Food Chemistry 172 (2015) 99–104

Contents lists available at ScienceDirect

Food Chemistry
journal homepage: www.elsevier.com/locate/foodchem

Short communication

Validation of an Ultraviolet–visible (UV–Vis) technique

for the quantitative determination of curcumin
in poly(L-lactic acid) nanoparticles
Rosana Aparecida da Silva-Buzanello a, Ana Caroline Ferro b, Evandro Bona a, Lúcio Cardozo-Filho c,
Pedro Henrique Hermes de Araújo d, Fernanda Vitória Leimann a, Odinei Hess Gonçalves a,⇑
a
Federal University of Technology – Paraná (UTFPR), Post-Graduation Program of Food Technology (PPGTA), Caixa Postal 271, BR 369, km 0.5, CEP 87301-006 Campo Mourão,
PR, Brazil
b
Federal University of Technology – Parana (UTFPR), Food Technology and Engineering Department (COEAL), Caixa Postal 271, BR 369, km 0.5, CEP 87301-006 Campo Mourão,
PR, Brazil
c
Maringá State University (UEM), Department of Chemical Engineering, Av. Colombo 5790, CEP 87020-900 Maringá, PR, Brazil
d
Federal University of Santa Catarina (UFSC), Department of Chemical and Food Engineering (EQA), Caixa Postal 476, Campus Reitor João David Ferreira Lima,
CEP 88040-970 Florianópolis, SC, Brazil

a r t i c l e i n f o a b s t r a c t

Article history: Curcumin is a natural yellow-orange pigment extracted from turmeric and is a potential substitute of
Received 20 February 2014 health-dangerous artificial dyes. Nanoencapsulation in biodegradable polymers is a promising alternative
Received in revised form 24 July 2014 to improve curcumin stability and water solubility but curcumin concentration inside the nanoparticles
Accepted 5 September 2014
must be precisely known. A reliable method to determine the actual curcumin concentration must be val-
Available online 16 September 2014
idated since the validation procedures warrant that the method is adequate and sufficient for the specific
application involved. This work describes the validation parameters given by the International Confer-
Keywords:
ence on Harmonisation (ICH) guidelines to adopt an analytical method based on Ultraviolet–visible spec-
Analytical validation
Nanoencapsulation
troscopy for the quantitative determination of curcumin encapsulated in poly(L-lactic acid) nanoparticles.
Curcumin This method was validated in respect to linearity, detection limit, quantification limit, accuracy and pre-
Nanoparticles cision. Studies on the analytical procedure validation warranted safety in final results obtained for the
PLLA curcumin concentration in the nanoparticles.
Miniemulsification Ó 2014 Elsevier Ltd. All rights reserved.

1. Introduction 2010), high performance liquid chromatography coupled to a mass

Curcumin is a yellow-orange phytochemical compound pre-
senting a polyphenolic structure and low molar mass (368.37 g/
gmol) that can be isolated from Curcuma longa. It is a diferuloylme-
thane with several phenolic groups and conjugated double bonds,
and has remarkably hydrophobic character (Basniwal, Buttar, Jain,
& Jain, 2011). Curcumin is used as a spice, flavouring and food dye
and its bioactivity has been extensively reviewed (Hatcher,
Planalp, Cho, Torti, & Torti, 2008; Maheshwari, Singh, Gaddipati,
& Srimal, 2006; Pari, Tewas, & Eckel, 2008).
Several methods have been proposed to determine the concen-
tration of curcumin and its derivatives in spices or in bulk drug and
pharmaceutical dosage forms such as Ultraviolet–visible spectros-
copy (UV–Vis) (Jasim & Ali, 1988, 1992; Sharma, Agrawal, & Gupta,
2012), ultra performance liquid chromatography (Cheng et al.,
⇑ Corresponding author. Tel.: +55 44 3518 1478.
E-mail address: odinei@utfpr.edu.br (O.H. Gonçalves).
spectrometer (Li et al., 2011), thin layer chromatography (Zhang
et al., 2008), high performance liquid chromatography (Koop,
Freitas, Souza, Martinez, & Silveira, 2013). Although each one
presents its advantages, UV–Vis is often used to the curcumin
determination on most encapsulation systems since it is fast
enough and can give reliable results.
Despite the increasing interest in the properties of curcumin, its
application is still limited by its low water solubility and its insta-
bility in alkaline conditions, thermal treatment, light, metallic ions,
enzymes, ascorbic acid and others (Lin, Lin, Chen, Yu, & Lee, 2009;
Paramera, Konteles, & Karathanos, 2011). The lack of bioavailability
is also an issue when in natura curcumin is administrated (Mishra,
Mohammada, & Mishra, 2008). Polymer nanoparticles and micro-
particles have been extensively studied aiming to improve the sta-
bility of several compounds and drugs. Many results have been
reported in increasing curcumin bioavailability and protection
against external conditions during processing and storage. In the
case of curcumin, encapsulation is a promising alternative to

http://dx.doi.org/10.1016/j.foodchem.2014.09.016
0308-8146/Ó 2014 Elsevier Ltd. All rights reserved.
100 R.A. da Silva-Buzanello et al. / Food Chemistry 172 (2015) 99–104
improve its application and there are a number of different meth-
ods to obtain nanoencapsulated curcumin such as cross-linking
(Bisht et al., 2007); emulsification and cross-linking followed by
solvent removal (Lertsutthiwong, Noomun, Jongaroonngamsang,
Rojsitthisak, & Nimmannit, 2008); emulsion (Shaikh, Ankolab,
Beniwal, Singha, & Kumar, 2009); nanoprecipitation (Anand et al.,
2010); ionotropic pre-gelation followed by polycationic cross-link-
ing; solvent emulsion–evaporation (Dandekar et al., 2010); ionic
cross-linking reaction (Anitha et al., 2011); production of micelles
by solid dispersion (Basniwal et al., 2011); and electrohydrody-
namic atomization (Gomez-Estaca, Balaguer, Gavara, &
Hernandez-Munoz, 2012). In all cases the encapsulant must be
carefully chosen since it directly affects the encapsulation effi-
ciency. Miniemulsification/solvent evaporation is a suitable tech-
nique to encapsulate hydrophobic compounds like curcumin and
presents the advantage that a wide range of encapsulants can be
used. The process consists of an organic phase composed of sol-
vent, polymer (the encapsulating agent) and the compound to be
encapsulated, while the aqueous phase that consists of distilled
water. A single surfactant or a mixture is added in the organic or
the aqueous phases as required. The organic phase is then dis-
persed in the water phase using high energy equipment such as
a sonicator. The solvent is then removed by extraction or
evaporation and the nanoparticles are formed (Leimann et al.,
2012; Musyanovych, Schmitz-Wienke, Mailänder, Walther, &
Landfester, 2008;). However, in all encapsulation methods some
amount of the encapsulated substance may be lost during the pro-
cedure steps depending on the interactions with the encapsulant
polymer and on the system stability. In the case of water dispersed
nanoparticles the amount of encapsulated substance is also influ-
enced by its partition between aqueous and organic phase and
by the encapsulant-substance interactions. The actual curcumin
concentration in the nanoparticles after the encapsulation process
must be known since loss or degradation may also occur during the
encapsulation process. However, most works present curcumin
concentration results but analytical validation is not mentioned.
A reliable analytical method must be employed so that the cur-
cumin concentration may be determined (Blanco, Campaiia, &
Barrero, 1996; Korany, Haggag, Ragab, & Elmallah, 2013; Nevado,
Cabanillas, & Salcedo, 1994) and the methodology may be vali-
dated (ICH, 2005). Validation is a continuous process that starts
at the planning of the analytical strategy and continues throughout
the method development and transference (Alves, Rolim, Fontes, &
Rolim-Neto, 2010). In the case of patenting new products, valida-
tion of analytical methodologies is also required. The validation
of an analytical procedure should warrant that the method meets
the requirements of analytical applications and ensures the reli-
ability of the results. According to the World Health Organization
Expert Committee on Specifications for Pharmaceutical Prepara-
tions (WHO, 1992), the systematic evaluation of an analytical pro-
cedure should demonstrate that it has the conditions for which it
must be applied.
The objective of this work was to validate the analytical method
based on Ultraviolet–visible spectroscopy to determine the actual
curcumin concentration in curcumin-loaded poly(L-lactic acid)
(PLLA) nanoparticles. Validation of the method was carried out as
per the International Conference on Harmonisation (ICH) guide-
lines Q2 (R1) (ICH, 2005).
2. Material and methods
2.1. Materials
Curcumin (Sigma–Aldrich, 99.5%), dichloromethane (Sigma–
Aldrich, 98%) and methanol (Sigma–Aldrich, 99.5%) were used as
received. Poly(L-lactic acid) (PLLA, Mw = 3780 g/mol) was synthe-
sized from L-lactide (Purac) by ring opening polymerization as
described by Bendix (1998) using tin octanoate (Sigma–Aldrich,
99%) as catalyst (10,000 molL-lactide/molcatalyst). Distilled water and
soybean lecithin (Alfa Aesar) were used as the continuous phase
and surfactant, respectively. Solutions were filtered through Milli-
poreÒ nylon membrane filters (diameter 0.45 lm) and centrifuged
in Amicon filters (100 kDa).
2.2. Analytical validation
Linearity, detection limit, quantification limit, accuracy and pre-
cision were evaluated following the procedures described by the
International Conference on Harmonisation (ICH, 2005) using an
Ultraviolet–visible spectrophotometer (PG Instruments, T70+)
operating with 1 nm spectral bandwidth, ±0.3 nm wavelength
accuracy and a splitbeam optical system. This equipment was used
in all experiments unless otherwise stated.
2.2.1. Linearity
Linearity is the capacity to obtain results directly proportional
to the analyte concentration in the sample within a determined
range (ICH, 2005; USP, 1999; WHO, 1992). Linearity was investi-
gated using curcumin solutions in dichloromethane and methanol
1:1 (v/v) in six different concentration levels ranging from
4 mg L1 to 40 mg L1. Investigations were carried out on three dif-
ferent days using the Ultraviolet–visible spectrophotometry tech-
nique (UV–Vis) at 465 nm.
2.2.2. Detection limit
Detection limit (DL) is defined as the minimum amount at
which the analyte may be detected and is often determined by
the analysis of samples with known analyte concentrations (ICH,
2005). DL must be determined from the linear equation of the cal-
ibration curve by using the angular coefficient (b) and the standard
deviation (s) calculated from seven blank samples according to Eq.
(1)(ICH, 2005).
3:3
DL ¼ s  ð1Þ
b
In Eq. (1), the number 3.3 is a constant chosen according to the
required confidence level indicating the probability of false posi-
tives a, and false negatives b, equal to 5% (95% confidence)
(Currie, 1995).
2.2.3. Quantification limit
Quantification limit (QL) is a parameter of quantitative assays
for low levels of compounds in the sample. It indicates the lowest
analyte concentration in a sample that may be calculated with pre-
cision and accuracy (ICH, 2005). QL was determined by the angular
coefficient (b) from the calibration curve and the standard devia-
tion (s) calculated from seven blank samples according to Eq. (2).
10
QL ¼ s  ð2Þ
6
According to the IUPAC recommendations (Currie, 1995), QL is
expressed in terms of relative standard deviation RSD, which is
the maximum tolerated value conventionally equal to 10%. The
constant used to calculate the detection limit is performed accord-
ing to the limit (kQ = 1/RSD), with a resulting value of 10.
2.2.4. Precision
The precision of an analytical procedure expresses the concor-
dance between a series of measurements from multiple samplings
of the same homogenous sample (Currie, 1995; ICH, 2005).
 R.A. da Silva-Buzanello et al. / Food Chemistry 172 (2015) 99–104
Table 1
Formulations of encapsulated curcumin nanoparticles.
Experimental condition Distilled water (g)
Formulation 1 22.4815
Formulation 2 22.4552
Formulation 3 22.4649
The intermediate precision (assays on different days), repeat-
ability (different assays on the same day) and reproducibility
(assays involving a different analyst and a different laboratory)
were determined with six scans of three curcumin solutions with
known concentrations (4, 20 and 40 mg L1). Precision levels were
calculated by the relative standard deviation percentage (RSD%)
from the analytical curves. All UV–Vis spectra were recorded using
PG Instruments equipment (model T70+) in the Laboratory of Food
Analysis (Federal University of Technology – Paraná). For the
reproducibility evaluation, a Hitachi UV–Vis spectrophotometer
was used (model U-1900, 4 nm spectral bandwidth, ±0.5 nm
wavelength accuracy and splitbeam optical system). This equip-
ment is in the Control Process Laboratory (Federal University
of Santa Catarina) and the experiments were carried out by the
same analyst.
2.2.5. Accuracy (recovery method)
Accuracy of an analytical procedure demonstrates a concor-
dance degree between the value (accepted as conventionally true
or an accepted reference value) and the obtained value (ICH,
2005). Accuracy was determined by preparing curcumin solutions
with known concentrations at three different levels as proposed by
Grillo et al. (2009). Curcumin solutions were prepared in dichloro-
methane/methanol (1:1, v/v) with concentrations of 4, 20 and
40 mg L1, which corresponds to the minimum, medium and max-
imum values of the previously determined standard calibration
curve. The samples were analyzed and concentrations were recal-
culated from the calibration curve. Assays were performed in trip-
licate on two different days.
2.2.6. Specificity
Specificity was evaluated by obtaining the UV–Vis spectra of
curcumin and the PLLA nanoparticles (without curcumin). The
objective was to guarantee that no interference occurred in the
absorbance region as a reference for curcumin quantification
according to the International Conference on Harmonisation (ICH,
2005). Curcumin was dissolved in dichloromethane:methanol 1:1
(v/v) to yield a 40 mg L1 solution. Additionally, 1 mL of PLLA
nanoparticles dispersion without curcumin was dried (70 °C) and
then 1 mL of dichloromethane and 1 mL of methanol were added.
The solution was filtered (Millipore membrane filter, 0.45 lm)
and1 mL was then diluted in dichloromethane:methanol 1:1 (v/v)
to 10 mL.
2.2.7. Statistical analyses
Calibration curve was evaluated by regression analysis (ANOVA,
adjusted determination coefficient and standard error of estimate)
along with the residual plots. The effects of analyst and laboratory
were evaluated by one-way ANOVA. The effects of concentration,
day ant their interaction were tested by factorial ANOVA. All anal-
yses were carried out using Statistica 7.1 (StatSoft, 2005).
2.3. Preparation of the curcumin-loaded PLLA nanoparticles
Nanoparticles were produced by the miniemulsification/solvent
evaporation technique following Musyanovych et al. (2008) and
Leimann et al. (2012) with some modifications as follows. PLLA
101
Lecithin (g) Dichloromethane (g) Curcumin (g) PLLA (g)
0.1800 11.7140 0.0040 0.3924
0.1796 11.7161 0.0115 0.3940
0.1799 11.7590 0.0237 0.3935
and lecithin were dissolved in dichloromethane for 5 min.
Curcumin was then added and mixed for 5 min. After this, distilled
water was added under moderate stirring and the system was son-
icated (Fisher-Scientific Ultrasonic Dismembrator 120 W with a 1/
800 tip) at 100% amplitude for 180 s in a pulsing regime (30 s soni-
cation and a pause of 10 s). The miniemulsions were poured into
250 mL Erlenmeyer flasks and the solvent was evaporated at
40 °C for 18 h. Table 1 presents the formulations of curcumin-
loaded PLLA nanoparticles.
2.4. Determination of the actual curcumin concentration in the
nanoparticles
1 mL of the nanoparticles dispersion was dried immediately
after their preparation in a circulation oven at 70 °C for 2 h. The
dried aliquots were then diluted in 1 mL dichloromethane and
the polymer was precipitated by adding 1 mL methanol. The solu-
tion was filtered with a Millipore membrane (0.45 lm) and 1 mL
was then diluted in dichloromethane:methanol 1:1 (v/v) to adjust
the absorbance. The procedure was also carried out for nanoparti-
cles produced without curcumin and the resulting solution was
used as blank in the UV–Vis analysis. Absorbance was determined
at 465 nm.
2.5. Nanoparticle characterization
Morphological characterization of the nanoparticles was per-
formed using Transmission Electron Microscopy (TEM; JEOL model
JEM-1011, 100 kV). Samples without dilution were dripped onto
300-mesh parlodium-copper grids. The grids were dried at room
temperature and stained with osmium tetroxide for 4 h.
3. Results and discussion
3.1. Analytical validation
UV–Vis spectra of pure curcumin and PLLA nanoparticles with-
out curcumin are presented in Fig. 1.
No signal at 465 nm was detected when the nanoparticles were
evaluated meaning that the method is specific to curcumin at this
absorbance value. This wavelength was then selected for the cali-
bration curve. Specificity is an important parameter because over-
lapped curves must be resolved before analysis. In some cases,
extraction or separation procedures must be necessary which
Fig. 1. UV–Vis spectra for: (a) pure curcumin; (b) PLLA nanoparticles (without
curcumin).
102 R.A. da Silva-Buzanello et al. / Food Chemistry 172 (2015) 99–104

increases the complexity of the analytical determination (Blanco Table 2

et al., 1996; Nevado et al., 1994; Zhang et al., 2008). Specificity is Accuracy study for the analytical validation of the curcumin determination method.

often ignored by authors which can lead to overestimated results Concentration Sample Concentration Average of Recovery
(Jasim & Ali, 1988). of added (day) of curcumin curcumin rate mean
The calibration curve found was ‘‘absor- curcumin found concentrations (%)
(mg L1) (mg L1) found (confidence
bance = 0.055 + 55.236 ⁄ Curcumin (mg mL1)’’. Angular and linear (mg L1)* interval
coefficients of the calibration curve were 55.236 ± 1.072 mL mg1 95%)*, 
and 0.055 ± 0.026, respectively. The correlation coefficient and 4 1a 3.11 3.09 77.21 ± 0.55a
the adjusted correlation coefficient of the straight line were 1b 3.11 RSD = 0.66% RSD = 0.68%
0.997 and 0.994, respectively, and the standard error of estimate 1c 3.09
was 0.058 meaning that linearity can be assumed. The residuals 2a 3.09
2b 3.07
presented normal distribution and no outliers were observed.
2c 3.06
Residuals are sections of data variability unexplained by adjust-
20 1a 19.54 19.46 97.28 ± 0.71b
ment. They can be interpreted with an error estimate and should
1b 19.37 RSD = 0.69% RSD = 0.70%
be normally and independently distributed around zero. In fact, 1c 19.27
they should present mean zero and constant variance. The analysis 2a 19.66
of variance showed that the calculated F value (2657.365) was 2b 19.45
higher than that of tabulated F at a significance level of 1% 2c 19.45

(8.531) and the probability of lack of linearity is lower than 106. 40 1a 38.64 38.53 96.32 ± 0.37b
The limit of detection was determined as 0.05 mg L1, meaning 1b 38.46 RSD = 0.37% RSD = 0.37%
1c 38.35
that this is the minimum level at which the analyte can be detected 2a 38.73
by the method. The limit of quantification found was 0.10 mg L1. 2b 38.43
Jasim and Ali (1992) used a spectrofluorometric method to deter- 2c 38.55
mine curcumin concentration in dry acetone solutions and found
RSD = relative standard deviation.
the limit of detection to be 0.34 lg L1. However, this method is *
n = 6.
very sensitive to temperature and to moisture present in the sol-  
Different letters in the same row indicate statistical differences (p > 0.05).
vent (acetone). Blanco et al. (1996) determined the curcumin con-
centration in mixtures of the food colourants using the derivative
spectrophotometric resolution technique. They found a linear recoveries from 84% to 97% depending on the curcumin concentra-
response from 1.2 to 20 mg L1 (R2 = 0.9995) and a limit of detec- tion and a relative standard deviation (RSD) of 4.37%. Jasim and Ali
tion of 1.23 mg L1. Cheng et al. (2010) validated a UPLC method (1988) studied curcumin concentrations of 1 and 15 mg L1 and
to determine curcumin concentration. They found the limit of found RSD percentages of 2.67% and 2.01%. The same authors
detection and the limit of quantification to be as low as 40.66 (Jasim & Ali, 1989) found RSD percentages for curcumin in 12 polar
and 134.38 picograms which is expected since UPLC is more sensi- solvents from 1.87% to 2.89%. An overall recovery percentage for
tive than UV–Vis spectroscopy. Nevado et al. (1994) found a deter- 3 mg L1 curcumin was found to be 99.2%, whereas that for
mination limit of 0.126 mg L1 and a linear response up to 9 mg L1 was 100.8%.
9 mg L1. Tables 3 shows the results for intermediate precision (inter-
Recovery rates and accuracy were calculated using the data pre- day), repeatability (intra-day), reproducibility (intra-laboratory)
sented in Table 2. and accuracy.
Recovery rate values varied from 97.28% to 77.2% and RSD from Reproducibility values for intermediate precision (inter-day)
0.37% and 0.69% meaning that good results were obtained. The and repeatability (intra-day) presented relative standard deviation
effect of curcumin concentration was statistically significant lower than 1.5%. Shaikh et al. (2009) evaluated the validation of a
(p < 0.05) and the effect of ‘‘day x concentration’’ interaction was methodology based on High-Performance Liquid Chromatography
not significant (p > 0.05). Ribani, Bottoli, Collins, and Jardim (HPLC) to determine the amount of encapsulated curcumin in
(2004) stated that an acceptable recuperation rate depends on poly(lactic acid-co-glycolic) by and obtained relative standard
the objective of the analysis. Acceptable intervals are generally deviation values for intermediate precision and repeatability lower
between 70% and 120% depending on the sample preparation than 3%. Grillo et al. (2009) also validated an HPLC based method
and on the analytical procedure. Blanco et al. (1996) presented to determine the amount of encapsulated benzocaine in

Table 3
Precision levels (intermediate precision, repeatability and reproducibility) for the analytical validation of the curcumin determination method.

Curcumin concentration (ppm) Laboratory I Laboratory II Reproducibility

1st day 2nd day Inter-day (n = 6) 1st day 2nd day Inter-day (n = 6)
RSD(%) RSD(%) RSD(%) RSD(%) RSD(%) RSD(%) RSD(%)
4 1.36 0.49 0.93 0.47 0.54 0.51 0.72
20 1.01 0.44 0.73 0.74 0.97 0.86 0.79
40 0.36 0.28 0.32 1.02 0.34 0.68 0.50
Analyst I Analyst II
1st day 2nd day Inter-day (n = 6) 1st day 2nd day Inter-day (n = 6)
RSD(%) RSD(%) RSD(%) RSD(%) RSD(%) RSD(%)
4 1.36 0.49 0.93 0.63 1.30 0.97 0.95
20 1.01 0.44 0.73 1.42 1.31 1.37 1.05
40 0.36 0.28 0.32 1.63 1.24 1.44 0.88

RSD: relative standard deviation (%)

 R.A. da Silva-Buzanello et al. / Food Chemistry 172 (2015) 99–104 103

Fig. 2. TEM image of the PLLA–curcumin nanoparticles and water dispersions: (A) PLLA nanoparticles (blank sample), (B) curcumin-loaded nanoparticles and (C) free
curcumin.

The capability of being dispersed in water is one of the most

Table 4
Curcumin concentration in the PLLA nanoparticles. important properties of the nanoparticles because it affects their
final application. Fig. 2 demonstrates that the nanoparticles (A
Formulation Added Curcumin Curcumin in the nanoparticles
 100 ð%Þ
Added curcumin and B) can be readily dispersed in water forming visually homoge-
curcumin concentration
(ppm) in the neous systems. It is worth noting that the samples (B) and (C) have
nanoparticles the same curcumin concentration (500 mg L1). Free curcumin was
*
(ppm) poorly dispersed in water and formed macroscopic aggregates due
1 173 170 ± 2 98.3 to its high hydrophobicity. Similar results were reported by Shao
2 499 491 ± 6 98.4 et al. (2011) for curcumin encapsulated in polyethylenoglycol
3 1028 689 ± 33 67.0 and poly(caprolactone) nanoparticles and also by Anitha et al.
*
Determined by the calibration curve. (2011) for curcumin encapsulated in ortho-carboxymethyl chito-
san nanoparticles.

poly(hydroxybutyrate-co-hydroxyvalerate). Relative standard 4. Conclusion

deviation for intermediate precision and repeatability was lower
than 2%.
The high specificity of the UV–Vis spectra of curcumin when
compared to the PLLA nanoparticles, the linearity of the calibration
curve and the precision values presented above indicate that the
proposed UV–Vis analytical method can be considered adequate
to determine the actual amount of curcumin encapsulated in the
poly(L-lactic acid) nanoparticles.
3.2. Nanoparticle characterization
Curcumin concentration in the nanoparticles is presented in
Table 4 and Fig. 2 shows a TEM image of the curcumin-loaded
nanoparticles (Formulation 2). Fig. 2 also presents curcumin-
loaded nanoparticles (1 mL), pure curcumin (5 mg) and PLLA nano-
particles (1 mL) dispersed in water.
Nanoparticles showed spherical shape with diameters of
approximately 200 nm (Fig. 2). The curcumin concentration in
the nanoparticles in formulations 1 and 2 presented values close
to the added curcumin concentration. The same was not observed
when high amounts of curcumin was added (Formulation 3). The
difference between added and actual curcumin concentrations is
not related to the proposed analytical method since it was vali-
dated for the absorbance range. This suggests that the system
remained stable throughout the nanoencapsulation without great
losses only when low amounts of curcumin were added.
Dandekar et al. (2010) produced curcumin nanoparticles encapsu-
lated in hydrogel hydroxyl propylmethylcellulose and poly(vinyl
pyrrolidone), also by using the technique of mini-emulsification/
solvent evaporation. The authors found values of encapsulation
efficiency of 72%, indicating a lower affinity of curcumin and the
encapsulants used. Yallapu, Gupta, Jaggi, and Subhash (2010)
encapsulated curcumin in poly(lactic-co-glycolic acid) by nanopre-
cipitation the technique of using stabilizers such as poly (vinyl
alcohol) and poly(L-lysine) and obtained encapsulation efficiency
between 49.6 ± 4.5% and 89.5 ± 3.2%.
An Ultraviolet–visible spectroscopy method to determine the
concentration of curcumin on biodegradable nanoparticles was
validated in respect of specificity, linearity and inter-day, intra-
day, inter-laboratory and inter-analyst precision. The method is
adequate to quantify the presence of curcumin in the nanoparticles
and presented a detection limit and limit of quantification of 0.04
and 1 mg L1, respectively.
Curcumin was encapsulated in poly(L-lactic acid) nanoparticles
and the curcumin losses were detected when high amount of cur-
cumin was initially added. The resulting curcumin-loaded nano-
particles could be completely dispersed in water, which was not
the case of free curcumin.
Acknowledgements
The authors would like to thank the financial support of CAPES
and CNPq and to LCME – Federal University of Santa Catarina for
TEM images.
Appendix A. Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.foodchem.2014.
09.016.
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