,Ying Hung,b, Xin Pn,＊,Qinghe Yo,Chunbin Wu,b

aSchool of Pharmaceutical Sciences,Sun Yat-sen University,Guangzhou 510006,China

bGuangDong Research Center for Drug Delivery Systems,Sun Yat-sen University,Guangzhou 510006,China

cSchool of Engineering,Sun Yat-sen University,Guangzhou 510006,China

dGuangzhou Neworld Pharm.Co.Ltd.,Guangzhou 510006,China

Studies on the spray dried lactose as carrier for dry powder inhalation

Linna Wua,1,Xu Miaoa,1,Ziyun Shana,Ying Huanga,Lu Lia,b, Xin Pana,＊,Qinghe Yaoc,＊,Ge Lid,Chuanbin Wua,b

aSchool of Pharmaceutical Sciences,Sun Yat-sen University,Guangzhou 510006,China

bGuangDong Research Center for Drug Delivery Systems,Sun Yat-sen University,Guangzhou 510006,China

cSchool of Engineering,Sun Yat-sen University,Guangzhou 510006,China

dGuangzhou Neworld Pharm.Co.Ltd.,Guangzhou 510006,China

A R T I C L E I N F O

Article history:

Received 31 March 2014

Received in revised form

14 July 2014

Accepted 14 July 2014

Available online 27 August 2014

Spray drying

The purpose of this study was to investigate the spray dried lactose as carrier for dry powder inhalation(DPI).The lactose particles were prepared by spray drying,then the particle size,shape and crystal form were characterized by laser diffraction,scanning electron microscopy(SEM),X-ray diffraction(XRD)and differential scanning calorimetry (DSC).The spray dried lactose particles were spherical and amorphous,but would transfer to crystal form when storage humidity was above 32%.Thus,the humidity of the storage environment should be controlled below 30%strictly in order to maintain the amorphous nature of spray dried lactose which is a great benef i t to DPI development.

? 2014 Shenyang Pharmaceutical University.Production and hosting by Elsevier B.V.All rights reserved.

1. Introduction

Inhalation of drugs is a form of administration that is actuated by patient to deliver drugs to the therapeutic site and to achieve local or systemic effect[1-5].The enormous surface area of the lung and the plentiful capillary vessels conduce to a rapid absorption rate for inhalation;meanwhile,the absorbed drug can directly go into the blood circulation to effectively avoid the elimination by the f i rst-pass metabolism in the liver [1,5,6].

Dry powder inhalation(DPI)is a dosage form that delivers the micronized drug particles to the lung in the assistance of carrier particles by the inspiration of patient.Compared with the traditional pMDI,DPI requires no propellant,has superior chemical stability and has good patient compliance[1,5,7,8], thus now it has been a hotspot in the research of inhalation.

To date,almost all marketed DPI products rely on jetmilled,micronized drugs[9].However,this method showed many def i ciencies including little control of particle size, shape and surface morphology,and highly cohesive powders produced with poor aerosolization properties[10].Spray drying technique is one of the methods to prepare medicinal particles intended for lung delivery using dry powder inhalers [2,11,12].The principle of spray drying is to disperse drug solution into very small droplets by the atomizer,then rapidly evaporate the solvent in a hot dry medium like hot air to obtain dry product of powder or granules.This process produces product with high degree of purity and narrow particle size distribution,and also is relatively easy to scale up for commercial production[2].

However,there are still some disadvantages for spray drying,such as,particles would be sucked by the airf l ow leading to low product yield and causing loss especially for expensive drugs;some drugs are not stable at elevated temperature and tend to degrade.To overcome these shortcomings,a widely used formulation approach is to blend drugs with carriers which can provide protection to drugs and then to process co-spray drying[13,14].Fig.1 showed the carrierbased and drug/excipient spray dried DPI dosage forms.Simply mix the drugs and carriers is the most favorite method for preparing DPI dosage forms due to the easy operation and avoidingthedrugloss duringthe spraydrying[3,6,8].The ideal carriers for DPI should have low density,well size distribution, good spherical degree,f l owability and reproducibility[6,9].For DPI carrier,the size acts a subsidiary role for the drug separate itself form the carrier and only the active ingredient deposits in the pulmonary[6,11].

Lactose is the only authorized carrier to be used in dry powder inhalation by FDA mainly because of its wellinvestigated toxicity prof i le and broad availability[2,15]. Commonly crystal form α-lactose monohydrate is used in marketed DPI products.The amorphous lactose,which could be manufactured by the spray drying,could increase the stability of drugs and the effectiveness of drugs delivering to the lung and absorbing in the blood circulation when acting as DPI carrier[18].The transformation of amorphous lactose to crystalline was a quick process especially in high humidity condition.The purpose of this article is to investigate characteristics of spray dried lactose in order to provide useful reference resources for future studies using this material as carrier for dry powder inhalation.

2. Materials and methods

2.1. Materials

α-lactose monohydrate was obtained from Meggle(Batch No: F20090032,Germany).Anhydrous α-lactose was obtainedfrom DMV Ltd.(SuperTab 20AN,Batch No:CR190021,Holland). Anhydrous β-lactose was obtained from DMV Ltd.(SuperTab 21AN,Batch No:10493390,Holland).

2.2. Spray drying

Microparticles of lactose were prepared by spray drying.Precalculated amount of lactose was solubilized in water to obtain lactose concentration of 5%(w/v).Then the solution was spray dried using a laboratory scale spray dryer(SD-1000 Spray Dryer,Eyela,Japan)with the following conditions:inlet temperature at 120°C,atomizing pressure of 180 kPa,air f l ow of 0.7 m3/min and solution feed rate at 3 rpm.Three batches of samples were prepared and stored in desiccator(under 25% relative humidity and room temperature)before use.

2.3. Particle size

Particle size was measured by laser diffraction(Malvern Mastersizer 2000,Malvern Instruments Ltd.,UK)using the Scirocco dry dispersion unit at a feed pressure of 4 bars and feed rate of 50%.All samples were analyzed in triplicate with the obscuration values between 0.5%and 5%.

2.4. Particle morphology

Particle morphology of spray dried lactose and α-lactose monohydrate was visualized by scanningelectronmicroscopy (SEM)(JSM-6330F,Japan)at 15 kV.Samples were mounted on carbon sticky tabs and gold-coated before imaging.

2.5. Crystal structure

The crystal structure of various forms of lactose was analyzed using X-ray diffraction(XRD)(D-MAX 2200 VPC,Rigaku Ltd., Japan)withCu-Kα asX-raysource.Settingswereasfollows:scan rate of 5°/min,voltage of 40 kV,current of 30 mA and scan range of 5-55°2θ.

Fig.1-The carrier-based and drug/excipient spray dried for DPI dosage forms.

Table 1-Production of spray dried lactose with particle size determined by laser diffraction and moisture content determined by thermogravimetric analysis(n=3).

Differential scanning calorimetry(DSC,STA-409,Netzsch, Germany)was simultaneously used to study the crystallinity of the samples.Approximately 5-10 mg of samples was accurately weighed into 40 μL aluminium pans.Samples in closed pans were run at a heating rate of 10°C/min from 30°C to 300°C under nitrogen purge of 20 ml/min.

2.6. Moisture content

Thermogravimetric analysis(TG/SDTA 851e,Mettler-toledo, German)was conducted on samples to determine the moisture content adsorbed on the surface of particles that could inf l uencetheparticle-particle interaction[16].About 5-10 mg of each sample was put into an alumina pan and was heated from 25 to 120°C at a heating rate of 10°C/min under a nitrogen atmosphere.

2.7. Effect of storage

Approximately 100 mg of spray dried lactose was laid uniformly in a glass vial with external diameter of 25 mm and height of 25 mm,the vials then were stored in sealed containers with a series of humidities created by different salt supersaturated solutions:32%RH,43%RH,53%RH,65%RH, 75%RH,85%RH at room temperature in duplicate.After 3 d, three samples were taken out from each container to study the crystallinity transformation.After 7 d,the same research was conducted on the rest samples.

100 mg of spray dried lactose sample was accurately measured on an analytical balance and put into the vial as described above.In each of desiccators with humidity set as above,three vials were put in to evaluate the moisture uptake of samples during storage.To avoid the inf l uence caused by moisture uptake of glass vial,the vials were pre-equilibrated in desiccators for at least 24 h.After 7 d of storage,all the vials were taken out to determine the weight changes.

3. Results and discussion

3.1. Production of spray drying

Table 1 summarized the production of three batches of spray dried lactose.It can be seen that the yield of spray drying was between 43%and 62%,the geometric diameter of the particles was in a range of 24-35 μm and the moisture content was around 6%.

3.2. Particle morphology

Representative electron micrographs of spray dried lactose were shown in Fig.2A-B.Generally,the spray dried particles appeared to be spherical in shape with smooth surface.In comparison,the SEM of α-lactose monohydrate in Fig.2C showed particles with an irregular morphology and a wide size distribution.

3.3. Crystallinity for different forms of lactose

As the most commonly used excipient for the formulation of DPI,lactose hasthree forms:α-lactose monohydrate,β-lactose and anhydrous lactose.Crystallinity of lactose with different forms was checked by DSC and the scans were shown in Fig.3.

Fig.3A was the DSC scan of commercial grade anhydrous lactose with a melting endotherm at 240°C indicating the existing of β-lactose.A slight depression at approximately 140°C suggested there might be a small amount of monohydrate in the sample.Fig.3B was the DSC pattern of lactose monohydrate with an endothermic peak at 140°C induced by dehydration and the melting endotherm around 220°C.In contrast,after spray drying,lactose became amorphous as evidenced by the exothermic peaks as shown in Fig.3C and this was in agreement with the previous report[17].

Commonly crystal form α-lactose monohydrate is used in marketed DPI products,however,it has been reported that the amorphous state of carriers can increase the stability of drugs and the effectiveness of drugs delivering to the lung and absorbing in the blood circulation[18].Nevertheless,theamorphous lactose is not stable which would transfer to αlactose monohydrate and β-lactose[11,19].Theuse of any new excipient in a DPI development would require full characterization and understanding of its functionality[20],and that is the main purpose of this work.

Fig.2-SEM images of spray dried lactose and a-lactose monohydrate.(A,B is spray dried lactose and C is a-lactose monohydrate).

Fig.3-DSC patterns of lactose with different forms. (A:anhydrous lactose;B:lactose monohydrate;C:spray dried lactose).

Analysis of XRD patterns for lactose before and after spray drying were shown in Fig.4.Fig.4B for spray dried lactose showed a signif i cant reduction in magnitude of XRD peaks compared with Fig.4A for lactose monohydrate and this clearly indicated the amorphous form of spray dried samples.

3.4. Crystallinity transformation under humidity

The reduction of particle size of spray dried products brings abouttheincreaseofspecif i csurfaceareas whichwouldresult in the increase of hydrophilism.Moisture is an important factor that can signif i cantly inf l uence the behavior of inhalation for DPI.It can act as plasticizing agent,change the surface of the particles and promote strong adhesion or agglomeration[2,21].Also,the existing of water would inf l uence the crystallinity of spray dried samples[2].Therefore,studying the inf l uence of environmental humidity on the crystallinity transformation of spray dried amorphous lactose becomes very important.

Fig.4-XRD patterns of lactose with different forms. (A:lactose monohydrate;B:spray dried lactose).

Fig.5-DSC patterns of spray dried lactose.(A:freshly spray dried sample;B-G:samples stored for 3 d under RH 32%,43%,53%,65%,75%,85%).

DSC and XRD researches were carried out on spray dried lactose samples which were stored under different humidities for 3 or 7 d.

The comparison of DSC scans of spray dried lactose samples stored under different humidities for 3 d with scan of freshly spray dried lactose was shown in Fig.5.After stored at 32%RH for 3 d(Fig.5B),the crystallinity transformation of the spraydried lactose was not apparent,while after stored at 43% RH for 3 d(Fig.5C),the slight depression at 140°C and the endothermic peaks at 220°C indicated the emergence of α-lactose monohydrate.When stored at higher humidity of 53%RH and above for 3 d(Fig.5D-G),the system became mixture of α-lactose monohydrate and β-lactose as the exothermic peaks for β-lactose appeared at around 240°C.

Fig.6-DSC patterns of spray dried lactose.(A:freshly spray dried sample;B-G:samples stored for 7 d under RH 32%,43%,53%,65%,75%,85%).

Fig.7-XRD patterns of spray dried lactose:(A:freshly spray dried sample;B-G:samples stored for 3 d under RH 32%,43%,53%,65%,75%,85%).

The DSC patterns of spray dried lactose stored under different humidities for 7 d were shown in Fig.6.No apparent changes were found as comparing patterns in Figs.5 and 6 which indicated after the crystallinity transformation in the f i rst 3 d,there was no further change during the extended storage.

X-ray diffraction was employed to further verify the results of DSC.It can be seen from Fig.7 that the spray dried lactose remained amorphous after stored at 32%RH for 3 d,while under higher humidity,the appearance of crystal peaks indicated the exist of crystallinity.Meanwhile,the XRD patterns after 7 d of storage as shown in Fig.8 are similar to those in Fig.7 which illustrated the crystallinity transformation for spray dried lactose in correspondence with the results of DSC study.

3.5. Weight changes under humidity

Fig.8-XRD patterns of spray dried lactose:(A:freshly spray dried sample;B-G:samples stored for 7 d under RH 32%,43%,53%,65%,75%,85%).

Fig.9-Weight change percentage of spray dried lactose stored under different humidities(n=3).

The percentage of weight change of spray dried lactose stored under different humidities was shown in Fig.9.It can be seen that when the humidity was increased from 32%to 43%,the samples absorbed water resulting in an increase of weight. While at humidity of 53%,the samples began to lose weight. This is because the amorphous lactose began to absorb water and recrystallized to form α-lactose monohydrate leading to the increase in weight when the atmosphere humidity was below43%.Underahigherhumidity,theamorphouslactosein the system completely transferred to the mixture of α-lactose monohydrateand β-lactose.Uponcrystallization,theabsorbed water was expelled from the material causing weight loss[22]. However,the specif i c mechanism should be further studied.

3.6. Investigation on storage condition

Spray dried lactose was stored in a silica gel dehydrator for 3 months,then the XRD pattern was obtained as shown in Fig.10.The single diffuse peak characteristic with no longrange order indicated the amorphous nature of the spray dried samples[23].Whether the crystallinity transformation would occur after 3 months requires longer term research. Consequently,it can be deduced that dry powder inhalation using spray dried lactose as carrier may not transfer to crystal at least for 3 months of storage when silica gel is used to control humidity under 30%RH at room temperature.

4. Conclusion

Fig.10-XRD pattern of spray dried lactose stored in desiccator with silica gel for 3 months.

Spray dried lactose was prepared as the carrier for dry powder inhalation and wascharacterizedforparticle sizeand morphology,moisture content,crystallinity and crystallinity transformation under humidity.The results suggested thatthe obtained spray dried lactose should be stored under humidity controlled strictly,best below 30%RH,in order to maintain its amorphous nature.To better understand the relationship between crystal form and the behaviour of the carriers,further studies are underway for the development of dry powder inhalation.

Acknowledgments

This work was supported by the National Natural Science Foundation of China(No.:81173002)and the National Science and Technology Support Program(No.:2012BAI35B02).

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*Corresponding authors.

E-mail addresses:pxin_1385@163.com,mercurypan@foxmail.com(X.Pan),yaoqhe@mail.sysu.edu.cn(Q.Yao).

1The f i rst two authors contributed equally to this work.

Peer review under responsibility of Shenyang Pharmaceutical University.

http://dx.doi.org/10.1016/j.ajps.2014.07.006

1818-0876/? 2014 Shenyang Pharmaceutical University.Production and hosting by Elsevier B.V.All rights reserved.

Lactose

Carrier

Dry powder inhalation