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Susceptibility of Escherichia coli to C2-C18 Fatty Acids

DOI:10.1007/BF02931506
Authors:
Milan Marounek at Institute of Animal Science, Prague
Milan Marounek
Eva Skřivanová at Institute of Animal Science, Prague
Eva Skřivanová
Vojtech Rada at Czech University of Life Sciences Prague
Vojtech Rada
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Abstract and Figures

The antimicrobial activity of C2-C18 fatty acids was determined in vitro in cultures of two strains of Escherichia coli grown on glucose. Antimicrobial activity was expressed as IC50 (a concentration at which only 50% of the initial glucose in the cultures was utilized). Utilization of glucose was inhibited by caprylic acid (IC50 0.30-0.85 g/L) and capric acid (IC50 1.25-2.03 g/L). Neither short-chain fatty acids (C2-C6) nor fatty acids with longer chain (C12-C18) influenced substrate utilization. Caproic acid, however, decreased cell yield in cultures of E. coli in a dose-dependent manner. No inhibition of glucose utilization was produced with unsaturated fatty acids, oleic and linoleic. Calcium ions added in excess reversed the antimicrobial effect of capric acid, but not that of caprylic acid. Antimicrobial activity of caprylic and capric acid decreased when the bacteria were grown in the presence of straw particles, or repeatedly subcultured in a medium containing these compounds at low concentrations. Counts of viable bacteria determined by plating decreased after incubation with caprylic and capric acid (30 min; 1 g/L) at pH 5.2 from > 10(9) to approximately 10(2)/mL. A reduction of a mere 0.94-1.96 log10 CFU was observed at pH 6.5-6.6. It can be concluded that caprylic acid, and to a lesser extent also capric acid, has a significant antimicrobial activity toward E. coli. Effects of other fatty acids were not significant or absent.
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Folia Microbiol. 48 (6), 731–735 (2003) http://www.biomed.cas.cz/mbu/folia/
Susceptibility of Escherichia coli to C
2
– C
18
Fatty Acids
M. MAROUNEK
a,b
*, E. SKŘIVANOVÁ
b
, V. RADA
c
a
Institute of Animal Physiology and Genetics, Academy of Sciences of the Czech Republic, 104 00 Prague-Uhříněves, Czechia
e-mail marounek@iapg.cas.cz
b
Research Institute of Animal Production, 104 01 Prague-Uhříněves, Czechia
c
Department of Microbiology and Biotechnology, Czech University of Agriculture in Prague, 165 21 Prague, Czechia
Received 7 May 2003
Revised version 6 October 2003
ABSTRACT. The antimicrobial activity of C
2
–C
18
fatty acids was determined in vitro in cultures of two
strains of Escherichia coli grown on glucose. Antimicrobial activity was expressed as IC
50
(a concentration
at which only 50 % of the initial glucose in the cultures was utilized). Utilization of glucose was inhibited
by caprylic acid (IC
50
0.30–0.85 g/L) and capric acid (IC
50
1.25–2.03 g/L). Neither short-chain fatty acids
(C
2
–C
6
) nor fatty acids with longer chain (C
12
–C
18
) influenced substrate utilization. Caproic acid, however,
decreased cell yield in cultures of E. coli in a dose-dependent manner. No inhibition of glucose utilization
was produced with unsaturated fatty acids, oleic and linoleic. Calcium ions added in excess reversed the
antimicrobial effect of capric acid, but not that of caprylic acid. Antimicrobial activity of caprylic and capric
acid decreased when the bacteria were grown in the presence of straw particles, or repeatedly subcultured in
a medium containing these compounds at low concentrations. Counts of viable bacteria determined by
plating decreased after incubation with caprylic and capric acid (30 min; 1 g/L) at pH 5.2 from >10
9
to
≈10
2
/mL. A reduction of a mere 0.94–1.96 log
10
CFU was observed at pH 6.5–6.6. It can be concluded that
caprylic acid, and to a lesser extent also capric acid, has a significant antimicrobial activity toward E. coli.
Effects of other fatty acids were not significant or absent.
Young animals are susceptible to the development of digestive disorders caused by Escherichia
coli, salmonellas and other feed-borne pathogens. Antibiotics are used to control enteritis infections but their
use has been viewed critically. This had led to a renaissance of the research into the use of nonantibiotic
compounds with bactericidal activity (Lauková and Mareková 2001; Morovský et al. 2001; Pantev et al.
2002). Fatty acids and their derivatives are among the candidate replacements for antibiotics (Jalč et al. 2002;
Ricke 2003). Some of them have a long history of being utilized as additives and preservatives, preventing
food and feed deterioration (acetic, propionic, lactic and sorbic acid). Antimicrobial activity of fatty acids
has been known for a long time. Fatty acids facilitate the transport of protons across the bacterial membrane
and thus collapse the proton gradient. This results in depletion of cellular energy followed by denaturation
of acid-sensitive proteins and nucleic acids. Nieman (1954) concluded that (i) fatty acids with a chain length
of around C
12
were the most active, (ii) inhibitory properties of unsaturated fatty acids were more pro-
nounced than those of saturated acids, and (iii) G
+
-bacteria were more susceptible to the action of fatty acids
than G
–
-bacteria. Antimicrobial activity of saturated C
4
–C
18
fatty acids and oleic acid in aerobically grown
cultures of E. coli was examined by Hassinen et al. (1951). Bacterial growth was inhibited by capric (C
10
)
and lauric (C
12
) acid. Caprylic acid (C
8
) inhibited the growth of E. coli only slightly. Other saturated fatty
acids and oleic acid did not inhibit the growth of the strain tested. In experiments of Sprong et al. (2001)
capric and lauric acid appeared to be effective bactericidal bovine milk fat components, active against
E. coli and other enteropathogens. As for short-chain fatty acids (SCFA; <C
8
), the results of Hassinen et al.
(1951) are not consistent with the results of some later studies. Prohászka (1980) suggested that undissocia-
ted SCFA were responsible for the inhibitory action of the rabbit cecal contents against E. coli strains, and
on the same basis Wallace et al. (1989) explained the failure of E. coli to colonize the ovine rumen. Cherring-
ton et al. (1990) observed a bacteriostatic effect of 5 mmol/L propionic acid in cultures of E. coli at pH 5.0.
Similarly, Mortensen et al. (1999) reported that SCFA in concentrations normally found in the human colon
had an inhibitory effect on the growth of E. coli.
*Present address: Institute of Animal Physiology and Genetics, Academy of Sciences of the Czech Republic, 142 20 Prague, Czechia.
732 M. MAROUNEK et al. Vol. 48
The aim of our study was to determine the susceptibility of two strains of E. coli to C
2
–C
18
fatty
acids. Furthermore, the effect of calcium ions, the presence of feed particles in the testing medium, adapta-
tion effect, and effect of pH were examined.
MATERIALS AND METHODS
E. coli strains CCM 3954 (ATCC 25922) and CCM 4225 (ATCC 35218) were obtained from the
Czech Collection of Microorganisms (Brno, Czechia). The former strain is a clinical isolate, both strains
being reference strains for testing of antimicrobials. The bacteria were grown on a medium containing
(in g/L): K
2
HPO
4
·3H
2
O 5.9, KH
2
PO
4
4.5, NaHCO
3
3.0, (NH
4
)
2
SO
4
2.9, NaCl 0.9, MgSO
4
·7H
2
O 0.09,
CaCl
2
0.09, bactopeptone 6, glucose 5, yeast extract 3. A trace metal solution (1 mL) containing nitrilotri-
acetic acid (Clark and Holms 1976) was also added. Experiments were carried out as quintuplicates.
1st experiment. The medium was dispensed to gas-tight glass flasks containing acetic, propionic,
butyric, isobutyric, valeric, isovaleric, caproic, caprylic, capric, lauric, myristic, palmic, stearic, oleic and
linoleic acid (0, 0.1, 0.2, 0.3, 0.5, 1, 2, 3, 5 g/L), together with an equivalent amount of 5 mol/L NaOH. The
flasks were filled with CO
2
, closed by rubber stoppers and autoclaved at 110 °C for 45 min. Inoculated cul-
tures were incubated in triplicates at 37 °C for 1 d. Then the pH was measured using the pH03 pH-meter
from Labio Praha (Prague) and residual glucose was determined enzymically by the oxidase–peroxidase
method using a commercial kit Oxochrom Glucose from Lachema Brno (Czechia). The amount of residual
glucose was expressed relative to the initial glucose concentration and plotted against the concentration of
the fatty acid. The IC
50
was the concentration at which only 50 % of the initial glucose was utilized. This
method avoids problems caused by opalescence or turbidity of media containing
≥
C
8
-fatty acids. In control
cultures and those containing C
2
–C
6
-fatty acids the growth was assessed by measuring absorbance A
640
at
the end of incubation.
2nd experiment. The bacteria were cultivated in media containing caprylic and capric acid (for con-
centrations see above), supplemented with CaCl
2
at 0.09 (control) or 1.8 g/L. After a 1-d incubation, the pH
was measured and residual glucose determined (see above).
3rd experiment. The bacteria were grown in media containing caprylic and capric acid and supple-
mented with particles of wheat straw at 0 or 10 g/L. The average size of straw particles (determined by dry
sieving) was 0.2 mm. the IC
50
was determined in 1-d cultures.
4th experiment. The bacteria were transferred 10
×
in media containing low concentrations of
caprylic and capric acid (3
×
lower than IC
50
found in the 1st experiment). IC
50
was then determined in the
cultures of both strains using adapted and nonadapted inocula.
5th experiment. Caprylic and capric acid were added to overnight-grown cultures of both strains at
1 g/L as 20 % (W/W) Me
2
SO solutions. Controls received an equivalent amount of the solvent. The cultures
were incubated at 37 °C in a water bath for 30 min; treated cultures were incubated at pH ≈5.2 and 6.5–6.6.
In the latter cultures, the pH was increased by addition of NaOH. After incubation, samples were removed
and serially diluted with the Wilkins–Chalgren broth (Oxoid). The number of viable bacteria was determi-
ned by streaking 0.1 mL of an appropriate dilution on Wilkins–Chalgren agar plates using sterile glass rods.
Inoculated plates were incubated at 37 °C for 1 d. Colonies were counted and means and SD calculated. The
significance of differences between control and treated cultures was evaluated by the t-test.
RESULTS
Table I top presents fatty acid concentration causing a 50 % decrease in the glucose utilization by
given E. coli strain. Both strains examined were susceptible to caprylic and capric acid. Control cultures and
those containing shorter (C
2
–C
6
) or longer (C
12
–C
18
) fatty acids utilized glucose completely. This was
accompanied by a pH decrease from 6.7 at the beginning of the incubation to 5.6 ± 0.1 at the end. The inhi-
bitory concentration of caprylic acid was not influenced by CaCl
2
addition; on the contrary, capric acid lost
its antimicrobial activity in cultures supplemented with CaCl
2
at 1.8 g/L (data not shown).
The antimicrobial activity of C
8
- and C
10
-acids decreased when the bacteria were grown in the pre-
sence of straw particles (Table I middle), or repeatedly grown in the presence of low fatty acid concentrat-
ions (Table I bottom). Caproic acid (C
6
) decreased the absorbance of E. coli cultures in a dose-dependent
manner while the effect of valeric acid (C
5
) was less pronounced (Fig. 1). Other SCFA had no influence on
growth of the two strains.
2003 EFFECT OF FATTY ACIDS ON E. coli 733
Incubation of bacteria with C
8
- and C
10
-acids at pH 5.2 led to a considerable reduction in the num-
bers of viable cells (Table II) while a reduction of a mere 0.94–1.96
log
10
CFU was observed at pH 6.5–6.6.
a
In which only 50 % of the initial glucose
was utilized within a 1-d incubation inter-
val.
b
Including branched isomers.
c
Stearic, oleic and linoleic acids.
d
Particles of wheat straw, 10 g/L.
e
Control values (i.e. without particles) in
parentheses.
f
Subcultured 10
×
in a medium containing
C
8
- or C
10
-acid at IC
50
/3.
Table II. Cell concentration of E. coli (c, 1/mL)
a
determined in strains CCM 3954 and CCM 4225 by plating after
a 30-min incubation with C
8
- and C
10
-acids at different pH
Treated cultures Control cultures
Strain
pH log c fatty acid
b
pH log c
CCM 3954 5.53 9.67 ± 0.09 C
8
6.52 8.37 ± 0.18*
5.20 <2*
C
10
6.49 8.63 ± 0.33*
5.18 6.35 ± 0.32*
CCM 4225 5.35 9.60 ± 0.33 C
8
6.60 7.64 ± 0.32*
5.21 2.56 ± 0.48*
C
10
6.54 8.42 ± 0.08*
5.19 <2*
a
Surviving after treatment with fatty acids; means of triplicates ±SD.
b
g/L.
*Significantly different from the control values (p < 0.01). Cell concentration <100/mL was considered as 100/mL
in statistical calculations.
DISCUSSION
Caprylic and capric acids, but not other acids prevented utilization of glucose in the cultures of
2 strains of E. coli. Unsaturated C
18
-fatty acids and saturated C
2
–C
6
- and C
12
–C
18
-fatty acids did not
influence substrate utilization.
The inhibitory effect of SCFA on E. coli reported by Prohászka (1980) and Wallace et al. (1989)
was not observed in this study with our strains. Caproic acid, and to a lesser extent also valeric acid, how-
ever, decreased cell yield expressed as turbidity of cultures. Both Prohászka (1980) and Wallace et al.
(1989) found that inhibitory effect of SCFA on E. coli was pH-dependent and ceased in the neutral pH
range. In our study the pH in cultures supplemented with SCFA fell from 6.71 at the beginning of the incu-
bation to 5.55 at its end. The SCFA were thus present in the cultures mostly in their active, nondissociated
form, except for the initial period. Therefore, also the activity of medium-chain fatty acids (C
8
, C
10
)
depends on pH, being much higher in an acid environment. It is noteworthy that a certain bactericidal acti-
vity of C
8
- and C
10
-acids exists at pH near 7. This has not been the case for formic and lactic acid, which
allowed growth of E. coli in the neutral pH range (Dibner and Buttin 2002). The acid milieu itself was not
Table I. Inhibitory concentration (g/L)
a
of C
2
–C
18
fatty acids against E. coli
strains CCM 3954 and CCM 4225 grown on glucose
Fatty acid CCM 3954 CCM 4225
C
2
, C
3
, C
4
b
, C
5
b
, C
6
>5 >5
C
8
0.45 0.30
C
10
2.03 1.25
C
12
, C
14
, C
16
, C
18
c
>5 >5
Growth in the presence of solid particles
d,e
C
8
1.72 (0.82) 1.83 (0.85)
C
10
2.57 (1.98) 5.00 (1.90)
Growth of adapted
e,f
strains
C
8
0.92 (0.63) 1.20 (0.47)
C
10
>5. (2.00) 2.90 (1.92)
734 M. MAROUNEK et al. Vol. 48
bactericidal as concentrations of viable bacteria taken from control cultures (pH 5.53 and 5.35) were >1/pL
(i.e. >10
9
/mL; Table II). Enterobacteriaceae while traveling through the gastrointestinal tract must endure
extremely low pH in the stomach. E. coli, salmonellas and shigellas possess specific survival mechanisms
that include amino-acid decarboxylases and shock proteins that prevent or repair macromolecular damage
(reviewed by Bearson et al. 1997). Apparently, medium-chain fatty acids are capable of overcoming this
barrier.
Several factors can influence the antimicro-
bial activity of fatty acids. Galbraith et al. (1971)
showed that calcium ions added in excess as CaCl
2
reversed the antimicrobial effects of lauric (C
12
)
and linoleic (C
18:2
) acid. Fatty acids differed in
the extent to which they reacted with cations to
form insoluble salts. Increasing chain length
increased the extent of insoluble calcium soap
formation (Jenkins and Palmquist 1982). In this
study CaCl
2
added in excess counteracted capric
acid, but not caprylic acid, apparently due to the
low affinity of the latter acid for Ca
2+
-ions. Har-
foot et al. (1975) and Maczulak et al. (1981) sug-
gested that the presence of a particulate phase in
a milieu reduced the inhibitory effect of fatty
acids on bacterial growth. This has also been
shown in Table I (middle). Food particles pro-
vide a site for the adsorption of fatty acids, which
would otherwise adsorb onto the bacteria and
inhibit bacterial metabolism and growth. As expec-
ted, an increase of the resistance of E. coli strains
was observed after their serial transfer in a me-
dium containing a subinhibitory concentration of
caprylic and capric acid.
Sprong et al. (2001) considered the bacteri-
cidal activity of lipids as biologically significant
when a reduction to >0.5 log
10
CFU/mL was
observed. Both caprylic and capric acid meet this
criterion. These acids were identified as the antimicrobial compounds of the stomach contents of suckling
rabbits (Cañas-Rodriguez and Smith 1966). Our previous work (Marounek et al. 2002) indicated a signifi-
cant antimicrobial activity of caprylic and capric acid against bacteria of the animal digestive tract. The results
presented here showed that these acids were active also against E. coli and may thus enhance the resistance
to certain types of gastrointestinal infections.
This work was supported by grant no. QF3134 of the Czech National Agency for Agricultural Research (NAZV).
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Medium chain organic acids (MCOA) show strong and specific antimicrobial activity and may be useful as part of an antibiotic reduction strategy. Two trials were conducted to evaluate the effect of MCOA on growth performance of pigs in comparison with benzoic acid (BA) and ZnO (ZnO). Each used 240 pigs in a randomised complete block design with six treatments in eight replicate pens. Trial 1 tested a blend of C6:0, C8:0, C10:0 and C12:0 for 56 d post-weaning. Treatments were (1) control basal diet (BD), (2) BD+ZnO (3) BD+ZnO+BA, (4) BD+MCOA (5) BD+MCOA+ZnO, (6) BD+MCOA+BA. Pigs receiving MCOA+BA showed higher average daily liveweight gain in the grower period (849 g/d) than pigs receiving the control diet (773 g/d) or the ZnO treatment (779 g/d; P=0.040), grew 7% more efficiently (P=0.008) and were 3.02 kg heavier at 56 d than control pigs. Trial 2 tested the MCOA blend at two inclusion levels in comparison with C6:0/C12:0 (50/50) and C6:0/C12 monolaurin (C12M; 50/50), for 28 d post-weaning. Treatments were (1) control basal diet, (2) BD+ZnO (3) BD+1,500 mg/kg MCOA (4) BD+750 mg/kg MCOA (5) BD+ C6/C12 (total 3,000 mg/kg), (6) BD + C6/C12M (total 3,000 mg/kg). Pigs receiving 750 mg/kg MCOA grew more slowly than those receiving C6/C12M or ZnO (P<0.05) to d 21. Pigs fed ZnO were 1.61 kg heavier than these pigs (P<0.05), while those fed C6/C12M were 1.29 kg heavier (P<0.05) at d 21. During 0-14 d pigs fed ZnO showed the best feed conversion ratio (FCR), but at 15-28 d the improvement was no longer seen. Both the C6/C12 and the C6/C12M treatments appeared to improve FCR during this second phase (P<0.01). The results suggested that supplementation with a combination of MCOA and BA may be a promising strategy to improve growth performance while replacing ZnO and reducing antibiotic dependence.
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... There are conflicting reports about the relative antimicrobial efficacy of different MCOA. Only caprylic acid (C8:0), and, to a lesser extent, capric acid (C10:0) have shown significant antimicrobial activity against two strains of Escherichia coli in one study (Marounek et al., 2003). However, in another study, both lauric acid (C12:0) and caprylic acid (C8:0) had the ability to damage the cytoplasmic structure of bacteria (Skrivanová et al., 2006). ...
Comparison of caproic acid and monolaurin with zinc oxide and benzoic acid; effect on growth performance of weaned pigs
Article
Full-text available
  • Feb 2022
Medium chain organic acids (MCOA) offer the potential to replace zinc oxide, without dependence on antibiotic use, and can replace or reduce the use of benzoic acid in pig production. A study was conducted to evaluate the effects of caproic acid (C6) and C12 monolaurin (C12M) on the growth performance of weaned pigs, in comparison with zinc oxide (ZnO) and benzoic acid (BA). Specifically, the trial examined whether MCOA, at a reduced inclusion level of BA, could maintain performance levels of pigs similar to those receiving a higher inclusion level of BA. In total, 240 pigs from weaning until 27 d post-weaning were used in a completely randomised block trial with six treatments and eight replicate pens per treatment. Treatments were: (1) control (CON), basal diet (BD); (2) BD+ZnO (ZnO); (3) BD+5,000 mg/kg BA (BA); (4) BD+2,500 mg/kg BA+2,000 mg/kg C12M+700 mg/kg C6 (BA low /MCOA); (5) BD+2,000 mg/kg C12M+700 mg/kg C6 (MCOA); (6) BD+1000 mg/kg C12M+700 mg/kg C6 (MCOA low ). Feed and water were provided ad libitum. Pigs receiving BA or BA low /MCOA had improved feed conversion ration (FCR) during the period 0-14 days, compared with control pigs (1.02 vs 1.17, P<0.05; 1.06 vs 1.17; P=0.084, respectively). The FCR of pigs receiving BA low /MCOA was similar to those receiving BA at the higher inclusion-level. Pigs showed good health throughout the study, as indicated by clinical and faecal scores. Replacement of ZnO was inconclusive and further studies with more challenging health conditions are required to conclude benefits.
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... On the other hand, the FA are also beneficial for human health, for example, C4:0 stimulates the proliferation of colonic cells and inhibits the growth of colon cancer cell lines (Chang et al., 2014;Dierick et al., 2002;Fauser et al., 2011;He et al., 2020). FA C8:0 and C10:0 are antimicrobial in nature (Marounek et al., 2003) and cause reduction of triacylglycerides and low-density lipoprotein levels, and they have been reported to induce thermogenesis (Terada et al., 2012). FA C12:0, C14:0 and C16:0 bring about increase in high-density lipoprotein cholesterol (Orsavova et al., 2015). ...
Effect of dehydration and butter-frying on chinicuil (Comadia redtenbacheri Hammershmidt) and maguey white worm (Aegiale hesperiaris Walker)
Article
Full-text available
  • Feb 2022
In the state of Hidalgo, México, chinicuil (CH) and maguey white worm (MW) are consumed as well as used in different culinary preparations. Few studies are available on the effect of butter-frying (BF) and dehydration (DN) on their chemical composition. The objective of this study was to compare the effect of DN and BF on the chemical composition and fatty acids (FA) content of CH and MW. CH and MW were purchased and then fried in butter. The fresh samples were dehydrated at 70 °C for 4 h, and their chemical analysis was performed. The Soxhlet method was used for the extraction of oils (DNCH, DNMW, BFCH, and BFMW), which were analysed with respect to the quality parameters (i.e. moisture, acidity, iodine, and peroxide). The FA contents were determined by gas chromatography coupled with mass spectrometer. The DNCH (72.82%) and DNMW (76.81%), revealed high concentrations of saturated FA. Free FA and hydro-peroxides were produced, which affected the percentage of saturated FA in the BFCH (15%) and BFMW (17%) samples, and unsaturated FA by 85 and 84%, respectively, along with the production of trans-FA. Therefore, it´s necessary to regulate the frying conditions, as BF causes harmful changes in their chemical composition and in their structure, in CH and MW, which can cause health problems.
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... On the other hand, the FA are also beneficial for human health, for example, C4:0 stimulates the proliferation of colonic cells and inhibits the growth of colon cancer cell lines (Chang et al., 2014;Dierick et al., 2002;Fauser et al., 2011;He et al., 2020). FA C8:0 and C10:0 are antimicrobial in nature (Marounek et al., 2003) and cause reduction of triacylglycerides and low-density lipoprotein levels, and they have been reported to induce thermogenesis (Terada et al., 2012). FA C12:0, C14:0 and C16:0 bring about increase in high-density lipoprotein cholesterol (Orsavova et al., 2015). ...
Effect of dehydration and butter-frying on chinicuil (Comadia redtenbacheri Hammershmidt) and maguey white worm (Aegiale hesperiaris Walker)
Article
Full-text available
  • Jun 2021
In the state of Hidalgo, México, chinicuil (CH) and maguey white worm (MW) are consumed as well as used in different culinary preparations. Few studies are available on the effect of butter-frying (BF) and dehydration (DN) on their chemical composition. The objective of this study was to compare the effect of DN and BF on the chemical composition and fatty acids (FA) content of CH and MW. CH and MW were purchased and then fried in butter. The fresh samples were dehydrated at 70 °C for 4 h, and their chemical analysis was performed. The Soxhlet method was used for the extraction of oils (DNCH, DNMW, BFCH, and BFMW), which were analysed with respect to the quality parameters (i.e. moisture, acidity, iodine, and peroxide). The FA contents were determined by gas chromatography coupled with mass spectrometer. The DNCH (72.82%) and DNMW (76.81%), revealed high concentrations of saturated FA. Free FA and hydro-peroxides were produced, which affected the percentage of saturated FA in the BFCH (15%) and BFMW (17%) samples, and unsaturated FA by 85 and 84%, respectively, along with the production of trans-FA. Therefore, it´s necessary to regulate the frying conditions, as BF causes harmful changes in their chemical composition and in their structure, in CH and MW, which can cause health problems.
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... Reference [13] states that there are two acid mechanisms in inhibiting bacterial development, namely, the undissociated form of citric acid will directly enter the lipid membrane of gram-negative bacteria and citric acid dissociates directly because of the ability of its hydrogen-free protons which can lower the pH so as to create acidic conditions, so that these conditions have a negative effect on the presence of pathogenic bacteria. Organic acids that enter the cytoplasm which has neutral pH conditions inhibit bacterial growth by inhibiting the oxidative phosphorylation process and increasing energy consumption. ...
The Effect of Acidifier-Dextrose against Hen Day Production and Feed Conversion Ratio in Laying Hens Infected with Avian Pathogenic Escherichia coli
Article
Full-text available
  • Mar 2021
Colibacillosis in Indonesia until now still appears frequently, so the case of colibacillosis laying hens cannot reach the peak of egg production; the egg production period is delayed and easily infected with other diseases. The purpose of this research is that the acidifier-dextrose combination is expected to be able to suppress the development of Avian Pathogenic Escherichia coli (APEC) bacteria in laying hens so that, in the end, the case of colibacillosis can be controlled in Indonesia. A total of 240 heads of laying hens were divided into 6 treatments and each consisted of 40 replications. The results of this research state that a combination of acidifier-dextrose can increase Hen Day Production (p
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Creating new water‐soluble Iso ‐ and n ‐fatty acid arginate hydrochloride with antimicrobial properties
Article
  • Jul 2023
Typically, short‐ and long‐chain lipids from oils exhibit different antimicrobial activities and therefore have been used in agriculture and aquaculture, biomedical therapeutic and antibacterial fields. However, these fatty acids have limitations in terms of bioactive efficacy, thermostability and aqueous solubility. In this study, water‐soluble iso ‐fatty acid arginate hydrochloride derivatives with antimicrobial properties were produced by introducing branched ( iso ‐) chain and other linear‐ ( n ‐) chain fatty acids to the “arginine” amino acid molecule. The two‐step synthetic route was straightforward and provided an efficient 88% and 76% product yields for ethyl n ‐oleoyl arginate hydrochloride and ethyl iso ‐oleoyl arginate hydrochloride, respectively. ATR‐FT‐IR, NMR, and LC‐MS‐Q‐TOF techniques were used to thoroughly characterize and confirm the products. These arginate products had strong antimicrobial activities against Listeria innucua , a Gram‐positive bacterium with minimum inhibitory concentrations and minimum bactericidal concentrations ranging from 1.8 µg mL ⁻¹ to 29.1 µg mL ⁻¹ . Therefore, the study demonstrated the development of a novel class of antimicrobial compounds from iso ‐fatty acids and arginates.
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Benzoxazine Monomers Grafted Poly(acrylic acid) as Novel Organic Additives with Fluorescence and Antibacterial Properties
Article
  • Apr 2022
Novel additives with dual fluorescence and antibacterial properties derived from benzoxazine monomers and poly(acrylic acid) (PAA) were prepared. Three benzoxazine monomers containing a hydroxyl group (BZXs) synthesized via the Mannich reaction were used as organic fluorophores and subsequently reacted with PAA to obtain poly(acrylic acid)-grafted benzoxazines (PAA-g-BZXs) via esterification. The results from Fourier transform infrared (FTIR) spectroscopy and ¹H nuclear magnetic resonance (NMR) showed that the BZXs were grafted onto the pendent groups of PAA only 25%. From UV-Vis, the maximum absorption wavelengths of PAA-g-BZXs were shown the same position of BZXs. By fluorescence spectroscopy, the excitation, emission, and fluorescence quantum yield of BZXs were found to be 375-480 nm, 550-585 nm, and 0.60, respectively. Meanwhile, PAA-g-BZXs showed the same excitation and emission wavelength positions around 360-485 nm and 500-585 nm, respectively. PAA-g-BZXs exhibited green emission light under a UV lamp due to the benzoxazine pendent groups. However, the fluorescence quantum yield of PAA-g-BZXs was decreased to be 0.40 BZXs due to only 25% of BZXs grafted on the pendent groups of PAA. Moreover, PAA-g-BZXs showed antibacterial activity against S. aureus and E. coli up to 100%. Only 1% by weight PAA-g-BZXs was an appropriate amount for the poly(lactic acid) (PLA) matrix to fabricate golden PLA/PAA-g-BZXs composite wires without adding any pigments since the obtained composite wires still showed a high optical performance by changing the color from gold under visible light to blue-green under UV irradiation.
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Effect of Added Fat and Calcium on in Vitro Formation of Insoluble Fatty Acid Soaps and Cell Wall Digestibility
Article
Full-text available
  • Jan 1982
Summary In vitro experiments were conducted to determine the extent to which fatty acids react with divalent cations to form insoluble soaps in rumen contents. The addition of 10% tallow to a hay and corn (70/30) basal substrate increased (P
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Use of Organic Acids as a Model to Study the Impact of Gut Microflora on Nutrition and Metabolism1
Article
Full-text available
Approvals for the use of nontherapeutic antibiotics in animal feed are fast disappearing worldwide. The primary effect of antibiotics is antimicrobial; all of the digestibility and performance effects can by explained by their impact on the gastrointestinal microflora. Among the candidate replacements for antibiotics are organic acids, both individual acids and blends of several acids. Like antibiotics, short-chain organic acids also have a specific antimicrobial activity. Unlike antibiotics, the antimicrobial activity of organic acids is pH dependent. Organic acids have a clear and significant benefit in weanling piglets and have been observed to benefit poultry performance. Organic acids have antimicrobial activity; however, there appear to be effects of organic acids beyond those attributed to antimicrobial activity. Reductions in bacteria are associated with feeding organic acids, which are particularly effective against acid-intolerant species such as E. coli, Salmonella and Campylobacter. Both antibiotics and organic acids improve protein and energy digestibilities by reducing microbial competition with the host for nutrients and endogenous nitrogen losses, by lowering the incidence of subclinical infections and secretion of immune mediators, and by reducing production of ammonia and other growth-depressing microbial metabolites. Organic acids have several additional effects that go beyond those of antibiotics. These include reduction in digesta pH, increased pancreatic secretion, and trophic effects on the gastrointestinal mucosa. Much more is known about these effects in swine than in poultry. There appears to be more variability in detecting an organic acid benefit in comparison to that observed with antibiotics. Lack of consistency in demonstrating an organic acid benefit is related to uncontrolled variables such as buffering capacity of dietary ingredients, presence of other antimicrobial compounds, cleanliness of the production environment, and heterogeneity of gut microbiota. Additional research can clarify the role of these factors and how to minimize their impact.
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Perspectives on the use of organic acids and short chain fatty acids as antimicrobials
Article
  • Apr 2003
Organic acids have a long history of being utilized as food additives and preservatives for preventing food deterioration and extending the shelf life of perishable food ingredients. Specific organic acids have also been used to control microbial contamination and dissemination of foodborne pathogens in preharvest and postharvest food production and processing. The antibacterial mechanism(s) for organic acids are not fully understood, and activity may vary depending on physiological status of the organism and the physicochemical characteristics of the external environment. An emerging potential problem is that organic acids have been observed to enhance survivability of acid sensitive pathogens exposed to low pH by induction of an acid tolerance response and that acid tolerance may be linked to increased virulence. Although this situation has implications regarding the use of organic acids, it may only apply to circumstances in which reduced acid levels have induced resistance and virulence mechanisms in exposed organisms. Evaluating effectiveness of organic acids for specific applications requires more understanding general and specific stress response capabilities of foodborne pathogens. Development and application of molecular tools to study pathogen behavior in preharvest and postharvest food production environments will enable dissection of specific bacterial genetic regulation involved in response to organic acids. This could lead to the development of more targeted strategies to control foodborne pathogens with organic acids.
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Effect of caprylic, capric and oleic acid on growth of rumen and rabbit caecal bacteria
Article
In a search for alternatives of in-feed antibiotics, the antimicrobial activity of caprylic (C8:0), capric (C10:0) and oleic (C18:1) acid was investigated in pure cultures of 19 strains of rumen and rabbit caecal bacteria, and in incubations of the rumen and rabbit caecal contents. In glucose-grown bacterial cultures the minimum inhibitory concentration (MIC) of caprylic acid ranged from 1 to 3 μl·ml-1. Two strains of Bacteroides ovatus were less susceptible to capric than to caprylic acid. In other strains, the MIC of capric acid was 0.25-0.50 μl·ml-1. The growth of most strains was not much affected by oleic acid. An exception to this were rumen bacteria Butyrivibrio fibrisolvens (MIC from < 0.05 to 1 μl·ml-1) and Lachnospira multiparus (MIC of 0.25 to 1 μl·ml-1). In incubations of the rumen and caecal contents caprylic and capric acid decreased the production of volatile fatty acids and gas, and increased production of lactate. In latter incubations the inhibitory effects of caprylic and capric acid were similar. In incubations of the rumen contents, capric acid was more efficient than caprylic acid when supplied at low concentrations (<1.25 μl·ml-1), but less efficient when supplied at 2.5 and 5 μl·ml-1. Effects of oleic acid in rumen and caecal cultures were not significant, except the increase in production of lactate by rumen microorganisms. It can be concluded that microorganisms of the animal digestive tract are susceptible to inhibition by caprylic and capric acid added to microbial cultures at fairly low concentrations. Oleic acid was far less effective.
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The bacteriostatic effect of saturated fatty acids
Article
1.1. The growth of gram-positive and gram-negative bacterial cultures were not significantly inhibited by 500 μg./ml. of C4, C6, C16, and C18 saturated fatty acids. Both gram-positive and gram-negative organisms were inhibited by the C8 and C10 saturated fatty acids. Only the gram-positive organisms tested were significantly inhibited by the C12 acid. The C14 acid caused growth inhibition at low concentrations in two of the microorganisms tested.2.2. With each increase of 2 carbon atoms from C8 through C12, the bacteriostatic effect increased an average of 3.5 times. A correlation is described between the increase in bacteriostatic effect and decrease in water solubility as the number of carbon atoms is increased.3.3. Increasing susceptibility to bacteriostasis by the C10 acid among the microorganisms tested was found to be correlated with an increasing number of the members of the saturated fatty acid series, C8, C12, C14, which cause bacteriostasis.
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Effects of short‐chain fatty acids on in vitro bacterial growth of Bacteroides fragilis and Escherichia coli
Article
Short-chain fatty acids (SCFA) are produced in the large bowel of nonruminant mammals by bacterial anaerobic fermentation. The aim of the present study was to investigate the effects of SCFA on the in vitro growth of Bacteroides fragilis and Escherichia coli. B. fragilis and E. coli isolated from fresh human clinical samples and a reference strain for each species were incubated in a meat infusion broth with increasing amounts of SCFA and grown under anaerobic conditions at a temperature of 37°C. Bacterial growth was estimated by spectrophotometry. Rate of growth was calculated from the logarithmic growth phase. SCFA, in concentrations normally found in the human colon, had a significant (p<0.01) inhibitory effect of the in vitro growth rate for E. coli, while they were without effect on the in vitro growth rate of B. fragilis. It may be concluded that under in vitro conditions SCFA had growth-inhibitory effects on E. coli, while they had no effect on B. fragilis.
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Toxicity of Volatile Fatty Acids at rumen pH prevents enrichment of Escherichia coli by sorbitol in rumen contents
Article
  • Jan 1989
Attempts were made to establishEscherichia coli ML308 in the sheep rumen by inoculating it in combination with the slowly metabolized sugar alcohol, sorbitol. Numbers were determined by plating dilutions on nutrient agar containing the chromogenic -galactoside, 5-bromo-4-chloro-3-indolyl--d-galactoside. This strain, alac-constitutive mutant, produced distinctive blue colonies.E. coli ML308 failed to grow in rumen fluid, despite being able to grow rapidly on sorbitol and in rumen fluid at pH 7.0. Its growth rate was depressed by relatively small drops in pH in the presence of volatile fatty acids (VFA), such that normal pH's of 6.2–6.6 in rumen contents were inhibitory. The indigenous remen bacterium,Streptococcus bovis, was much more resistant to the combination of high VFA concentrations and low pH. The success of this and similar strategies for the introduction of new organisms with beneficial new properties will, therefore, depend on the organism's having a tolerance to VFA over a range of rumen pH that enables them to survive in the same way as native species.
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Control of the Sequential Utilization of Glucose and Fructose by Escherichia coli
Article
In Escherichia coli (ATCCI5224; ML308), glucose and fructose phosphotransferase systems (PT-systems) are constitutive but activities are increased five and 10-fold respectively by aerobic growth on their respective substrates in defined media. In mixtures, glucose is used preferentially and the fructose PT-system activity is kept at its minimum; but, on glucose exhaustion, it overshoots its steady-state level and growth continues on fructose without lag. Cyclic AMP prevents overshoot. Continuous cultures operating as turbidostats on mixtures of glucose and fructose do not use fructose if sufficient glucose is present to support growth. If less glucose is available, it is all used and sufficient fructose is metabolized concurrently to maintain the growth rate characteristic of glucose. Both PT-systems are inhibited by hexose phosphates. Presence of homologous substrate specifically sensitizes each PT-system to inactivation by N-ethylmaleimide (NEM). Glucose diminishes the ability of fructose to sensitize its PT-system to NEM. This effect parallels the inhibition of fructose utilization by glucose and suggests that glucose denies fructose access to the fructose-specific part of the PT-system.
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The role of plant particles, bacteria and cell-free supernatant fractions of rumen contents in the hydrolysis of trilinolein and the subsequent hydrogenation of linoleic acid
Article
  • Feb 1975
The role of different fractions of rumen contents in the hydrolysis of trilinolein and the subsequent hydrogenation of the linoleic acid has been investigated by a series ofin vitro incubations. Hydrolysis of the trilinolein to free linoleic acid occurs almost wholly in the cell-free supernatant; the liberated linoleic acid in the supernatant can be rapidly adsorbed onto food particles where it is hydrogenated to stearic acid via the C18trans-11 monoene. Some 25% of the trilinolein added as substrate was taken up by the bacteria and of this a small percentage appeared to be hydrolysed and the free linoleic acid hydrogenated to stearic acid intracellularly. No conclusive evidence was obtained to suggest that this intracellular hydrogenation proceeded by a route other than that which took place on the food particles.
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