Abstract
The influence of mPEG-PCL and mPEG-PLGA on encapsulation efficiency and drug-loading of nanoparticles was very important. SN-38 NPs were prepared from a series of diblock copolymers: mPEG1000-PLGA2000, mPEG2000-PCLs, mPEG5000-PCLs, mPEG2000-PLGAs, and mPEG5000-PLGAs by the thin film-hydration method. The prepared nanoparticles were characterized by morphology, size, encapsulation efficiency, drug-loading, and in vitro release behavior. This experiment suggested that the encapsulation efficiency and drug-loading of SN-38 NPs were attained the maximum values when the ratio of hydrophilic to hydrophobic block was between 1:2 and 1:3.
Introduction
Irinotecan hydrochloride or camptothecin-11 (CPT-11) is a topoisomerase I (Topo I) inhibitor by trapping the enzyme during the cleavage of DNA (Hsiang et al. Citation1985). Chemically, it is a semisynthetic analog of the natural alkaloid camptothecin (Kunii et al. Citation2008). The 7-Ethyl-10-hydroxycamptothecin (SN-38), a biologically active anticancer drug, is derived from CPT-11 by carboxylesterases in the liver and in tumors, which is particularly effective against many malignancies including colorectal, lymphoma, lung, gastric, cervical, and ovarian cancer (Satoh et al. Citation1994, Slatter et al. Citation1997). Moreover, in vitro cytotoxicity studies indicate that SN-38 is up to 100- to 1000-fold more potent than CPT-11 against several tumor cell lines (Chabner and Longo Citation2001, Prasad and Danqi Citation2015). Nonetheless, the clinical use of SN-38 is limited by its hydrophobicity and instability at physiological pH. SN-38 is poorly soluble in aqueous solution, and is hardly insoluble in most physiologically compatible and pharmaceutically acceptable excipients, including polysorbate 80 and cremophor (Wadkins et al. Citation1999). Additionally, Crow et al. (Crow and Crothers Citation1992, Wani et al. Citation1987) show that a closed lactone ring moiety is an important structure for active drug interaction with the Topo-I target and antitumor potency. Rapidly opening the lactone ring of camptothecins and their derivatives will lead to the complete loss of biological activity (Crow and Crothers Citation1992). It is reported that SN-38 incorporated into nanoparticles can improve its solubility and stability (Ebrahimnejad et al. Citation2009, Ebrahimnejad et al. Citation2010, Ebrahimnejad et al. Citation2011, Gu et al. Citation2012, Vangara et al. Citation2013, Zheng et al. Citation2014).
Recently, nanoparticulate drug delivery systems have attracted much attention due to their unique accumulation behavior at the site of the tumor (Bawarski et al. Citation2008). Several nanocarriers have been reported, such as SN-38 conjugates (Bawarski et al. Citation2008, Ebrahimnejad et al. Citation2010, Ebrahimnejad et al. Citation2011, Williams et al. Citation2012), liposomes (Fu and Luan Citation2008, Pal et al. Citation2005, Zhang et al. Citation2004), and nanoparticles (Atyabi et al. Citation2009, Ebrahimnejad et al. Citation2009, Gu et al. Citation2012, Vangara et al. Citation2013, Wang et al. Citation2013, Zheng et al. Citation2014). The nanoparticulate carriers prepared by amphiphilic copolymers for the delivery of antitumor therapeutics have drawn much attention since 1990s with a large number of novel drug delivery systems developed (Li et al. Citation2009). In aqueous media, the amphiphilic lipids and polymers self-assemble to form nano-sized micelles with a hydrophobic core surrounded by a hydrophilic corona (Liu et al. Citation2006). In most cases, the hydrophilic block of micelles forming copolymers refers to polyethylene glycol (PEG) owing to its highly hydrophilic and biocompatible properties. Among biodegradable aliphatic polyesters, polyɛ-caprolactone (PCL) is one of the most attractive and promising hydrophobic blocks for its good biocompatibility, drug permeability, and nontoxicity (Valizadeh et al. Citation2016; Xie et al. Citation2007). In addition, polylactic-coglycolic acid (PLGA) nanoparticles have received intensive attention due to their biodegradability, biocompatibility, controllable particle diameters, well-described formulations, and sustained release behavior (Anari et al. Citation2015). Furthermore, PLGA has been approved by the European Medical Agency (EMA) and Food and Drug Administration (FDA) in drug delivery systems for parenteral administration (Vangara et al. Citation2013). Moreover, copolymers composed of PEG and PCL(mPEG-PCL), and PEG and PLGA (mPEG-PLGA), have been chosen as drug delivery vehicles in various biomedical applications. For example, SN-38-loaded Pluronic F-108 and PEG-b-PCL nanoparticles were successfully prepared and showed a significant enhancement of the SN-38 NPs drug efficiency in killing cancer compared with the free SN-38 (Gu et al. Citation2012). Naringenin-loaded MPEG-PCL nanoparticles and their formulation into buccal tablets exhibited an improvement of the solubility of naringenin and the treatment of oral inflammatory and ulcerative diseases (Wang et al. Citation2014). And, the SN-38-loaded PLGA-PEG-FOL nanoparticles demonstrated a cytotoxicity against HT-29 cancer cells (Ebrahimnejad et al. Citation2010). Vincristine sulfate-loaded PLGA-PEG-folate nanoparticles showed significant in vitro targeting effects for MCF-7 breast cancer cells, which resulted in enhanced cellular uptake and higher cytotoxicity in comparison with the free drug and PLGA-mPEG NPs (Zhao et al. Citation2008). However, there is still a lacking of studies on influence of different hydrophobic chain length, ratio of hydrophilic to hydrophobic segment, and different hydrophilic chain length on encapsulation efficiency and drug-loading of SN-38 NPs.
Herein, SN-38 nanoparticles (SN-38 NPs) were prepared from a series of diblock copolymers: mPEG1000-PLGA2000, mPEG2000-PCLs, mPEG5000-PCLs, mPEG2000-PLGAs, and mPEG5000-PLGAs. The prepared nanoparticles were characterized by morphology, size, encapsulation efficiency, drug-loading, and in vitro release behavior. The influence of different hydrophobic chain length, ratio of hydrophilic to hydrophobic segment, and different hydrophilic chain length on encapsulation efficiency and drug-loading of SN-38 NPs were systematically examined to exploit the potential actions of mPEG-PCL and mPEG-PLGA in the delivery system of anticancer drugs. This preliminary study attempts to provide more promising insights that the copolymer might act as an important factor for nanoparticle preparation.
Experimental
Materials
SN-38 (≥99.0%, 140205) was obtained from Jiangyuan Product Co. (Sichuan, China). mPEG1000-PLGA50/50 (molecular weight, Mw, 2000), mPEG2000-PLGA50/50 (Mw 1000, 2000, 4000, 5000, 9000, 18000), mPEG5000-PLGA50/50 (Mw 1000, 2000, 4000, 5000, 10000, 20000), mPEG2000-PCL (Mw 1140, 2000, 4000, 5300, 6000, 8000, 10000), and mPEG5000-PCL (Mw 1000, 2000, 4000, 5000, 10000, 15000, 20000, 45000) were purchased from Daigang Biotechnology Engineering Co. (Jinan, China). Pyrene (≥99.0%, 0001426999) was supplied by Sigma-Aldrich (Taufkirchen, Germany). Dichloromethane (DCM) and dimethyl sulfoxide (DMSO) were acquired from Damao Chemical Industry Co. (Tianjin, China). Acetone was from East Chemical Industry Co. (Tianjin, China).
Acetonitrile and methanol used as mobile phase in high-performance liquid chromatography (HPLC) were obtained from Fisher Scientific (Fair Lawn, NJ). All other chemicals used were of analytical grade.
Preparation of SN-38-loaded NPs
SN-38-loaded NPs were prepared by the thin film-hydration method. Briefly, 1 mg of SN-38 and 20 mg of different polymer were dissolved in the mixture of DCM and acetone (1:4, V/V) and mixed by a magnetic stirrer (Yuhua Apparatus Co., Gongyi, China) at room temperature for 1 h. The mixture was transferred into a suitable round- bottom flask and dried in a rotary evaporator (RE-2000, Yarong Biochemical Analyzer Industry, Shanghai, China) to form a homogeneous film. The dry film was maintained overnight under air to remove the residue of DCM and acetone. Then the film was hydrated with 5 mL of water at 260 W using a probe sonication (XO-650, Xian’ou Instrument manufacturing Co., Nanjing, China) for 2.5 h at 2-s pulse in ice bath. The nanoparticle dispersion was extruded through 0.22-μm polyamide filter (Yibo Filtering Equipment Factory, Haining, China). The filtered formulation was then centrifuged at 3000 × g for 5 min (Centrifuge 5804 R, Eppendorf, Germany) to separate the unloaded material from nanoparticles. Here, SN-38-loaded nanoparticles produced from mPEG2000-PCL1000 and mPEG5000-PLGA1000 were named SP2C1 and SP5G1, respectively, and so on, for each of nanoparticles.
HPLC assay
The concentration of SN-38 was analyzed by HPLC system (Agilent 1200, USA) with a UV detector (265 nm). The chromatographic separations were achieved using an Agilent Zorbax SB-C18 column (4.6 × 250 mm, pore size 5 μm) at 30 °C. The mobile phase consisted of a 35:20:45 (v/v/v) mixture of acetonitrile: methanol: Na2HPO4 buffer (pH 3.8) at a flow rate of 1.0 mL/min. The calibration curve of SN-38 was linear over the range of 0.05–10 μg/mL with R2 >0.999, and no interference was observed with mPEG-PLGA and mPEG-PCL at 265 nm.
Particle size and zeta potential
The particle size, size distribution, polydispersity, and zeta potential of the prepared SN-38 NPs were measured by Zetasizer Nano ZS-90 (Malvern Instruments, Malvern Worcestershire, U.K.). All the analyses were carried out in triplicate.
Measurement of morphology
The morphology of SN-38 NPs was examined using transmission electron microscopy (TEM). A drop of SN-38 NPs was stained with 2% phosphotungstic acid and dried on copper grids for 24 h before SN-38 NPs were visualized under TEM (H-7500, Hitachi, Tokyo, Japan).
Encapsulation efficiency and drug-loading
Encapsulation efficiency and drug-loading of SN-38 NPs were determined by filtration. Free SN-38 mainly existed as a precipitation, which could be directly removed by 0.22-μm filters. Furthermore, free SN-38 dissolved in water could be ignored compared with encapsulated SN-38 because SN-38 is almost insoluble in water (7.2 μg/mL; Chen et al. Citation2011). Unfiltered and precipitated SN-38 was dissolved in DMSO and determined by HPLC. On the other hand, SN-38 encapsulated in the nanoparticles was diluted by methanol and mixed for 1 min before HPLC determination.
The percentage of the encapsulation efficiency and drug-loading of SN-38 NPs were calculated as follows:
In vitro drug release
In vitro release of SN-38 from nanoparticle formulation was analyzed by membrane dialysis against phosphate-buffered saline (PBS, pH 7.4, with 30% ethanol) at 37 °C. Briefly, an 1-mL aliquot of SN-38 NPs was placed in the dialysis tube (Green Bird Science dialysis tubes with a Mw cutoff of 14,000 Da) and then suspended in an erlenmeyer flask containing 150 mL of PBS. The flask was then put in a water bath shaker (SHA-C, Hualong Experimental Apparatus Industry, China), which was maintained at 37 °C and shaken horizontally at 100 rpm. At 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 5, 6, 8, 10, 12, 24, 48, 72 h, an aliquot of 4 mL was withdrawn from the flask and replaced with the same amount of fresh release medium; 10 μL of the formic acid was added into 1 mL of the aliquot and mixed for 5 min. The concentration of SN-38 was assayed by the HPLC described previously after 0.5 h.
Measurement of the critical micelle concentration of mPEG-PCL and mPEG-PLGA
The critical micellar concentration (CMC) of mPEG-PCL and mPEG-PLGA nanomaterials was investigated by fluorescence spectroscopy using pyrene as fluorescence probe (Zhang et al. Citation2014). The fluorescence spectrum was recorded on a fluorescence spectrophotometer (F-4600, Hitachi, Tokyo, Japan). Aliquots of pyrene solution (6 × 10 − 5 mol/L in acetone, 100 μL) were added to 10-mL volumetric flasks, and acetone was allowed to evaporate under a stream of nitrogen. A series of aqueous mPEG-PCL and mPEG-PLGA solutions ranging from 10 − 2 to 10 − 6 mg/mL were added into the flasks to give a final pyrene concentration of 6 × 10 − 7 mol/L. The solutions were sonicated 30 min and then placed overnight at room temperature before measurements. The excitation spectra were recorded from 200 to 400 nm when emission was carried out at 390 nm. Both excitation and emission slit widths were 10 nm. The intensity ratios (I337/I303) were analyzed as a function of amphiphilic copolymer concentrations. CMC values were estimated as the intersection point of the tangent to the curve at the inflection with the horizontal tangent through the points at low concentration.
Results and discussion
Influence of PCL chain length of mPEG-PCLs on the encapsulation efficiency and drug-loading of SN-38 NPs
To investigate the effect of the PCL chain length, a series of nanoparticles were produced from diblock copolymers of mPEG2000-PCLs in which the Mw of the PEG chain was kept constant (2000), and the Mw of the PCL chain was gradually increased from 1140 to 2000, 4000, 5300, 6000, 8000, and up to 10000. As shown in and , and , the nanoparticles possessed different physicochemical characteristics. They were nearly spherical in shape, uniformly distributed; the mean diameter of mPEG2000-PCLs nanoparticles was in the range of 155–250 nm; The samples exhibited zeta potentials between −2.29 and −12.57 mV; with augmenting of PCL chain length, encapsulation efficiency and drug-loading increased first and then decreased, in which the maximum were 83.21% (mPEG2000-PCL5300) and 3.37% (mPEG2000-PCL6000) for encapsulation efficiency and drug-loading, respectively.
Figure 1. Morphological characteristics of SN-38 NPs containing mPEG2000-PCL1140 (A), mPEG2000-PCL2000 (B), mPEG2000-PCL4000 (C), mPEG2000-PCL5300 (D), mPEG2000-PCL6000 (E), mPEG2000-PCL8000 (F), mPEG5000-PCL1000 (G), mPEG5000-PCL2000 (H), mPEG5000-PCL4000 (I), mPEG5000-PCL5000 (J), mPEG5000-PCL10000 (K), mPEG5000-PCL15000 (L), mPEG2000-PLGA2000 (M), mPEG2000-PLGA4000 (N), mPEG2000-PLGA5000 (O), mPEG5000-PLGA1000 (P), mPEG5000-PLGA2000 (Q), mPEG5000-PLGA4000 (R), mPEG5000-PLGA5000 (S), mPEG5000-PLGA10000 (T), and mPEG5000-PLGA20000 (U), respectively.
Figure 2. Profiles of encapsulation efficiency and drug-loading of SN-38-loaded nanoparticles with different hydrophobic segments of mPEG2000-PCLs.
Table 1. Physicochemical characteristics of SN-38-loaded nanoparticles with different hydrophobic segments of mPEG2000-PCLs.
A batch of mPEG5000-PCLs, where the PEG Mw was kept constant (5000) and the Mw of the PCL block was progressively increased from 1000 to 2000, 4000, 5000, 10000, 15000, 20000, and at the most 45000, were applied to form diverse particles of mPEG5000-PCL1000, mPEG5000-PCL2000, etc. The basic physicochemical characteristics of the prepared nanoparticles were summarized in . The nanoparticles distributed uniformly with spherical structure seen from TEM micrographs. The mean diameters of mPEG5000-PCLs nanoparticles were in the range of 161–346 nm as shown in . The zeta potentials were between −4.13 and −10.45 mV. Accompanied by PCL chain length extending, the encapsulation efficiency and drug-loading first increased and finally decreased; and the former reached a peak value of 80.71% at mPEG5000-PCL10000 and the latter reached a peak value of 3.19% at mPEG5000-PCL15000, as shown in .
Figure 3. Profiles of encapsulation efficiency and drug-loading of SN-38-loaded nanoparticles with different hydrophobic segments of mPEG5000-PCLs.
Table 2. Physicochemical characteristics of SN-38-loaded nanoparticles with different hydrophobic segments of mPEG5000-PCLs.
This indicated that the encapsulation efficiency and drug-loading contents increased with the increase of the PCL length on the condition that the Mw of PCL was less than 6000 at PEG2000 and 15000 at PEG5000. Diab and coworkers also found that the longer the PCL block chain in the mPEG-PCL copolymer used in the microparticle, the higher the encapsulation efficiency of cytarabine, when all the other preparation conditions were fixed (Diab et al. Citation2010). However, the encapsulation efficiency and drug-loading content decreased later when the Mw of PCL block was increased from 6000 to 10000 and 15000 to 45000 with the PEG block controlled in 2000 and 5000, respectively.
It was reported that drug-loading capacity of nanoparticles was affected by the critical micellar concentration (CMC; Shuai et al. Citation2004a, Shuai et al. Citation2004b), partition coefficient (Kozlov et al. Citation2000, Liu et al. Citation2006), Flory–Huggins interaction parameter (χsp) (Patel et al. Citation2008), hydrophobic–lipophilic balance (HLB; An et al. Citation2015) and crystalline phases of hydrophobic chains (Shuai et al. Citation2004a). Herein, the CMC was determined using pyrene as a hydrophobic fluorescent probe. HLB was calculated as follows (Wen et al. Citation2014):
The CMC and HLB values of copolymers were shown in and , respectively. It was showed that CMC and HLB values of mPEG-PCL decreased with the length of hydrophobic chain increasing, which led to the increase of encapsulation efficiency and drug-loading of nanoparticles. Moreover, the length of the hydrophobic block could also influence the partition coefficient (Hurter et al. Citation1993). If the hydrophilic block was kept constant, an increase in the Mw and length of the hydrophobic block had been found to increase the partition coefficient for a particular micelle system (Kozlov et al. Citation2000, Liu et al. Citation2006) which resulted in an increased drug-loading capacity per micelle (Allen et al. Citation1999). In addition, the increased Mw of PCL chains could decrease χsp values, which resulted in higher drug solubility in PEO-b-PCL micelles when PCL chains were longer (Patel et al. Citation2008). Nevertheless, PCL chains in the micelle cores with crystalline state would decrease the loading content of doxorubicin-loaded micelles because amorphous phase was much more likely to accommodate drug molecules; crystallization ability of PCL chains was obviously improved by the increasing of PCL length in the diblock copolymers (Shuai et al. Citation2004a, Xie et al. Citation2007). It is suggested that encapsulation efficiency and drug-loading of SN-38 NPs first increased and then decreased with longer PCL length here, which can be explained by CMC, χsp, HLB, and crystallization state of different PEG-PCLs.
Table 3. CMC values of different copolymers.
Table 4. HLB values of different copolymers.
Influence of PLGA chain length of mPEG-PLGAs on the encapsulation efficiency and drug-loading of SN-38 NPs
In regards to the effect of the PLGA chain length, SN-38-loaded nanoparticles were prepared with assorted mPEG2000-PLGAs where the Mw of the PLGA block was increased from 1000 to 2000, 4000, 5000, 9000, and 18000, respectively, but that of PEG block was kept constant 2000. exhibited that the shape of the SN-38 NPs was spherical and they were uniformly distributed.
summed up physicochemical characteristics of these particles. described that the mean diameter of mPEG2000-PLGA nanoparticles was in the range of 173–274 nm. The zeta potentials were between −10.70 and −16.43 mV. depicted that with the increasing of PLGA chain length, the encapsulation efficiency and drug-loading primarily increased, secondarily decreased, and further got 79.59% and 3.36%, respectively, at mPEG2000-PLGA5000.
Figure 4. Profiles of encapsulation efficiency and drug-loading of SN-38-loaded nanoparticles with different hydrophobic segments of mPEG2000-PLGAs.
Table 5. Physicochemical characteristics of SN-38-loaded nanoparticles with different hydrophobic segments of mPEG2000-PLGAs.
Plus, another kinds of SN-38 NPs were also produced by mPEG5000-PLGAs, where the Mw of the PEG block was remained at 5000 and the Mw of the PLGA block was increased from 1000 to 2000, 4000, 5000, 10000, and 20000. The TEM images of SN-38 NPs were shown in , which were nearly spherical in shape and uniformly distributed. The basic physicochemical properties of these particles were summarized in . The mean diameter of mPEG5000-PLGA nanoparticles was in the range of 134–272 nm (). Their zeta potentials were between −4.27 and −12.10 mV. With the increasing of PLGA chain length, at the beginning the encapsulation efficiency and drug-loading increased, decreased in the end, and their highest values were 76.54%, and 2.93%, respectively at mPEG5000-PLGA10000 ().
Figure 5. Profiles of encapsulation efficiency and drug-loading of SN-38-loaded nanoparticles with different hydrophobic segments of mPEG5000-PLGAs.
Table 6. Physicochemical characteristics of SN-38-loaded nanoparticles with different hydrophobic segments of mPEG5000-PLGAs.
The above-mentioned findings demonstrated that the encapsulation efficiency and drug-loading contents increased with the increase of the PLGA length when the Mw of PLGA was less than 5000 and that of PEG block stayed at 2000; the similar phenomena also occurred when the Mw of PLGA was less than 10000 and that of PEG block stayed at 5000. But yet, the encapsulation efficiency and drug-loading contents had decreased when the Mw of PLGA block was being increased to 9000 from 5000 at PEG2000 and to 20000 from 10000 at PEG5000.
It was inferred that the increasing of PLGA length could cause the decrease of CMC (), HLB (), and χsp values and the increase of partition coefficient, which contributed to the increase of the encapsulation efficiency and drug-loading contents of SN-38 NPs. This resembling occurrence was repeatedly observed in SN-38 NPs prepared with PEG-PLGAs or PEG-PCLs. In addition, it was reported that denser particles with larger PLGA blocks drew together the PEG blocks, leading to condensed PEG blocks with high PEG volume fraction in the PEG/water phase (Yang et al. Citation2015), which explained the fall in the encapsulation efficiency and drug-loading of SN-38 NPs in the rear parts of and .
Influence of different ratio of hydrophilic to hydrophobic segment on the encapsulation efficiency and drug-loading of SN-38 NPs
The encapsulation efficiency and drug-loading of SN-38 NPs were practically independent of the ratio of the diblock copolymer. The ratio of hydrophilic to hydrophobic segment on encapsulation efficiency and drug-loading of SN-38 NPs were shown in . It was observed that encapsulation efficiency and drug-loading of SN-38 NPs were changed with different ratio of hydrophilic to hydrophobic block. On the one hand, for mPEG-PCLs, the maximum encapsulation efficiency was 83.21% at mPEG2000-PCL5300, and the maximum drug-loading was 3.37% at mPEG2000-PCL6000; on the other hand, for mPEG-PLGAs, the encapsulation efficiency and drug-loading reached a top value of 79.59% and of 3.09%, respectively at PEG2000-PLGA5000. These results demonstrated that the maximum encapsulation efficiency and drug-loading were observed when the ratio of hydrophilic to hydrophobic block was between 1:2 and 1:3. It indeed confirmed that ratio of hydrophilic to hydrophobic block can significantly exert impact on the encapsulation efficiency and drug-loading of nanoparticles.
Influence of PEG chain length of mPEG-PLGA2000s on the encapsulation efficiency and drug-loading of SN-38 NPs
To identify the influence of the hydrophilic block (PEG) of mPEG-PLGAs on encapsulation efficiency and drug-loading, SN-38 NPs were prepared from mPEG1000-PLGA2000 (data were not shown), mPEG2000-PLGA2000, and mPEG5000-PLGA2000, respectively. The mean diameter of mPEG-PLGA2000 nanoparticles was in the range of 173–230 nm. The samples exhibited a zeta potential between −9.61 and −14.57 mV. The encapsulation efficiency and drug-loading of SN-38 NPs were not obviously changed as the length of hydrophilic PEG block increased when the Mw of PLGA block was remained at 2000. This indicated that changes in the length of the PEG block had no significant effect on the encapsulation efficiency and drug-loading of SN-38 NPs. Lee and coworkers (Lee et al. Citation2003) found that changes in the length of the hydrophilic block of copolymer had no significant effect on the CMC. Patel et al. (Citation2008) indicated that an increase in the Mw of the PEO block (same PCL Mw) did not almost affect the χsp values. Hence, the encapsulation efficiency and drug-loading had been hardly mediated by the length of the PEG block because of PEG-unaffected CMC and χsp values.
In vitro drug release
Release profiles of SN-38 from NPs were shown in . More than 77% and 95% of loaded SN-38 were released from nanoparticles prepared from mPEG2000-PCLs and mPEG5000-PCLs, respectively in PBS at 37 °C at 72 h. And the releasing rate of SN-38 from nanoparticles formed from mPEG2000-PLGAs and mPEG5000-PLGAs in PBS at 37 °C was larger than 93% and 89%, respectively at 72 h.
Figure 6. In vitro release curve of SN-38-loaded nanoparticles with different molecular weight of hydrophobic segments (mPEG2000-PCL1140, mPEG2000-PCL2000, mPEG2000-PCL4000, mPEG2000-PCL5300, mPEG2000-PCL6000, and mPEG2000-PCL8000).
Figure 7. In vitro release curve of SN-38-loaded nanoparticles with different molecular weight of hydrophobic segments (mPEG5000-PCL1000, mPEG5000-PCL2000, mPEG5000-PCL4000, mPEG5000-PCL5000, mPEG5000-PCL10000, and mPEG5000-PCL15000).
Figure 8. In vitro release curve of SN-38-loaded nanoparticles with different molecular weight of hydrophobic segments (mPEG2000-PLGA2000, mPEG2000-PLGA4000, and mPEG2000-PLGA5000).
Figure 9. In vitro release curve of SN-38-loaded nanoparticles with different molecular weight of hydrophobic segments (mPEG5000-PLGA1000, mPEG5000-PLGA2000, mPEG5000-PLGA4000, mPEG5000-PLGA5000, mPEG5000-PLGA10000, and mPEG5000-PLGA20000).
Additionally, there were no significant effects of different hydrophobic and hydrophilic chain length, ratio of hydrophilic to hydrophobic segment on the release of SN-38.
It has been reported that the lactone ring of SN-38 hydrolyses to give pharmacologically inactive carboxylate form at pH >6 (Chen et al. Citation2011). Actually, an acidic pH promoted the formation of the active lactone ring; 100% of SN-38 carboxylate was converted back to the SN-38 lactone form when pH was adjusted to 3 (Zhang et al. Citation2004), as shown in . Here, formic acid was used to transform an inactive carboxylate form, which mainly exist in PBS (pH 7.4), into an active α-hydroxyl-δ-lactone ring analyzed by HPLC.
Figure 10. pH-dependent equilibrium of SN-38.
Conclusion
In this study, SN-38-loaded nanoparticles were prepared from different diblock copolymers of mPEG-PCLs and mPEG-PLGAs of different molecular weights and ratio by the thin-hydration method. The PCL and PLGA Mw were varied from 1000 to 45,000, when the PEG Mw was kept constant (2000 or 5000). It was thus possible to investigate the influences of the hydrophobic block and the ratio of hydrophilic to hydrophobic block, as well as the hydrophilic block, on encapsulation efficiency and drug-loading of SN-38 NPs. It was observed that the encapsulation efficiency and drug-loading increased first and then decreased, with the increasing of PCL and PLGA chain length. Our data suggested that the encapsulation efficiency and drug-loading of SN-38 NPs were changed with different ratios of hydrophilic to hydrophobic block, and attained the maximum values when the ratio of hydrophilic to hydrophobic block was between 1:2 and 1:3. The length of the PEG block had no significant effect on the encapsulation efficiency and drug-loading of SN-38 NPs. In conclusion, the encapsulation efficiency and drug-loading of nanoparticles can be impacted by hydrophobic chain length and ratio of hydrophilic to hydrophobic segment, which is attributable to the alterations of CMC, HLB, χsp, and crystallization state.
Funding information
This work was supported by Natural Science Foundation of Ningxia Province, China (NZ13171).
Disclosure statement
The authors report no declarations of interest.
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