supplement (Table 4). Paired t tests revealed that _µmaxdata obtained with the retentates were on the average 4% lower than with the filtrates, and that this difference was significant. The same was obtained with the OD_max data, which were 5% lower with the retentates as compared to the filtrates (data not shown).

In conclusion, the 2.5× retentates of the 10 kDa UF membrane do not have the same growth-promoting abilities on LAB as do the filtrates, on an equal solids basis, but the difference is not major. Therefore, retentates could still prove useful in some microbiological media.

Biological value of YE between strains

The variable growth response of the strains with respect to the YE was noted earlier. It was nevertheless examined whether trends could be found. As can be seen in Tables 5 and 6, correlations between strains were not high. No clear

Table 4. Maximum specific growth rates ( max) obtained by spectrophotometric analyses for various lactic cultures in basal media containing 2.5× retentates or filtrates of a YE ultrafiltration process with a membrane cutoff of 10 kDa.

					Yeast extract
		A		B		C		D		E
Strain	Filtrate	Retentate	Filtrate	Retentate	Filtrate	Retentate	Filtrate	Retentate	Filtrate	Retentate
Lc. cremoris Wg2L	0.132	0.125	0.101	0.119	0.025	0.066	0.01	0.02	0.122	0.119
Lc. cremoris R2	0.148	0.138	0.217	0.211	0.189	0.178	0.212	0.206	0.142	0.136
Lb. casei EQ28	0.215	0.207	0.251	0.237	0.233	0.221	0.219	0.223	0.218	0.207
Lb. casei EQ85	0.267	0.259	0.312	0.314	0.287	0.272	0.285	0.281	0.29	0.234
Lb. casei EQ70	0.156	0.158	0.185	0.189	0.17	0.157	0.164	0.168	0.149	0.161
Lb. plantarum EQ27	0.307	0.292	0.378	0.37	0.308	0.271	0.359	0.342	0.288	0.279
Lb. acidophilus EQ57	0.211	0.209	0.061	0.082	0.205	0.2	0.132	0.132	0.203	0.181
P. acidilactici MA 18/5M	0.104	0.165	0.107	0.103	0.129	0.092	0.192	0.159	0.1	0.096
P. pentosaceus EQ44	0.285	0.247	0.423	0.416	0.374	0.31	0.331	0.316	0.28	0.257

Table 5. Determination coefficients (R²) of maximum optical density (OD_max) data obtained by automated spectrophotometry between different lactic acid strains

different lactic acid strains.
Strain	EQ28	EQ85	EQ70	EQ27	EQ57	Wg2L	R2	Ma 18/5M	EQ44
Lb. casei EQ28	1
Lb. casei EQ85	0.06	1
Lb. casei EQ70	0.02	0.56	1
Lb. plantarum EQ27	0.06	0.59	0.55	1
Lb. acidophilus EQ57	0.36	0.08	0.45	0.39	1
Lc. cremoris Wg2L	0.09	0.59	0.11	0.25	0.01	1
Lc. cremoris R2	0.31	0.01	0.12	0.07	0.6	0.28	1
P. acidilactici Ma 18/5M	<0.01	0.14	0.03	0.04	0.01	0.39	0.1	1
P. pentosaceus EQ44	0.39	0.1	0.05	0.03	0.5	0.37	0.81	0.09	1

Table 6. Determination coefficients (R²) of maximum growth rate ( max) data obtained by automated spectrophotometry between different lactic acid strains.

Strain	EQ28	EQ85	EQ70	EQ27	EQ57	Wg2L	R2	Ma 18/5M	EQ44
Lb. casei EQ28	1
Lb. casei EQ85	0.37	1
Lb. casei EQ70	0.59	0.56	1
Lb. plantarum EQ27	0.07	0.56	0.31	1
Lb. acidophilus EQ57	<0.01	0.37	0.11	0.54	1
Lc. cremoris Wg2L	0.17	0.04	0.08	0.12	<0.01	1
Lc. cremoris R2	0.17	0.53	0.3	0.8	0.38	0.4	1
P. acidilactici Ma 18/5M	<0.01	<0.01	0.01	0.13	0.07	0.42	0.21	1
P. pentosaceus EQ44	0.31	0.59	0.42	0.58	0.19	0.18	0.73	<0.01	1

trends could be observed. Although determination coefficients (R²) between strains of the same species were never below 0.31, they were not necessarily the highest. The limited number of different YE studied might be responsible for these results. In a study involving 26 different commercial YE, Champagne et al. (1999) had found better relationships between _µmaxand _ODmaxof strains of the same species.

Conclusions

The industrial production of LAB is often carried out in media containing YE. This study shows that bakers’ YE seems more appropriate then brewers’ YE, potentially because of their higher nitrogen content, and that UF with 10 kDa membranes may be useful in improving YE nitrogen

contents to that end. The disadvantage of bakers’ YE is that it is more expensive than brewers’ YE. In order to determine the most appropriate product, economically and qualitywise, the variability of responses of LAB to YE make it necessary for producers or users of these cultures to test lots of available YE. This points to the need for cooperation between YE suppliers and users in determining the most suitable lots for their uses.

Acknowledgements

Gratitude is expressed to Melanie Bilodeau and Nancy Gardner for technical assistance, as well as Laurent Bazinet for useful comments on the manuscript.

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