Introduction

Nanoparticles gained therapeutic importance flourishing respiratory organ trental delivery due to its ability to enter into deeper parts of the lung, to elude powerful pulmonic dendrites along with mucociliary consent methods, sequent in prolonged residence time (Daniher & Zhu, 2008; Daraghmeh, Al-Omari, Sara, Badwan, & Jaber, 2002). Statistical data on asthma showed that children missed 12.8 million school days, with 444,000 patients hospitalized, 1.7 million emergency visits, physician visits of about 10.6 million and 3,613 deaths (NIH Publi­cation 10-7542., 2010). Due to the unique features like surface to mass ratio, the ability to absorb creates an exterior region to carry various compounds, and nanoparticles gained much importance in acupuncture in reference to an asthma attack. Dry powder inhaler (DPIs) finds up to be effective to deliver a drug into the lung efficiently (Dolovich & Dhand, 2011; Edwards, Ben-Jebria, & Langer, 1998). DPIs can be easily carried with less drug loss as well as provides spectacular drug effectively to the lung (Hickey & Garcia-Contreras, 2001; Roy & Vij, 2010). The particles with a mass median aerodynamic diameter (MMAD) under 5µ hit in order to be the best candidates for a pulmonic orphan drug. Particles in the above range will deposit in stroke and therefore increases systemic drug absorption (Li & Mansour, 2011; Zhang, Wu, Chan, & Watanabe, 2011).

Aspergillus fumigatus is a fungus which is likely one of the major cause for asthma. Hence in this comprehensive study, Terbutaline Sulphate (a bronchodilator) and Itraconazole (an antifungal) was used for spectacular acupressure going from asthama attack. This study was planned to design, optimize and develop a novel dry powder aerosol using physical mixing and spray drying.

Materials and Methods

Terbutaline Sulphate and Itraconazole were obtained from KP Labs Hyderabad. Lactose was obtained from Drugs India Hyderabad. Trehalose from Hyashibara Co Ltd Japan. Methanol was obtained from Himalaya Scientific Nellore. Other chemicals used in the entire work owe allegiance analytic shortlist.

Pre-formulation

Preformulation parameters like organoleptic properties, Solubility, the Melting point was determined. Compatibility between drug and excipients and thermal properties of the drug was determined using FT-IR Spectroscopy and Differential Scanning Calorimetry. All the Drugs and Excipients have been verified as various physical properties like Bulk Density, Tapped Density, Angle of Repose, Hausner’s Ratio, Compressibility Index and the results revealed the poor flow properties of drug and excipients. So, in order to improve the flow properties, drugs and excipients were milled to nano size and further evaluations were done (Ali & Lamprecht, 2014; Meenach et al., 2012).

Formulation

Physical Mixing

Terbutaline Sulphate, Itraconazole and the sugar used as carriers (Lactose, Trehalose) were taken in specified quantities shown in Table 2 and dried at 37⁰ c using vacuum oven for 12 Hrs. The size of drug and carriers were reduced using grinding mill (Jet mill, air is supplied at 110 psig with air classifier speed of 4200 rpm) for 3 Hrs for nanosized particles and for 2 hours for fine particles (Jet mill, air is supplied at 90 psig with air classifier speed of 3800 rpm). Calculated amount of Terbutaline Sulphate and Itraconazole was mixed separately in every formulation with fine lactose and micronized lactose, fine trehalose and micronized trehalose in geometric dilutions and passed through 60# screen, blended and filled into size "3" hard gelatin capsules in addition to booklet silique weft roadster (SS handbook space capsule woof machine by Pharmafill Technologies) with a weight of 25.5 mg per capsule of Terbutaline Sulphate, 100 mg per capsule of Itraconazole, 125.5 mg per containing Terbutaline Sulphate and Itraconazole in combination (NIH Publi­cation, 2010; Newman, Anderson, Byron, & Dalby, 2009).

Spray Drying

Mini spray dryer (SS Laboratory Spray Dryer (LSD 01)) using a high-performance cyclone separator

was used for performing a spray drying process using dilute solutions. All the ingredients were taken in specified quantities shown in Table 3.

Solution-1

Dilute way out used to be mapped out by means of dissolving each ingredient in calculated quantities, consisting of Terbutaline Sulphate with lactose and Terbutaline Sulphate with trehalose using methanol to make total powder concentrations of 0.4%W/v.

Solution-2

Dilute way out turned into mapped out with the aid of dissolving each ingredient in calculated quantities, consisting of Itraconazole with lactose and Itraconazole with Trehalose using methanol to make total powder concentrations of 0.4%W/v (Olsson, Borgström, Lundbäck, & Svensson, 2013).

Solution-3

Dilute solution way out used to be mapped out by means of dissolving each ingredient in calculated quantities, consisting of Terbutaline Sulphate and Itraconazole with lactose, Terbutaline Sulphate and Itraconazole with Trehalose using methanol to make total powder concentrations of 0.8%W/v (Pritchard, 2001).

Spray Drying- Conditions

• Atomization Rate-600 L/Hr

• Pump Rate-15 ml/min (Medium Pump Rate)

• Inlet Temperature-150ºc

• Nozzle Diameter-0.7mm

Sprayer dehydrated vectors have been separated by cyclone separator and sealed in the glass vials and stored in dessicator under ambient pressure. Outlet temperatures were given in Table 4.

Evaluation

Average fill weight per capsule

20 capsules were randomly taken and the overall table of contents was taken way externally, shedding a bit much a part of the over shell as completely as possible (Rogueda & Traini, 2007). 20 capsules were weighed for content and determined using the following formulae.

Average fill weight (mg) = Content present in 20 capsules (mg)/20 ____(1)

Moisture content

In a 5 ml volumetric flask, 6-10 mg of sample was dissolved in methanol and hydranal KF reagent was filled in the reaction cell and test solutions were injected into it. The moisture content present in the sample was calculated by conducting all the experiments in triplicate (n = 3) (Meenach et al., 2013). Moisture content of the sample was calculated using the formula

SF*100/W ______(2)

Where W = Coefficient the Sample, in mg, S = Volume of the KF reagent, in ml, F = the water compare factor of KF reagent, in mg.

Percentage Yield

Percentage yield was determined by taking the weights of reactants (drug and excipients) and products and calculated in line with the formula.

% Y = (Weight of Product / Weight of initial drug and excipients)*100 _____(3)

Drug Content

10 capsules were taken and transferred into a 100 milliliter volumetrically flask (Terbutaline dose equivalent to 50mg and Itraconazole dose equivalent to 500mg) and capsule was dissolved under sonication by adding the suitable volume of diluent (Water for Terbutaline sulphate and Methanol for Itraconazole), for about 10 minutes as well as intermittent quaking and final volume was made with diluents. Solution turned into selected through a 0.45μ membrane and estimated Terbutaline Sulphate at 276nm and Itraconazole at 264nm using UV spectrophotometer (2060 Plus UV-VIS Dual Beam, Analytical Technologies Pvt. Ltd.).

Scanning electron microscopy (SEM)

Hitachi S-4300 microscope was used to analyze the shape and morphology. Double-sided mucilaginous atomic number 6 directories have always been compensate alum stubs and samples were gold coated using Emscope SC400 coating system. Working distance with 13-15mm was set and a beam of electrons with fast electricity of 5-10 kV was used. Similar conditions were maintained for all the magnifications (Shekunov, Chattopadhyay, Tong, & Chow, 2007).

Size distribution using laser light diffraction

Shimadzu laser light diffractor was used to analyze the size plus filler redistribution of the particles in all the formulations. The samples were ultrasonicated for 10 min by dissolving the samples in chloroform. The dispersion was transferred into a measuring radical cell plus stored stirring. Volume-based measurements like Dv1mw980, Dv50, and Dv90 were used for the characterization of particles (Mansour, Wu, Hayes, Zwischenberger, & Kuhn, 2013). The span value was calculated using the formula below (Mansour et al., 2013; Zhang et al., 2011).

((Dv90-Dv10)/Dv50) _______(4)

Particle Size and Zeta potential

Malvern zeta sizer was used to measure the electrophoretic mobility of the particles present in all the formulations by diluting the sample with water.

Determination of MMAD using cascade impactor

Seven ultimateness fall impactor was used to determine the MMAD. Randomly selected capsules (3 per each formulation) were needle pierced and was drawn into the cascade impactor with a controlled flow rate of 28 L/min with delay time of 10s. A total of 25.5mg (TER-A to TER-D) total per run, 100mg (ITR-A to ITR-D) total per run and 125.5mg (TER: ITR-A to TER: ITR-D) total per run. After deposition of DPI, Terbutaline Sulphate and Itraconazole content was determined in each chamber by UV Spectroscopy. Absorbance was detected at 276nm for Terbutaline Sulphate and 264nm for Itraconazole. MMAD and GSD at each stage for all the samples were calculated using MMAD calculated.

In vitro drug release studies

The dissolution studies are ordered palmy phosphate buffer with pH 7.4 at 37 °C using a rotating basket apparatus at 50rpm. The studies were performed with milled and spray-dried particles containing Terbutaline Sulphate and Itraconazole. At a predetermined time interval, 5ml aliquots had been withdrawn and a fresh 5 ml buffer was added simultaneously. Cumulative release of Terbutaline Sulphate and Itraconazole by UV/Vis-spectroscopy (λmax: 276 nm for Terbutaline Sulphate, λmax: 264 nm for Itraconazole) (Li et al., 2014).

Results and Discussion

Preformulation

FTIR and DSC studies were performed for individual drugs and excipients. From the results, no incompatibility was observed between the drugs and excipients. The results were shown in Figure 8; Figure 7; Figure 6; Figure 5; Figure 4; Figure 3; Figure 2; Figure 1. All the drugs, excipients and formulations were subjected to various physical properties like Bulk Density, Tapped Density, Angle of Repose, Hausner’s Ratio and Carr’s Index to evaluate the density and flow of powder. Average fill weight also determined. All the formulations were found to have good flow properties when compared to plain drug and excipients. Due to the good flow property, all the formulations can be effectively delivered through the dry powder inhaler, which is very important in formulating as DPIs. Results were shown in Table 1 and Table 5.

Moisture Content

Moisture content for raw drugs and formulations was done by Karl Fisher serial dilutions. The residual water content for raw Terbutaline Sulphate and raw Itraconazole was found that one may be 4.41±0.18 and 7.62±0.11, respectively. Moisture content of all the formulations prepared by physical mixing and spray drying was decreased when compared to the raw drugs, which may be due to the general ubiety of lactose and trehalose in all the formulations.

Moisture content of formulations prepared by physical mixing ranges from 3.80±0.44 to 4.89±0.22. Formulations (Physical mixing) containing the combination drugs showed a slight increase, which may be due to the powerful ubiquity of Itraconazole (Fluffy Powder). Formulations prepared by way of atomizer brushing showed less moisture content when compared to the formulations prepared by physical mixing and ranges from 3.12±0.18 to 4.21±0.27. Results record in Figure 9 & Figure 10.

Percentage Yield and Drug Content

The spray-dried formulation showed less percentage yield (ranges from 77.50–88.75) when compared to milled formulations (ranges from 95.69–98.90). Less percentage yield for spray-dried formulations, when compared to milling, maybe due to the conditions used in the spray drying process. Drug content comes across in order to be within limits (95.8±0.15 - 102.3±0.25) and results reflect in Table 6.

Scanning Electron Microscopy

The particle size of raw material was found to have a maximum particle size that may interrupt the respiratory passages. Hence the particle size has been decreased to the respirable-size using milling and spray drying. Particles were found that one may be smooth and spherical for spray-dried formulations. Formulations prepared through milling were found to have rough and irregular in shape. Results reflect in Figure 13; Figure 12; Figure 11.

Particle sizing and Size distribution

All formulations tried and true nonparametric littleness distributions. The formulations prepared with trehalose as the carrier showed less D_v90, D_v50 and D_v10 values due to the fineness in the particles of trehalose when compared to lactose. The D _v50 values were in the variety of 0.43-0.89 µm for all formulations. The D_v10 values were in the range of mountains of 0.21–0.49 µm. This shows that all the particles were found planned in very particulate matter range and results reflect in Table 7 and Figure 14.

Zeta Potential

From the results, it was found that the formulations prepared by using milled trehalose as carrier attained less particle size when compared to milled lactose. Further, individual formulations attained less particle sizes when compared to the formulations in combination. Polydispersity index was found impending in a very range of mountains of 0.101 to 0.467, which explains the particle size distribution as monodisperse. Zeta potential was found so be between +17±0.13 to +32±0.12, which explains the particles as moderately stable. Results had been shown in Table 8 and Figure 15 &Figure 16.

Determination of MMAD (Mean Median Aerodynamic diameter)

Particle deposition on lower degrees containing ultimate twain totally that one may phase 7 (the lowest stage) is said. General, spectacular percent deposit towards phase a meg increased for unmilled formulations and formulations in which the drugs are given in combination. The exception to this, spray-dried formulations showed very slight differences in % deposition at stage 1. This trend was opposite for stage 4, where the amount of powder deposited on this stage increased in spray-dried systems when compared to physically mixed systems. From the results, it was found that significant deposition was seen in lower stages (Stages 4–7), which explains that the particle may deposit in the deep lung region, thereby increasing the therapeutic benefit. Results were shown in Table 9 and Figure 17.

In-vitro drug release studies

Dissolution studies indicate that formulations prepared using milled trehalose release the drug at a faster rate due to its very fine particle size. TER-A(sd) and ITR-A(sd) was found that one may be the best formulations which released the drug upto 98.9% and 99.1% in 180 mts. All the formulations prepared viva atomizer drying has high % depositions from stage 2 to 7 when compared to other formulations, which made the spray-dried formulations the most effective drug delivery throughout the lung. Results were shown in Figure 18 & Figure 19.

Conclusions

Demanding tasks in formulating dry powder nanoparticles is achieving the respirable particle size that avoids the physical limitations of the general stroke and delivering the synergist to a powerful target site. From the results obtained, it was evident that the prepared formulations can be prepared as DPIs that releases the drug directly into the lung efficiently for the acupressure going from asthma. Spray drying was found to be the best method suitable for the readying containing dry powder nanoparticles when compared to physical mixing of milled drug and excipients. Combination foams could provide coeval delivery to the synoptical computer epithetical legal action maximizing the capability outcome in reference to the medicinal drugs.