The lower amounts of intracellular and extracellular peptidoglycan
detected could be due to fragments of peptidoglycan being
released into the medium during cell growth and elongation.
It has been previously stated that peptidoglycan is steadily
broken down by peptidoglycan-cleaving enzymes during
cell growth, especially during the exponential phase, where
a cell wall turnover process occurs, resulting in the release
of peptidoglycan fragments from the wall (Reith et al.
2011). It has also been reported that the turnover rate and
the amount of cell wall material vary among species and
even strains. Reith et al. (2011) reported that the magnitude
of cell wall turnover is also dependent on the growth conditions. In addition, the capability of microorganisms in the
recycling of cell wall material also affected the amount of
peptidoglycan detected in the extracellular and intracellular
extracts (Litzinger et al. 2010). Our current study demonstrated that all strains contain a total peptidoglycan amount
ranging from 0.241 to 0.425 μg/mL. Although larger
amounts of peptidoglycan, in the range of 10–100 μg/mL
are necessary to stimulate cellular responses, peptidoglycan
has also been reported to be effective at lower concentrations, via synergism with LTA (Yang et al. 2001). It is
postulated that peptidoglycan at concentrations as low as
0.24 μg/mL in the presence of LTA is sufficient to induce
production of antimicrobial peptides in keratinocytes.
HA is synthesized intracellularly according to the proposed biosynthetic pathway by Matsubara et al. (1991) and
transported out across the cellular membrane upon synthesis. Thus, the varying concentrations of HA in the intracellular and extracellular extracts among strains studied would
be dependent on the efficiency of transport across the membrane. In addition, it has been suggested that biosynthesis of
HA was dependent on the glucose uptake of the microorganism (Cooney et al. 2008). Our current study showed that
lactobacilli were able to produce higher amounts of HA.
Parche et al. (2006) revealed that, unlike many other bacteria, glucose utilization of some bifidobacteria was impaired
in the presence of glucose and lactose such as in milk, and
thus may have resulted in the lower production of HA. In
our current study, strains of lactobacilli and bifidobacteria
were able to produce HA at concentrations ranging from 0.4
to 1.4 mg/mL. Hyaluronic acid has been used as an ideal
bio-component in dermatological and pharmaceutical products, mainly due to its remarkable rheological, hygroscopic
and viscoelastic properties which are relevant for dermal
tissue function. It is worth noting that, in this current
study, L. rhamnosus FTDC 8313 and L. gasseri FTDC 8131
produced HA at concentrations of more than 1 mg/mL.
Kobayashi and Terao (1997) reported that HA at concentration of 1 mg/mL enhanced the release of interleukin-1β,
which is responsible for the secretion of RNase 7, which is
an antimicrobial peptide that enhances the innate immune
defense system of keratinocytes.
Citrate utilization and production of diacetyl among lactobacilli has been extensively studied over the past few
decades, and these studies showed that homofermentative
species produced diacetyl more readily and in larger volumes than the heterofermenters (Christensen and Pederson
1958). Interestingly, our results showed that the heterofermenter, L. casei BT 1268 strains produced higher amounts
of diacetyl than the homofermenter, L. gasseri FTDC 8131.
Table 5 Acid and neutral sphingomyelinases activity in extracellular
and intracellular extracts of bifidobacteria and lactobacilli cultured in
8 % (w/v) reconstituted skimmed milk (RSM) for 20 h at 37 °C
Strains Sphingomyelinase activity
(mU/mL of RSM)
Intracellular Extracellular
Acid sphingomyelinase
L. casei BT 1268 0.22±0.02 aB 0.55±0.08 bA
B. animalis subsp. lactis BB 12 0.19±0.03 aB 0.50±0.13 bA
B. longum BL 8643 0.25±0.09 aB 0.48±0.11 bA
L. rhamnosus FTDC 8313 0.28±0.06 aB 0.64±0.15 bA
L. gasseri FTDC 8131 0.27±0.04 aB 1.25±0.17 aA
Neutral sphingomyelinase
L. casei BT 1268 1.16±0.05 aB 2.40±0.63 aA
B. animalis subsp. lactis BB 12 1.29±0.40 aA 0.66±0.12 bB
B. longum BL 8643 0.92±0.17 aA 0.94±0.21 bA
L. rhamnosus FTDC 8313 0.35±0.14 bB 1.10±0.12 bA
L. gasseri FTDC 8131 1.31±0.25 aB 2.57±0.61 aA
Results are expressed as mean±standard deviation. Values are means
of duplicates from three separate runs (n03)
Means in the same column with different lowercase letters are significantly different (P