Figure 11.2a
223
Biomimetic Mineralization of Apatite on Bacterial Cellulose
Mass (mg/L)
0
1
2
3
4
5
6
7
AJ2000
M1604 1604A
M1604 1010B
M1604 0416B
M1604 0020B
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
Cultivation Time (day)
Figure 11.2a
Cellulose. production. in. the. presence. of. glucose,. mixed. sugars,. or. GlcNAc. as. a. function. of.
cultivation. time.. Source:. From. Nge. et. al.,. Biopolymers. (ISBN:978-953-307-109-1,. InTech.,. 2010),.
345–368,.with.permission.
concentrations.at.a.fixed.total.carbon.source..Shirai.et.al..(1994).also.reported.
the.accumulation.of.GlcNAc.during.incubation.in.mixed.sugar.media..For.
their. analysis,. the. authors. determined. the. time. course. of. sugar. consumption. by. using. Schales’s. modified. procedure. for. total. sugar. and. a. glucose.
oxidase/peroxidase.system.for.glucose.
Based. on. the. results. shown. in. Figure 11.2a,. the. cultivation. time. in. further.experiments.was.fixed.to.7–9.days.for.the.media.containing.GlcNAc.of.
all. starter. cultures.. Cellulose. produced. from. AJ2000. and. M1604A. showed.
a.similar.mass.profile,.whereas.a.slightly.lower.mass.profile.was.observed.
for.cellulose.produced.from.M0020A.starter.culture..Therefore.cell.adaptation.was.not.critical.in.starter.cultures.containing.small.amounts.of.GlcNAc.
(M1604A)..The.amount.of.incorporated.GlcNAc.was.determined.by.a.labeling.
method.based.on.the.use.of.radioactive.GlcNAc.(1.85.MBq.N-acetyl-D-[1- 14 C].
glucosamine;. Moravek. Biochemicals,. Inc.,. Brea,. CA,. USA). and. liquid. scintillation. counting. (LSC;. Packard. Tri-Carb,. Perkin. Elmer,. Inc.). (Nge. et. al..
2010a)..Small-scale.labeling.was.carried.out.in.1.ml.culture.media.containing.
different. amounts. of. nonradioactive. GlcNAc. and. inoculated. using. starter.
. cultures AJ2000,.M1604,.M1010,.M0416,.and.M0020..The.molar.concentration.
of.added.radioactive.GlcNAc.was.negligible.compared.to.the.concentration.
of. nonradioactive. sugar. in. the. culture. medium.. The. static. cultures. were.
incubated.for.7.days.under.the.same.conditions.as.for.bulk.production..The.
small.pellicle.formed.was.taken.out.at.days.2,.3,.5,.and.7.followed.by.purification.as.described.for.bulk.production..After.addition.of.the.scintillation.
cocktail.(Ultima.Gold,.Perkin.Elmer).in.the.vials.containing.the.purified.BC.
pellicles,. the. radioactivity. corresponding. to. the. incorporated. GlcNAc. was.
223
Biomimetic Mineralization of Apatite on Bacterial Cellulose
Mass (mg/L)
0
1
2
3
4
5
6
7
AJ2000
M1604 1604A
M1604 1010B
M1604 0416B
M1604 0020B
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
Cultivation Time (day)
Figure 11.2a
Cellulose. production. in. the. presence. of. glucose,. mixed. sugars,. or. GlcNAc. as. a. function. of.
cultivation. time.. Source:. From. Nge. et. al.,. Biopolymers. (ISBN:978-953-307-109-1,. InTech.,. 2010),.
345–368,.with.permission.
concentrations.at.a.fixed.total.carbon.source..Shirai.et.al..(1994).also.reported.
the.accumulation.of.GlcNAc.during.incubation.in.mixed.sugar.media..For.
their. analysis,. the. authors. determined. the. time. course. of. sugar. consumption. by. using. Schales’s. modified. procedure. for. total. sugar. and. a. glucose.
oxidase/peroxidase.system.for.glucose.
Based. on. the. results. shown. in. Figure 11.2a,. the. cultivation. time. in. further.experiments.was.fixed.to.7–9.days.for.the.media.containing.GlcNAc.of.
all. starter. cultures.. Cellulose. produced. from. AJ2000. and. M1604A. showed.
a.similar.mass.profile,.whereas.a.slightly.lower.mass.profile.was.observed.
for.cellulose.produced.from.M0020A.starter.culture..Therefore.cell.adaptation.was.not.critical.in.starter.cultures.containing.small.amounts.of.GlcNAc.
(M1604A)..The.amount.of.incorporated.GlcNAc.was.determined.by.a.labeling.
method.based.on.the.use.of.radioactive.GlcNAc.(1.85.MBq.N-acetyl-D-[1- 14 C].
glucosamine;. Moravek. Biochemicals,. Inc.,. Brea,. CA,. USA). and. liquid. scintillation. counting. (LSC;. Packard. Tri-Carb,. Perkin. Elmer,. Inc.). (Nge. et. al..
2010a)..Small-scale.labeling.was.carried.out.in.1.ml.culture.media.containing.
different. amounts. of. nonradioactive. GlcNAc. and. inoculated. using. starter.
. cultures AJ2000,.M1604,.M1010,.M0416,.and.M0020..The.molar.concentration.
of.added.radioactive.GlcNAc.was.negligible.compared.to.the.concentration.
of. nonradioactive. sugar. in. the. culture. medium.. The. static. cultures. were.
incubated.for.7.days.under.the.same.conditions.as.for.bulk.production..The.
small.pellicle.formed.was.taken.out.at.days.2,.3,.5,.and.7.followed.by.purification.as.described.for.bulk.production..After.addition.of.the.scintillation.
cocktail.(Ultima.Gold,.Perkin.Elmer).in.the.vials.containing.the.purified.BC.
pellicles,. the. radioactivity. corresponding. to. the. incorporated. GlcNAc. was.