In luence o calcium ni a e iming on he s uc u al and ex u al
cha ac e is ics o mesopo ous SiO
2
-CaO nanopa icles
Ona Bas
¸ak
a,*
, Fa ih Ku uldu
a,*
, Jan Ila ský
b
, Ma ina Vi ´
azko ´
a
a
, Ana M. Bel ´
an
c
,
F ancisco Mu˜
noz
d
, Yolanda Cas o
d
, Ma in Mich´
alek
a
, Ma ia Ch omˇ
cíko ´
a
a,e
a
FunGlass, A. Dubˇ
cek Uni e si y o T enˇ
cín, ˇ
S uden sk´
a 2 911 50 T enˇ
cín, Slo akia
b
X- ay Science Di ision, Ad anced Pho on Sou ce, A gonne Na ional Labo a o y, A gonne 60439, IL, USA
c
Depa amen o de Ingenie ía y Ciencia de los Ma e iales y del T anspo e, Escuela Poli ´
ecnica Supe io , Uni e sidad de Se illa, Se illa, 41011, Spain
d
Ins i u e o Ce amics and Glass (CSIC), Kelsen 5, Mad id 28049, Spain
e
VILA – Join Glass Cen e o he IIC SAS, TnUAD, FChPT STU, ˇ
S uden sk´
a 2 911 50 T enˇ
cín, Slo akia
ARTICLE INFO
Keywo ds:
Mesopo ous bioac i e nanopa icles
Mic oemulsion assis ed sol-gel syn hesis
Calcium ni a e e ahyd a e
ABSTRACT
Mesopo ous bioac i e glass nanopa icles (MBGNPs) a e p omising ma e ials o d ug deli e y due o hei high
po e olume and speci ic su ace a ea. This s udy in es iga es how he iming o calcium ni a e addi ion a ec s
he s uc u al and ex u al cha ac e is ics o MBGNPs syn hesized ia a mic oemulsion-assis ed sol-gel me hod.
Delayed calcium ni a e addi ion educed CaO inco po a ion om 14.2 o 9.5 mol% and inc eased pa icle size
om 178 ±51 nm o 256 ±30 nm. The speci ic su ace a ea alues inc eased wi h he delayed addi ion o
calcium ni a e, as obse ed h ough BET and USAXS/SAXS measu emen s. The p opo ion o Q
Si
n
uni s sligh ly
changed, bu no cy o oxici y was obse ed in os eoblas -like cells. These indings p o ide aluable insigh s in o
op imizing MBGNP syn hesis o biomedical applica ions.
1. In oduc ion
Bioac i e glasses (BGs) ha e been ex ensi ely s udied o biomedical
applica ions since hei de elopmen in he la e 1960s due o hei
ema kable abili y o bond wi h bone [1,2]. This unique p ope y a ises
om he o ma ion o a hyd oxyapa i e laye on hei su ace h ough
dissolu ion in con ac wi h physiological luids, which mimics he
mine al composi ion o bone. BG dissolu ion can be ailo ed by doping
he apeu ic ions and modi ying chemical composi ion, which can
enhance biological unc ions such as angiogenesis, os eogenesis, and
an ibac e ial ac i i y [3]. These p ope ies ha e expanded he BG ap-
plica ions in wound healing and so issue enginee ing [4]. Addi ion-
ally, he ab ica ion me hod, mel ing o sol-gel, no ably in luences he
ion elease beha io o BGs [5].
Mel quenching was he i s me hod o ab ica ing BGs [6]. How-
e e , i has ce ain limi a ions, including he use o high p ocessing
empe a u es and a na ow composi ion ange o main ain bioac i i y
[4]. Fo ins ance, mel -de i ed glasses can inco po a e only up o 60 mol
% SiO
2
, whe eas sol-gel syn hesis allows he ab ica ion o glasses wi h
up o 90 mol% SiO
2
a lowe empe a u es, while s ill p ese ing
bioac i i y [7]. This me hod in ol es hyd olysis and condensa ion e-
ac ions o p ecu so s (alkoxydes o sal s) o ob ain and in e connec ed
ino ganic ne wo k a oom empe a u e and ambien p essu e using, o
example, e ae hyl o hosilica e (TEOS) as he p ecu so [8]. Adjus ing
he ca alys and p ecu so s concen a ion can modi y he pa icle size
and mo phology, he eby in luencing bioac i i y [9]. These ad an ages
make he sol-gel me hod ideal o syn hesizing ad anced ma e ials such
as po ous sca olds, hyb id BGs, and mesopo ous bioac i e glass nano-
pa icles (MBGNPs) [10]. MBGNPs a e p omising hos s o a ious
d ugs, genes, and g ow h ac o s due o hei supe io su ace a ea,
la ge po e olume compa ed o con en ional BGs [11–13]. Addi ion-
ally, MBGNPs exhibi highe pu i y and homogenei y han mel -de i ed
glasses and can be u he modi ied due o he p esence o silanol g oups
[14].
In sol-gel syn hesis, calcium ni a e is widely used as a calcium
p ecu so due o i s solubili y in wa e and low cos [15]. Howe e , i s
inco po a ion in o he silica ne wo k p esen s signi ican challenges.
Calcium ni a es a e only inco po a ed in o he glass s uc u e a em-
pe a u es abo e 450 ◦C, whe eas silica ne wo k o ma ion begins om
oom empe a u e [16]. This misma ch in eac ion condi ions o en
* Co esponding au ho s.
E-mail add esses: [email p o ec ed] (O. Bas
¸ak), [email p o ec ed] (F. Ku uldu).
Con en s lis s a ailable a ScienceDi ec
Open Ce amics
jou nal homepage: www.sciencedi ec .com/jou nal/open-ce amics
h ps://doi.o g/10.1016/j.oce am.2025.100807
Recei ed 14 Ap il 2025; Recei ed in e ised o m 15 May 2025; Accep ed 3 June 2025
Open Ce amics 23 (2025) 100807
A ailable online 12 June 2025
2666-5395/© 2025 The Au ho (s). Published by Else ie L d on behal o Eu opean Ce amic Socie y. This is an open access a icle unde he CC BY-NC-ND license
(
h p://c ea i ecommons.o g/licenses/by-nc-nd/4.0/ ).
leads o he o ma ion o calcium- ich egions a he han a homoge-
neous dis ibu ion wi hin he silica e ne wo k [17]. Fu he mo e, dis-
c epancies be ween nominal and expe imen al composi ions ha e been
equen ly epo ed in s udies using calcium ni a e as a p ecu so
[18–20]. These disc epancies a e p ima ily a ibu ed o he washing
and cen i uga ion p ocesses, which a e essen ial o emo ing un eac-
ed p ecu so s and su ac an s, bu can also esul in he loss o calcium
ions. While hese s eps a e necessa y o main ain pa icle dispe sion and
p e en unwan ed side eac ions, hey exace ba e he gap be ween he
desi ed and achie ed composi ions.
The mic o-emulsion assis ed sol-gel me hod is one o he mos
common and acile app oaches o he syn hesis o MBGNPs, o e ing he
ad an age o p e en ing pa icle agglome a ion du ing syn hesis [8,21].
In his me hod, hexadecyl ime hylammonium b omide (CTAB) ac s as a
su ac an o acili a e he o ma ion o mesopo ous s uc u es. Unde
basic condi ions, CTAB, combined wi h e hyl ace a e (EA), o ms
oil-in-wa e (O/W) mic oemulsion d ople s, which se e as con ined
eac ion si es o he syn hesis p ocess [22]. Wi hin hese d ople s, he
hyd olysis and condensa ion o TEOS occu in he p esence o aqueous
ammonia, which unc ions as bo h a ca alys and a egula o o ex u al
p ope ies [23]. The concen a ions o ammonia and CTAB play a c i ical
ole in con olling pa icle size, mesopo e s uc u e, and mo phology.
Fo example, Liang e al. [24] obse ed ha a ying ammonia concen-
a ions led o di e en po e ypes (e.g., wo m-like, adial, and lamella )
and an inc ease in pa icle size om 28 o 255 nm. Simila ly, Li e al.
[25] epo ed di e ences om sphe es o od-shaped pa icles wi h
inc easing CTAB concen a ion. These indings highligh he unabili y
o MBGNP p ope ies h ough p ecise con ol o syn hesis pa ame e s.
Se e al s udies ha e shown ha he iming o calcium ni a e addi-
ion signi ican ly in luences he ex u al and s uc u al p ope ies o
MBGNPs. Fo ins ance, Zheng e al. [26] obse ed ha a ia ions in
calcium ni a e addi ion iming du ing syn hesis ia he modi ied S ¨
obe
me hod esul ed in no able changes in bioac i e glass nanopa icle ag-
g ega ion, composi ion, and mo phology. Simila ly, Kesse e al. [27]
epo ed compa able indings using a one-po syn hesis app oach,
highligh ing he c i ical ole o calcium ni a e iming on he pa icle
cha ac e is ics such as composi ion and mo phology. Kesse e al. [28]
also explo ed he bioac i i y o MBGNPs wi h high su ace a ea by
op imizing he Ca/Si a io.
In his s udy, he in luence o iming o calcium ni a e addi ion on
he s uc u al, ex u al, and biocompa ibili y p ope ies o bina y
MBGNPs syn hesized ia he mic oemulsion-assis ed sol-gel me hod was
e alua ed. The mo phology, su ace a ea, and po e s uc u e o MBGNPs
we e cha ac e ized using ad anced analy ical echniques. Addi ionally,
hei biological esponse was assessed in os eoblas -like cells o e alua e
hei po en ial o bone issue enginee ing applica ions. The esul s we e
compa ed wi h p e ious s udies o u he unde s and how syn hesis
pa ame e s in luence ma e ial p ope ies and biological pe o mance.
2. Ma e ials and me hod
2.1. Syn hesize o MBGNPs
Te ae hyl o hosilica e (TEOS, 99 %, Sigma-Ald ich), hexadecyl-
ime hylammonium b omide (CTAB, BioX a, ≥99 %, H9151, Sigma-
Ald ich), calcium ni a e e ahyd a e (Ca(NO₃)₂⋅4H₂O, 99.1 %, VWR),
e hyl ace a e (99.7 %, l-00,028, Cen alchem), and ammonium hy-
d oxide (ACS eagen , 28.0–30.0 % NH
3
basis, 221,228, Sigma-Ald ich)
we e used as p ecu so s in he syn hesis o MBGNPs wi h a nominal
composi ion o 70SiO
2
–30CaO (in mol %) ia a mic oemulsion-assis ed
sol-gel me hod [29]. In b ie , 1.4 g o CTAB was dissol ed in 66 ml o
deionized wa e , ollowed by adding 20 ml o e hyl ace a e (EA) and
s i ing a 25 ◦C o 30 min. Nex , 1.8 ml o ammonium hyd oxide (28 %)
was added and s i ed o 15 min. Then, 7.2 ml o TEOS was added,
ollowed by 4.26 g o calcium ni a e e ahyd a e, wi h a ying addi ion
imes (15 o 90 min). The mix u e was s i ed o 4 h a 25 ◦C. A e -
wa ds, he samples we e washed wice wi h deionized wa e , once wi h
e hanol by cen i uga ion, and d ied o e nigh a 60 ◦C. The d ied
samples we e calcined a 650 ◦C o 3 h wi h a hea ing a e o 1 ◦C/min.
The samples we e labelled as MBGNP_15min, MBGNP_30min,
MBGNP_45min, MBGNP_60min, and MBGNP_90min, co esponding o
calcium ni a e addi ion imes o 15, 30, 45, 60, and 90 min,
espec i ely.
2.2. Cha ac e iza ion o MBGNPs
The mo phology and pa icle size o MBGNPs we e analyzed using
ield emission scanning elec on mic oscopy (FESEM, Hi achi S4700) a
20 kV, wi h gold spu e -coa ing o imp o e conduc i i y. Pa icle size
was de e mined by analyzing a leas 100 pa icles using ImageJ
so wa e.
The chemical composi ions o MBGNPs we e analyzed using Wa e-
leng h X- ay Fluo escence (WXRF) wi h a PANaly ical MagicX de ice
(PW-2424) equipped wi h a Rh anode RX ube and a 2.4 kW gene a o .
MBGNPs we e homogeneously mixed wi h anhyd ous li hium e abo-
a e (Li₂B₄O₇) lux and li hium b omide (LiB ) as a elease agen . The
mix u e was mel ed a 1100 ◦C using a Philips Pe l’X3 pea lize and a
pla inum-gold c ucible. The mel ed mix u e was hen cas on o molds o
ma ching composi ion o p oduce anspa en glass beads. Calib a ion
cu es we e cons uc ed using ce i ied silico-aluminous s anda ds o
co ela e he measu ed X- ay in ensi ies wi h elemen al concen a ions.
Mic os uc u al p ope ies o MBGNPs we e analyzed using ans-
mission elec on mic oscopy (TEM, Talos F200S G2, The mo ishe USA),
N
2
adso p ion-deso p ion (ASAP 2020, Mic ome i ics, USA) and Ul a-
Small and Small Angle X-Ray Sca e ing (USAXS/SAXS). Fo TEM, he
samples we e dispe sed in e hanol ia ul asonic ba h, hen moun ed on
ca bon g ids and imaged a 200 kV. P io o N
2
adso p ion-deso p ion,
MBGNPs we e degassed a 150 ◦C o 1200 min. The speci ic su ace
a ea was de e mined using he B unaue -Emme -Telle (BET) me hod,
while he Ba e -Joyne -Halenda (BJH) me hod was used o po e size
dis ibu ion analysis. USAXS/SAXS measu emen s we e conduc ed a
12-ID-E beamline a Ad anced Pho on Sou ce, A gonne Na ional Labo-
a o y. The combined sca e ing ec o magni ude (q) ange was be-
ween 1 and 1 ×10
−4
Å
−1
; he e q =4
π
/λ sin(θ), λ is he wa eleng h and
θ is ½ o he sca e ing angle. The X- ay ene gy was 21 keV (λ =0.5895
Å) and he X- ay pho on lux densi y was ≈5 ×10
12
mm
−2
⋅s
−1
. Be o e
measu emen s, he weigh o he sample holde was eco ded bo h wi h
and wi hou he MBGNP powde s, ensu ing he weigh o he ape
windows was accoun ed o o acili a e absolu e in ensi y calib a ion.
The olume occupied by he powde was de e mined om he di-
mensions o he sample holde . The olume and weigh di e ences
allowed he calcula ion o he packing densi y o he sample du ing
measu emen . The esul ing packing densi y was hen used o calib a e
he measu ed signal ela i e o MBGNP weigh .
Fo s uc u al cha ac e iza ion o silica e uni s, Fou ie T ans o m
In a ed (FTIR) spec a we e eco ded in abso p ion mode using a
Pe kin-Elme Spec um 3 FTIR Spec ome e (USA) wi hin he
400–4000 cm
−1
wa eleng h ange, wi h a esolu ion o 4 cm
−1
and 32
scans. Magic-Angle Spinning (MAS) NMR spec a we e eco ded using a
B uke AV-400-WB NMR spec ome e , employing 4 mm Z O₂ o o s
wi h Kel-F caps. ²⁹Si MAS-NMR spec a we e measu ed a 79.49 MHz
wi h
π
/2 pulses a 62.5 kHz, a spec al wid h o 40 kHz, and a MAS speed
o 10 kHz. A elaxa ion delay o 60 s was applied be ween scans, wi h
kaolin as a seconda y e e ence, calib a ed o a chemical shi o 91.2
O. Bas¸ak e al.
Open Ce amics 23 (2025) 100807
2
ppm ela i e o e ame hylsilane (TMS). MAS-NMR spec a we e
simula ed using Dm i so wa e [30].
2.3. Ion elease beha io o MBGNPs
The ion elease es was pe o med in a T is/HCl bu e (50 mM, pH
7.4). MBGNPs we e imme sed in he bu e a a concen a ion o 1.5 mg/
mL and incuba ed a 37 ◦C wi h con inuous s i ing a 120 pm o up o
7 days. A p ede e mined ime in e als, aliquo s we e collec ed, sy inge
il e ed, and supe na an s analyzed using induc i ely coupled plasma
op ical emission spec oscopy (ICP OES Agilen 5100 SVDV, Agilen
Technologies, USA). Calib a ion solu ions we e p epa ed using ce i ied
ICP e e ence s anda ds (Analy ika, P ague, Czech Republic), and
scandium was employed as an in e nal s anda d o minimize non-
spec al in e e ences. Da a we e epo ed as mean alues ±s anda d
de ia ions om a leas h ee eplica es o each elemen .
2.4. In- i o cy o oxici y assay
In i o cy ocompa ibili y was es ed using MG-63 human os eoblas -
like cells line (Sigma-Ald ich, Ge many). The cells we e cul u ed in
Dulbecco’s Modi ied Eagle Medium (DMEM, Gibco, Ge many) wi h 10
% e al bo ine se um (FBS, Gibco, Ge many) and 1 % Penicillin-
S ep omycin (P/S, Gibco, Ge many), in a humidi ied a mosphe e wi h
5 % CO
2
a 37 ◦C. Indi ec es ing was pe o med using supe na an s.
MBGNPs we e s e ilized by hea ea men a 160 ◦C o 3 h, added o he
cell cul u e medium a 10 % w/ concen a ion, and hen incuba ed a
37 ◦C o 24 h. Supe na an s we e collec ed ia cen i uga ion, il e ed
wi h a 0.22
μ
m sy inge il e , and dilu ed o 1 %, 0.1 %, and 0.01 % (w/
) in comple e medium o he cy ocompa ibili y assay.
MG-63 cells we e seeded in o 96-well pla es a an inoculum densi y
o 5 ×10
4
cells/mL and incuba ed o 24 h. The cells we e hen exposed
o ex ac s a 1 %, 0.1 %, and 0.01 % w/ concen a ions o 48 h. The
samples we e p epa ed in iplica e, and cell iabili y was assessed using
he WST-8 assay (CCK-8 Ki , Sigma-Ald ich, Ge many). Cells we e
incuba ed wi h 1 % / WST-8 in a cell cul u e medium o 3 h. A blank
Fig. 1. FESEM images and pa icle size dis ibu ion o MBGNPs.
O. Bas¸ak e al.
Open Ce amics 23 (2025) 100807
3
sample con aining only 1 % / WST-8 in a cell cul u e medium was also
incuba ed. A e incuba ion, 100
μ
L aliquo s om each well we e
ans e ed o a new 96-well pla e o spec ome ic analysis using a
mic opla e eade (NanoEnTek EVE Au oma ic cell coun e ) a 450 nm.
Rela i e cell iabili y was calcula ed using Eq (1).
S a is ical analysis was conduc ed using O igin 2018b so wa e wi h
one-way ANOVA and Bon e oni’s es . S a is ically signi ican di e -
ences we e ep esen ed by p <0.05. The esul s we e p esen ed as mean
±s anda d de ia ion (SD).
3. Resul s and discussion
3.1. Physicochemical p ope ies
Fig. 1 p esen s FESEM images and pa icle size dis ibu ion his o-
g ams o MBGNPs, illus a ing how he iming o calcium ni a e addi-
ion in luences nanopa icle mo phology and size dis ibu ion. The
FESEM images e eal ha he MBGNP_15min sample exhibi s a b oad
size dis ibu ion and p edominan ly non-sphe ical, elonga ed nano-
pa icles, wi h he highes aspec a io (1.44) among all samples. In
con as , o he MBGNP samples display a uni o m size dis ibu ion and
sphe ical mo phology, as con i med by he pa icle size dis ibu ion
his og am. In his s udy, he only a iable was he ime in e al be ween
TEOS and calcium ni a e addi ion, while all o he pa ame e s we e held
cons an . This ensu ed silica o ma ion began be o e calcium ni a e was
inco po a ed in all MBGNPs. The non-sphe ical shape o MBGNP_15min
is a ibu ed o insu icien eac ion ime o comple e pa icle o ma-
ion. Resul s indica e ha a minimum 30-minu e in e al be ween TEOS
and calcium ni a e addi ion is necessa y o achie e uni o mi y in
MBGNPs, consis en wi h indings om mic oemulsion-assis ed sol-gel
syn hesis s udies [10,14,31]. Addi ionally, he mean pa icle size o
MBGNPs inc eased wi h delayed calcium ni a e addi ion while main-
aining dispe si y and uni o mi y, excep o MBGNP_15min. A simila
end was obse ed in s udies on MCM-41, which epo ed a co ela ion
be ween eac ion ime and pa icle g ow h [32].
The chemical composi ions o MBGNPs a e summa ized in Table 1.
The analyzed SiO
2
/CaO a ios a e signi ican ly lowe han he nominal
alues (70SiO
2
–30CaO), due o calcium loss du ing he washing s eps
employed du ing syn hesis [19]. Ne e heless, he analyzed composi-
ions a e consis en wi h hose epo ed in he exis ing li e a u e o he
bina y SiO
2
–CaO sys em [10,29]. The CaO con en in MBGNPs is no ably
in luenced by he amoun o calcium ni a e deposi ed on he silica
nanopa icle su aces. This deposi ion p ocess is go e ned by elec o-
s a ic in e ac ions be ween he nega i ely cha ged Si–OH g oups on
silica and Ca
2+
ions [26]. When pa icle o ma ion is incomple e, silica
pa icles exhibi highe eac i i y, enhancing elec os a ic in e ac ions.
As p e iously epo ed, Ca
2+
ions ac as c oss-linke s be ween silica
pa icles [27]. Unde hese condi ions, an inc eased Ca
2+
deposi ion is
obse ed. Howe e , hese ions a e p one o agg ega ion due o hei
en apmen wi hin he MBGNP s uc u e, complica ing hei emo al
du ing washing s eps. This phenomenon explains he highe CaO con-
cen a ion and po en ial agglome a ion o Ca
2+
ions in MBGNP_15min,
which also exhibi s an i egula shape. The indings indica e ha ea lie
calcium ni a e addi ion enhances Ca
2+
deposi ion, leading o ion
en apmen and in luencing he chemical composi ion and mo phology
o MBGNPs. In con as , a sligh dec ease in CaO con en is obse ed
wi h delayed calcium ni a e addi ion. As shown in Fig. 1,
MBGNP_90min exhibi s he la ges pa icle size. La ge pa icles a e
epo ed o exhibi g ea e esis ance o elec os a ic in e ac ions
compa ed o smalle pa icles, wi h he addi ion o elec oly es such as
calcium sal s [33].
In he case o s uc u al cha ac e iza ion o silica e uni s, FTIR and
MAS-NMR echniques a e employed. The FTIR spec a o MBGNPs
(Fig. 2) show h ee cha ac e is ic bands: one a app oxima ely 443–451
cm
−1
co esponding o Si–O–Si bending ib a ions, ano he a 803 cm
−1
a ibu ed o Si–O symme ic s e ching ib a ions, and a las b oad
band be ween 1000 and 1250 cm
−1
assigned as asymme ic s e ching.
This mos in ense band has wo shoulde s due o ans e se op ical mode
(TO, 1050 cm
−1
) and longi udinal op ical (LO, 1250 cm
−1
) mode o
asymme ic s e ching ib a ion o he Si–O–Si bond in SiO
4
e ahed a.
These spec al ea u es a e consis en wi h hose epo ed o MBGNPs in
p e ious s udies [10,29].
The s uc u al uni s in MBGNPs a e desc ibed using he Q
Si
n
no a ion,
whe e "n" deno es he numbe o b idging oxygens (BO) pe SiO
4
e -
ahed on, which anges om 0 o 4 [34]. Q
Si
4
uni s can ans o m in o
Q
Si
3
, Q
Si
2
, Q
Si
1
, o Q
Si
0
uni s, p omo ing he depolyme iza ion o he silica e
ne wo k s uc u e a e CaO addi ion. This p ocess can be induced by
ne wo k-modi ying ca ions such as Ca
2+
o H
+
, which gene a e
non-b idging oxygens (NBOs) wi hin he s uc u e [35]. Fu he mo e,
he
29
Si MAS-NMR spec a we e pe o med on he MBGNP_15min and
MBGNP_90min samples, ep esen ing end-membe composi ions due o
he consis en CaO con en ac oss all MBGNPs. The asymme ic na u e o
Table 1
Chemical composi ion o MBGNPs (mol %) measu ed by WXRF.
Samples SiO
2
CaO
MBGNP_15min 85.8 (±0.5) 14.2 (±0.1)
MBGNP_30min 88.4 (±0.6) 11.6 (±0.2)
MBGNP_45min 88.9 (±0.4) 11.1 (±0.2)
MBGNP_60min 89.2 (±0.6) 10.8 (±0.2)
MBGNP_90min 90.5 (±0.5) 9.5 (±0.1) Fig. 2. FTIR Spec a o MBGNPs as a unc ion o calcium addi ion ime.
Cell iabili y (%) = (Abso bance o sample −Abso bance o blank)
(Abso bance o posi i e con ol −Abso bance o blank)×100 (1)
O. Bas¸ak e al.
Open Ce amics 23 (2025) 100807
4
he
29
Si MAS-NMR spec a equi ed a decon olu ion p ocess o de e -
mine he dis ibu ion o Q
Si
n
s uc u al uni s. Fig. 3 p esen s he expe i-
men al and decon olu ed spec a, while Table 2 p o ides he
co esponding peak assignmen s, wi h obse ed chemical shi s om
−110.49 o −110.30 ppm o Q
Si
4
and om −101.09 o −100.70 ppm o
Q
Si
3
uni s, o 15- and 90-minu es samples, espec i ely. The ull wid h a
hal maximum (FWHM) alues anged om 11.50 o 11 ppm o Q
Si
4
and
8 ppm o Q
Si
3
uni s. The esul s con i m ha he decon olu ion me hod
aligns wi h he p o ocol es ablished o MBGNPs by Leono a e al. [36].
In his app oach, he p ima y Q
Si
4
esonance cen e ed nea −110 ppm,
subs i u ing hyd oxyl g oups wi h BO a oms, esul ing in an inc ease in
he chemical shi o app oxima ely 8–12 ppm. The p esence o Q
Si
3
uni s
in MBGNPs indica es hyd oxyl g oups and BO a oms wi hin he s uc-
u e, sugges ing ha H
+
ca ions ac as ne wo k modi ie s like Ca
2+
ions.
Fig. 3.
29
Si MAS-NMR Spec a o a) MBGNP_15min and b) MBGNP_90min.
Table 2
Peak Assignmen s in
29
Si MAS-NMR spec a.
Sample Q
Si
4
Q
Si
3
δ (ppm) FWHM (ppm) Popula ion ( %) δ (ppm) FWHM (ppm) Popula ion ( %)
MBGNP_15 min −110.15 11 75.30 −100.95 8 24.70
MBGNP_90 min −110.50 11.85 77.54 −101.05 8 22.46
Fig. 4. TEM images o a) MBGNP_15min, b) MBGNP_30min, c) MBGNP_45min, d) MBGNP_60min and e) MBGNP_90min.
O. Bas¸ak e al.
Open Ce amics 23 (2025) 100807
5
In he MBGNP_15min sample, a highe popula ion o Q
Si
3
uni s han
MBGNP_90min sugges s a mo e open silica e ne wo k due o he g ea e
inco po a ion o CaO, as consis en wi h Table 1. Fu he mo e, he
29
Si
MAS-NMR spec a co ela es wi h he deg ee o ne wo k condensa ion,
whe e Q
Si
4
is he mos abundan uni , e lec ing apid silica ne wo k
condensa ion [37]. The delayed addi ion o calcium ni a e inc eases he
Q
Si
4
uni s, indica ing a mo e condensed and highly c oss-linked silica
ne wo k cha ac e ized by an inc eased numbe o siloxane b idges [38].
This obse a ion is consis en wi h indings ha ex ended eac ion imes
enhance ne wo k condensa ion [39].
3.2. Mic os uc u e o MBGNPs
Fig. 4 shows he TEM images o MBGNPs, illus a ing he in luence o
calcium ni a e addi ion iming on he pa icle mo phology and meso-
po osi y o he MBGNPs. The pa icles in he MBGNP_15min sample
exhibi a mo e i egula shape han o he MBGNPs, which is consis en
wi h he obse a ion om FESEM images. Addi ionally, he agglome -
a ion o Ca
2+
ions is mo e p onounced in he MBGNP_15min sample. A
no able ea u e ac oss all MBGNP samples is he p esence o al e na ing
g ey and pale s ipes in he TEM images. This pa e n sugges s ha he
pa icles a e o med h ough a laye -by-laye sel -assembly p ocess. This
s uc u al cha ac e is ic ag ees wi h p e ious s udies on MBGNPs, as
e e enced in sou ces [22,24].
The N₂ adso p ion-deso p ion iso he ms o MBGNPs a e shown in
Fig. 5. Fo MBGNP_15min, he adso p ion pa e n sugges s a Type VI
iso he m (Fig. 5, inse ), cha ac e ized by s epwise mul ilaye adso p ion
be ween 0 and 0.2 P/P
o
, consis en wi h he IUPAC classi ica ion o
non-po ous ma e ials [40]. In con as , o he MBGNP samples exhibi
Type IV iso he ms wi h Type 3 hys e esis loops, cha ac e is ic o mes-
opo ous ma e ials (2–50 nm) and indica i e o sli -shaped po es. A low
ela i e p essu es (0–0.1 P/P
o
), a sligh inc ease in N
2
adso p ion sug-
ges s he p esence o mic opo es (<2 nm) in all MBGNPs. A high ela-
i e p essu es (0.9–1.0 P/P
o
), N
2
up ake inc eases p opo ionally wi h
delayed calcium ni a e addi ion (MBGNP_45min, MBGNP_60min and
MBGNP_90min). This de ia ion om he expec ed ho izon al pla eau o
mesopo ous ma e ials indica es he p esence o mac opo osi y (>50 nm)
in hese samples. Consequen ly, i can be concluded ha all MBGNPs,
excep o MBGNP_15min, exhibi a combina ion o mic o-, meso‑, and
mac opo osi y. The po e size dis ibu ion u he con i ms he meso-
po ous na u e o he MBGNPs, wi h a p ima y peak a app oxima ely 3.7
nm and mino peaks be ween a ound 4.35 and 12.22 nm. Addi ionally, a
clea p opo ional ela ionship was iden i ied be ween he iming o
calcium ni a e addi ion and he o ma ion o a bimodal po e dis ibu-
ion. The de elopmen o la ge po es p o ides a no able ad an age by
imp o ing he ma e ial’s capaci y o accommoda e and anspo la ge
biomolecules [41]. Fo example, i has been epo ed ha e ec i e
p o ein loading in o mesopo es equi es po e sizes exceeding 5 nm [42].
As shown in Table 3, all MBGNPs wi h a delayed calcium ni a e
addi ion exhibi highe su ace a eas han MBGNP_30min. The speci ic
su ace a ea alue o MBGNP_30min a e co ela ed wi h alues epo ed
in he li e a u e [29,43]. No ably, he MBGNP_45min demons a es he
highes su ace a ea among all MBGNP samples, which can be a ibu ed
o a mo e p onounced hys e esis loop be ween app oxima ely 0.4 and
0.9 P/P
o
. This ea u e is indica i e o capilla y condensa ion wi hin
mesopo es. In con as , he speci ic su ace a ea o MBGNP_60min is he
lowes among all MBGNPs wi h delayed calcium ni a e addi ion
(MBGNP_45min, MBGNP_60min, and MBGNP_90min). The smalle po e
olume alue can explain his educ ion in su ace a ea obse ed o
MBGNP_60min compa ed o he o he samples.
The USAXS/SAXS absolu e calib a ed in ensi y p o iles o all
MBGNPs, analyzed using I ena so wa e [44], e eal h ee dis inc
sca e ing egions ac oss he q- ange o 1 Å
−1
o 10
−4
Å
−1
(Fig. 6a). All
p o iles a e also p esen ed in Fig. 6b. In he high-q egion (q >0.02 Å
−1
),
he sca e ing signal o igina es om nanopo es (<2 nm) wi hin indi-
idual pa icles, quan i ied using Po od’s law unde he assump ion o
sphe ical po e geome y. The in e media e q- ange (0.002–0.02 Å
−1
)
co esponds o sca e ing om p ima y pa icle ea u es. A he lowes
q- alues (q <0.002 Å
−1
), he obse ed powe -law sca e ing indica es
mass ac al a angemen s be ween pa icle clus e s, cha ac e is ic o
di usion-limi ed agg ega ion du ing syn hesis. The o al speci ic su ace
a ea alues p esen ed in Table 4 combine con ibu ions om in a-
g anula mic opo osi y (de i ed om high-q analysis) and in e pa icle
oid spaces quan i ied h ough medium-q modeling. The sphe ical po e
model applied o high-q da a demons a es a good co ela ion wi h N₂
physiso p ion ends (Fig. 5), alida ing he me hodology o meso-
po ous sys ems. MBGNPs syn hesized wi h delayed calcium ni a e
addi ion exhibi sys ema ically highe su ace a eas (15–20 % inc ease)
compa ed o MBGNP_30 min, accompanied by educed ac al dimen-
sionali y, sugges ing mo e open a chi ec u al a angemen s when
modi ying he p ecipi a ion sequence. No ably, MBGNP_15 min equi ed
exclusi e cha ac e iza ion h ough USAXS/SAXS measu emen s due o
Fig. 5. a) N
2
adso p ion-deso p ion iso he ms and b) po e size dis ibu ion (ob ained om deso p ion b anch o MBGNPs).
Table 3
Tex u al p ope ies o MBGNPs wi h N
2
adso p ion-deso p ion me hod. S
BET
:
Speci ic su ace a ea, V
P
: To al po e olume (deso p ion b anch).
Sample S
BET
(m
2
/g) V
P
(cm
3
/g)
MBGNP_30min 466.1 (±2.7) 0.45
MBGNP_45min 746.4 (±5.2) 0.74
MBGNP_60min 515.1 (±8.3) 0.69
MBGNP_90min 698.4 (±3.2) 0.87
O. Bas¸ak e al.
Open Ce amics 23 (2025) 100807
6
N₂ adso p ion’s inhe en limi a ions in de ec ing sub-2 nm po es and
po en ial mic opo e collapse du ing degassing p e ea men . Despi e
his me hodological di e gence, bo h echniques consis en ly iden i y
MBGNP_15min as possessing he lowes o al po osi y (Table 4), indic-
a i e o apid pa icle agg ega ion kine ics du ing i s ea lie -s age syn-
hesis. The combined USAXS/SAXS p o iles and N
2
adso p ion da a
es ablish ha pos poning calcium ni a e in oduc ion enhances po e
accessibili y while mode a ing ac al clus e densi y, p o iding c i ical
s uc u e-p ope y ela ionships o op imizing MBGNP design.
Modeling o egions 2 and 3 in USAXS/SAXS p o iles p o ides
aluable insigh s in o he spa ial a angemen o pa icles, wi h he e-
sul s summa ized in Table 5. The analysis assumes a Gaussian dis i-
bu ion o pa icle sizes, p ima ily sphe ical, excep o MBGNP_15 min,
whe e a sphe oidal pa icle dis ibu ion was applied. This adjus men
was necessa y due o he aspec a io o 1.44, de e mined om FESEM
images (Fig. 1), which indica ed sligh aniso opy in pa icle shape. The
local a angemen o g ains was u he cha ac e ized by wo key pa-
ame e s: he olume ac ion (φ), which e lec s he densi y o pa icle
packing, and he co ela ion leng h (
η
), which desc ibes he a e age
dis ance o e which s uc u al o de ing pe sis s. Fo MBGNPs syn he-
sized wi h delayed calcium ni a e addi ion, bo h φ and
η
alues
inc eased compa ed o ea lie -s age addi ions. This end indica es ha
he g ains we e dis ibu ed o e a b oade spa ial ange while main-
aining localized clus e ing o e longe dis ances. Speci ically, highe φ
alues sugges close packing o nea es neighbou s, while la ge
η
alues imply mo e ex ended o de ing ac oss he pa icle ne wo k.
Toge he , hese indings sugges ha delaying calcium ni a e addi ion
enhances bo h local densi y and long- ange o ganiza ion wi hin he
mesos uc u e. The compac ness o ac al agg ega es was e alua ed
using he pa ame e P, which e lec s he densi y o ac al clus e s.
Highe P alues indica e educed emp y space wi hin agg ega es, co -
esponding o dense packing. Howe e , no di ec co ela ion be ween P
alues and delayed calcium ni a e addi ion was obse ed. Ins ead, he
non-linea end in P alues wi h inc eased φ alues likely a ises om
he dynamic na u e o he O/W emulsion sys em used du ing syn hesis.
In his sys em, equen d ople collisions and andom B ownian mo ion
in luence he dis ibu ion o he oil phase, leading o a ia ions in
po osi y and g ain a angemen [23]. These s ochas ic p ocesses in o-
duce a iabili y in o agg ega e densi y, decoupling P om syn hesis
iming. These indings also highligh he diso de ed na u e o he po e
s uc u e in MBGNPs p oduced wi h he mic oemulsion-assis ed sol-gel
me hod [43]. The in e play be ween emulsion dynamics and sol-gel
chemis y c ea es me as able a angemen s ha a e e lec ed in he
s uc u al pa ame e s de i ed om USAXS/SAXS modelling. A sche-
ma ic 3D ep esen a ion o he modeling is p oposed in Figu e S1 o he
Supplemen a y In o ma ion.
To unde s and he unde lying mechanism o he delayed addi ion o
calcium ni a e, i is essen ial o emphasize ha he o al eac ion ime
has a mo e signi ican in luence on he p ope ies o MBGNPs han he
amoun o CaO inco po a ed. This is e iden om he subs an ial in-
c ease in su ace a ea and po e olume wi hou no able changes in
chemical composi ion. In a basic medium, po osi y is p ima ily go -
e ned by elec os a ic in e ac ions be ween posi i ely cha ged su ac-
an s (S
+
) and nega i ely cha ged silica clus e s (I
−
) [45]. Howe e , he
p esence o coun e ions (M
+
X
−
), such as sal s, can in e e e wi h hese
elec os a ic in e ac ions, po en ially al e ing he po osi y. Recen
s udies sugges ha high po osi y is p ima ily a ibu ed o a o able
elec os a ic in e ac ions be ween CTAB and silica in a basic medium
(pH >8), pa icula ly o la ge pa icles (>100 nm) whe e minimal pH
Fig. 6. (a) Schema ic ep esen a ion o dis inc egions obse ed in he USAXS/SAXS p o iles. (b) USAXS/SAXS p o iles o all MBGNP samples.
Table 4
Speci ic su ace a ea alues o MBGNPs de e mined by he USAXS/SAXS
me hod.
Sample S
o al
(m
2
/g) S
ine
(m
2
/g) Po e Volume (cm
3
/g)
MBGNP_15min 27 21.4 0.02
MBGNP_30min 34 32.5 0.03
MBGNP_45min 144 130.3 0.13
MBGNP_60min 125 107.2 0.15
MBGNP_90min 109 105.3 0.12
Table 5
Fi ing pa ame e s o egions 2 and 3 om USAXS/SAXS p o iles.
Sample Diame e
(nm)
Wid h
(nm)
P (uni less)
η
(nm) φ
MBGNP_15
min
151 22 2.34 84*1.36
*
MBGNP_30
min
175 26 1.90 126.5 2.62
MBGNP_45
min
185 39 1.80 180 3.50
MBGNP_60
min
180 38 1.63 225 3.44
MBGNP_90
min
175 37 1.92 255 4.00
*
=no eliable.
O. Bas¸ak e al.
Open Ce amics 23 (2025) 100807
7
d op occu s o e ime [46]. These in e ac ions a e c i ical o mesophase
o ma ion. Howe e , inco po a ing Ca
2+
ions can educe hese elec o-
s a ic in e ac ions, po en ially ac ing as a “blocking e ec ” ha impedes
mesophase de elopmen [47]. This supp ession e ec can be co ela ed
wi h TEM images (Fig. 4) and N
2
adso p ion-deso p ion esul s (Fig. 5).
No ably, MBGNP_15min exhibi s incomple e deso p ion beha io , clas-
si ying i as a non-po ous ma e ial based on N
2
adso p ion-deso p ion
iso he ms. Fu he mo e, USAXS/SAXS esul s con i m ha
MBGNP_15min has he lowes po osi y among all MBGNP samples. In
con as , delayed calcium ni a e addi ion (MBGNP_45min,
MBGNP_60min, and MBGNP_90min) esul s in mo e p onounced hys-
e esis loops in he N
2
adso p ion-deso p ion iso he ms. These loops
indica e capilla y condensa ion wi hin mesopo es and con ibu e o an
enhanced speci ic su ace a ea compa ed o MBGNP_30min. USAXS/-
SAXS p o iles con i m ha he delayed addi ion o calcium ni a e leads
o an inc ease in ine po osi ies, esul ing in highe speci ic su ace a ea
alues ha a e consis en wi h N
2
adso p ion-deso p ion iso he ms. This
obse a ion also sugges s ha Ca
2+
ions hinde mesophase o ma ion,
which co ela es wi h he inc eased S
ine
alues in MBGNPs wi h delayed
calcium ni a e addi ion compa ed o MBGNP_30min. Fu he mo e,
delayed calcium ni a e addi ion leads o he o ma ion a bimodal po e
dis ibu ion, as e idenced by he coexis ence o mesopo es and mac o-
po es, which is obse ed wi h N
2
adso p ion-deso p ion iso he ms.
Mac opo es become inc easingly appa en wi h delayed calcium ni a e
addi ion, as demons a ed in Fig. 1. This obse a ion aligns wi h he
b oade po e size dis ibu ion obse ed (Fig. 5). The p onounced o -
ma ion o mac opo es can be co ela ed wi h he USAXS/SAXS p o iles
in egions 2 and 3, as well as wi h he N
2
adso p ion-deso p ion iso-
he ms a high ela i e p essu es. Howe e , a disc epancy exis s be-
ween he speci ic su ace a ea alues o MBGNP_60min and
MBGNP_90min, as he esul s om USAXS/SAXS p o iles and N
2
adso p ion-deso p ion iso he ms do no ully co ela e. The highe spe-
ci ic su ace a ea ob ained om N
2
adso p ion-deso p ion o
MBGNP_90min compa ed o MBGNP_60min (Table 3) may be a ibu ed
o a g ea e p opo ion o ine po osi ies, as indica ed by he USAXS/-
SAXS measu emen s (Table 5). These indings u he suppo ha
MBGNPs exhibi bimodal po osi y, wi h he dynamic na u e o O/W
emulsions being mo e dis inc ly obse ed in USAXS/SAXS
measu emen s.
3.3. Ion elease beha io in T is/HCl bu e
The ion elease beha io o nanopa icles was s udied using T is/HCl
bu e a pH 7.4, as shown in Fig. 7. Among he samples, MBGNP_15min
exhibi ed he highes elease o Ca
2+
and Si
4+
a e 1 day o imme sion.
In e es ingly, he concen a ion o Si
4+
is simila a e 3 and 7 days o
incuba ion. The ion elease beha io o MBGNPs is p ima ily in luenced
by wo ac o s: speci ic su ace a ea and chemical composi ion [48].
Howe e , i s chemical composi ion (Table 1) shows he highes calcium
oxide con en (14 mol%), and
29
Si MAS-NMR esul s indica e a g ea e
numbe o NBOs. These ac o s likely con ibu e o i s accele a ed ion
elease. The MBGNP_30min and MBGNP_45min samples exhibi ed
simila ion elease beha io o bo h Ca
2+
and Si
4+
ions. Speci ically,
MBGNP_30min showed highe Si⁴⁺ elease a e 1 day o incuba ion, bu
a e 3 and 7 days, he silicon concen a ions became compa able. Fo
Ca
2+
elease, bo h samples demons a ed simila le els a e 1 and 3
days, wi h MBGNP_30min showing a sligh ly highe elease on day 7. In
con as , he MBGNP_60 min and MBGNP_90min samples exhibi ed
signi ican ly lowe ion elease o bo h ions han he o he samples. A e
7 days, bo h samples eleased simila concen a ions o calcium ions.
Howe e , MBGNP_90min showed a slowe elease a e. A simila end
was obse ed o Si
4+
ions, wi h MBGNP_90min showing an app oxi-
ma ely 25 % slowe elease a e han MBGNP_15 min.
The combined in luence o chemical composi ion and speci ic su -
ace a ea on ion elease beha io p esen s challenges o explici in e -
p e a ion due o hei simul aneous a ia ion [49]. The e ec o
chemical composi ion is pa icula ly e iden when compa ing he ion
elease a es o MBGNP_15min and MBGNP_90min. As shown in Table 3,
he MBGNP_15min has a highe numbe o NBOs and a less connec ed
silica ne wo k han MBGNP_90min, leading o accele a ed dissolu ion
despi e i s lowe su ace a ea. This obse a ion is consis en wi h ind-
ings epo ed by Schumache e al. [50]. Con e sely, he impac o spe-
ci ic su ace a ea is mo e p onounced when compa ing MBGNP_45min
and MBGNP_60min. Despi e hei nea ly iden ical chemical composi-
ions (Table 1), he as e ion elease obse ed o MBGNP_45min can be
a ibu ed o i s highe speci ic su ace a ea [51]. While MBGNP_30min
is a e e ence poin in his s udy, no able di e ences in ion elease
beha io a e e iden ac oss he MBGNPs. Despi e delayed calcium ni-
a e addi ion esul ing in highe speci ic su ace a eas (MBGNP_45min,
MBGNP_60min and MBGNP_90min), he as es ion elease among hese
samples was obse ed o MBGNP_30min. No ably, he highes ion
elease was obse ed o MBGNP_15min. This inding highligh s ha
su ace eac i i y plays a c ucial ole in ion elease beha iou alongside
speci ic su ace a ea and chemical composi ion. The ele a ed elease
a es obse ed o MBGNPs, such as MBGNP_15min and MBGNP_30min,
can be a ibu ed o hei highe le els and mo e homogeneous dis i-
bu ion o CaO on pa icles, as also con i med by USAXS/SAXS p o iles.
Fu he mo e, T is/HCl bu e main ains physiological pH condi ions
due o i s bu e ing capaci y; howe e , he ion elease beha iou o
nanopa icles may di e unde eal-li e condi ions, whe e he
Fig. 7. Ion elease p o iles o MBGNPs o a) Ca
2+
(mg/L) and b) Si
4+
(mg/L) ions in TRIS/HCl Bu e solu ion.
O. Bas¸ak e al.
Open Ce amics 23 (2025) 100807
8
en i onmen may p omo e p ecipi a e o ma ion o p o ein adso p ion
on he ma e ial’s su ace [52,53]. The e o e, i is impo an o conduc
ion elease s udies in a ious media, such as simula ed body luid o cell
cul u e medium, o be e unde s and he in e ac ions o he nano-
pa icles. Fu u e s udies should also e alua e he long- e m ion elease
kine ics in dynamic biological en i onmen s o mo e accu a ely simu-
la e in- i o condi ions and assess po en ial e ec s on bioac i i y and
biocompa ibili y.
3.4. In- i o biocompa ibili y
Fig. 8 p esen s he cell iabili y o MG-63 os eoblas -like cells
cul u ed o 48 h wi h dilu ed ex ac s o MBGNPs. Based on he ela i e
cell iabili y, all samples demons a ed non-cy o oxic beha io in
acco dance wi h he ISO 10,993–5 s anda d, which de ines ma e ials
wi h cell iabili y abo e 80 % as non-cy o oxic. The lowes cell iabili y
was obse ed a he 1 % w/ ex ac concen a ion o all MBGNPs.
These indings a e consis en wi h he s udy by Ku uldu e al. [29],
which e alua ed he cy ocompa ibili y o MBGNPs ex ac s based on a
bina y SiO
2
–CaO sys em. Thei esul s indica ed p omising cell iabili y
a ex ac concen a ions o 1 % w/ and lowe . Fo samples
MBGNP_15min, MBGNP_30min, and MBGNP_45min, an inc ease in cell
iabili y was obse ed wi h dec easing ex ac concen a ions. This
ou come highligh s he an icipa ed ole o Ca
2+
ions in enhancing
os eogenic cell p oli e a ion, such as MG-63, while he elease o Si
4+
ions u he enhance cell iabili y [54]. As shown in Fig. 6, he mos
eac i e ma e ial is MBGNP_15min, which exhibi ed he as es ion
elease du ing imme sion in TRIS/HCl bu e solu ion. Fo
MBGNP_45min, he highes cell p oli e a ion occu ed a an ex ac
concen a ion o 0.1 % w/ , sugges ing ha his concen a ion p o ides
an op imal mic oen i onmen o os eogenic cells, likely due o he
balanced he apeu ic ion elease. Con e sely, he lowes cell iabili y
was obse ed o MBGNP_60min and MBGNP_90min, which exhibi ed
he slowes ion elease a es. This educed iabili y may be a ibu ed o
insu icien elease o Ca
2+
and Si
4+
ions, essen ial o os e ing a
s imula o y en i onmen o os eogenic cells. Subop imal ion elease
likely esul s in less a o able condi ions o cell p oli e a ion [55].
O e all, MBGNPs wi h as e ion elease (MBGNP_15min,
MBGNP_30min, and MBGNP_45min) mo e e ec i ely suppo os eo-
genic cell p oli e a ion by p o iding adequa e le els o he apeu ic ions.
In con as , hose wi h slowe ion elease (MBGNP_60min and
MBGNP_90min) exhibi sligh ly educed cell iabili y, likely due o
insu icien Ca
2+
and Si
4+
ion elease, which diminishes hei
s imula o y e ec s on os eogenic cells.
4. Conclusion
This s udy demons a es ha he iming o calcium ni a e addi ion
conside ably impac s he po osi y o MBGNPs. Delayed addi ion o
calcium ni a e inc eased speci ic su ace a ea, po e size, olume, and
pa icle size. While he disc epancy be ween nominal and analyzed
composi ions was p ima ily a ibu ed o he washing s eps, he iming
o calcium ni a e addi ion also in luenced he chemical composi ion
due o pa icle enla gemen and la ge po e sizes. Despi e his, he
p opo ion o Q
Si
n
uni s sligh ly changed, indica ing minimal a ia ion in
he chemical composi ion ac oss he samples. The esul s o he s udy
indica ed ha he p esence o any deg ee o oxici y in os eoblas -like
cells was no obse ed among all MBGNPs. These indings sugges ha
adjus ing he iming o calcium ni a e addi ion is a simple ye e ec i e
s a egy o ailo ing he ex u al p ope ies o MBGNPs o a ious
biomedical applica ions. Addi ionally, a 45-minu e in e al be ween he
addi ion o TEOS and calcium ni a e seems op imal o p oduce high-
quali y MBGNPs, o e ing a aluable guideline o u u e esea ch.
CRediT au ho ship con ibu ion s a emen
Ona Bas¸ak: W i ing – o iginal d a , Visualiza ion, Me hodology,
In es iga ion, Fo mal analysis, Da a cu a ion, Concep ualiza ion. Fa ih
Ku uldu: W i ing – e iew & edi ing, Visualiza ion, Me hodology,
In es iga ion, Fo mal analysis, Concep ualiza ion. Jan Ila ský: W i ing
– e iew & edi ing, Visualiza ion, In es iga ion. Ma ina Vi ´
azko ´
a:
W i ing – e iew & edi ing, In es iga ion. Ana M. Bel ´
an: W i ing –
e iew & edi ing, In es iga ion. F ancisco Mu˜
noz: W i ing – e iew &
edi ing, Me hodology, In es iga ion. Yolanda Cas o: W i ing – e iew
& edi ing, Supe ision, Resou ces, In es iga ion. Ma in Mich´
alek:
W i ing – e iew & edi ing, Supe ision, Resou ces. Ma ia
Ch omˇ
cíko ´
a: W i ing – e iew & edi ing, Supe ision, Resou ces,
P ojec adminis a ion, Funding acquisi ion.
Decla a ion o compe ing in e es
The au ho s decla e ha hey ha e no known compe ing inancial
in e es s o pe sonal ela ionships ha could ha e appea ed o in luence
he wo k epo ed in his pape .
Fig. 8. Cell Viabili y esul s o MG-63 cells cul u ed wi h he ex ac o 1, 0.1 and 0.01 % cell cul u e medium dilu ions (*p <0.05).
O. Bas¸ak e al.
Open Ce amics 23 (2025) 100807
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