1.

Introduction

Currently hypertension is one of the most significant risk factors for cardiovascular

disease, affecting especially the adult population. One of the main classes of antihypertensive

drugs for the oral route are the beta blockers (Larochelle, 2014). In this specific therapeutic class,

bisoprolol fumarate (BSP), chemically known as 1-[4-[[2-(1-methylethoxy) ethoxy]-methyl]-3-

[(1-methylethyl) amino]-2-propanol salt (The British Pharmacopoeia, 2012), is a cardioselective

β1-adrenergic blocking agent used for secondary prevention of myocardial infarction, heart

failure, angina pectoris and mild to moderate hypertension (Lin et al., 2012; Metra et al., 2007;

Poldermans, 2002). The daily dose of bisoprolol is 5 to 10 mg, which can be increased, if

necessary, to up to 20 mg/day. After oral administration bisoprolol is rapidly absorbed in

gastrointestinal tract, the absolute bioavailability after a 10 mg oral dose of BSP being about

80% (Saini-Chohan et al., 2009; Ahad et al., 2015). The maintenance of a constant plasma level

of a cardiovascular drug is important for the desired therapeutic response. Because the half-life

of the bisoprolol is about 3-4 h, multiple doses need to be administered in order to maintain a

constant plasma concentration. The immediate release tablets with bisoprolol (2.5, 5 or 10

mg/tablet) have been found to have many associated drawbacks such as diarrhea, weakness,

fatigue, anxiety, headache, dry mouth, nausea, vomiting, stomach pain, etc. Therefore, to reduce

frequency of administration and improve patient compliance, maintain a uniform drug

therapeutic level and reduce side-effects, a sustained release dosage formulation of bisoprolol is

desirable. Drug release from the dosage form is controlled by the type and proportion of

modified-release polymer used in the preparations. The preparation of matrix tablets with rate

controlling polymers is the simplest and most widely used method for achieving desired

extended release rate for drugs.

 Several studies have investigated different matrix tablets (hydroxypropyl methylcellulose

(HPMC) in association with sodium bicarbonate, Carbopol 974, glycerin, propylene glycol,

polyethylene glycol and ethyl cellulose, with the purpose of controlling the release of bisoprolol

(Mhetre et al., 2014; Malakar et al., 2012; Ankit et al., 2015; Kavitha et al., 2015; Abo Enin et

al., 2014; Sarath Chandran et al., 2013).

Precirol ATO5 (PREC), a mixture of mono-, di-, and triglyceryl esters of palmitic and

stearic acids, with a HLB value of 2, is a multipurpose pharmaceutical ingredient recommended

to be used in topical and oral pharmaceutical formulations as a taste masking agent, lubricant and

modified-release excipient (https://www.gattefosse.com/precirol-ato-5). Several research studies

have been conducted to investigate the proportions of Precirol ATO5 for lipophilic matrix tablets

and capsule formulations (Çabasi et al., 2001; Popa et al. 2013). For modified release purposes

the recommended proportions vary between 10 and 50%, more likely 10-25%. Innovative

formulations have been investigated also for orally disintegrating tablets containing PREC as

binder for taste masking purpose in proportions of 4.5-5%, which do not prolong the

disintegration times (Kibbe, 2000; Rowe et al., 2009). Given its highly fatty nature, PREC can be

combined in modified release tablets with hydrophilic polymers such as hydroxypropyl

methylcellulose (HPMC).

HPMC is a semisynthetic, inert, viscoelastic polymer widely used in solid oral dosage

forms as a binder, a film coating, and a controlled-release matrix (Brady et al., 2017). HPMC of

various viscosity grades has been used as a polymer for hydrophilic matrix tablets, due to its

swelling capacity in water. For achieving an optimal kinetics of tablets it is necessary a fast

hydration of the outer layer of the HPMC matrix to form a gel barrier, which provides

subsequent slow release of the drug (Li et al., 2005). In this context the drug release takes place
mainly by diffusion through, and/or subsequent erosion of the gel layer; while the drug from the

outer surface of the matrix is released firsty by a burst effect. Nevertheless, it has been

demonstrated that modified drug release from HPMC matrix tablets depends also on the drug

solubility (Siepmann and Peppas 2012; Hardy et al., 2007). The water-soluble drugs are released

mainly through diffusion mechanism, whereas insoluble and low-soluble drugs are released

through erosion (Lin et al., 2016).

Bisoprolol fumarate is a BCS class 1 drug, with high solubility and low permeability. For

highly water soluble drugs (Charoo et al., 2014), literature data suggest an inflexibility of release

kinetics, which may lead to sub-optimal in vivo release profiles (Lin et al., 2016). This

characteristic has recommended modulation of release kinetics of HPMC matrix tablets by

combining it with other excipients (PVP, carbopol, glyceryl esters, fatty acids) or by designing

multi-layered tablets. The processing of the tablets in solid-state can lead to reactions between

the drugs and formulations, possibly initiating alterations in their stability, solubility, dissolution

rates and bioavailability. Therefore, structural evaluation of the matrix and compatibility studies

are needed to ensure that potential physical and chemical interactions between a drug and

excipient do not compromise the stability of the drug (Qiu et al., 2005; Guerrieri and Taylor,

2009).

The purpose of the present study was to obtain modified release matrix tablets with

bisoprolol fumarate using an innovative combination of Precirol ATO5 (PREC) and

hydroxypropyl methylcellulose, in order to investigate the optimal ratio between these

ingredients as matrix formers and to analyze the pharmaceutical properties and the kinetics

release profile of bisoprolol modified release tablets, with a view to maximize duration and

bioavailability of this drug and to improve efficacy and better patient compliance.
2. Materials and methods

2.1. Materials
The substances used for the pharmaceutical formulations were: Bisoprolol fumarate 20

mg/tablet weight (BSP, provided by Unichem Laboratories LTD, India), Precirol ATO5

(glyceryl distearate - PREC, kindly offered by Gatefossé) and hydroxypropyl methylcellulose

(HPMC K4M, 4000 cps, Premium, Colorcon).

2.2. Preparation of the pharmaceutical formulations

The preparation of tablets was done by melt granulation process (Royce, 1997), using the

property of PREC excipient to melt at low temperatures and consequently to bind the mixture

particles. The powder (containing BSP, PREC and HPMC) was mixed and heated to 65°C in an

oven. The resulted agglomerated mass was passed through a 600 μm aperture pharmaceutical

sieve (Fisherbrand, 8̎’-FH-SS-SS-US-30). The obtained granules were further compressed on a

Korsch EK0 tablet press (9 mm flat punches, 5 kN tabletting pressure). Since PREC is

recommended as a good tablet lubricant, the addition of an extra lubricant in the granulation

mixture was not considered (https://www.gattefosse.com/precirol-ato-5). Eight formulations of

tablets containing BSP were prepared with various excipients ratios according to Table 1.

Table 1. The formulations of tablets containing BSP and their tested pharmaco-technical

properties.

Formula F1 F2 F3 F4 F5 F6 F7 F8
Composition
PREC (%) 10 15 20 25 30 35 40 45
HPMC (%) 76,66 71,66 66,66 61,66 56,66 51,66 46,66 41,66
Total tablet
150 150 150 150 150 150 150 150
weight (mg)
 In order to be pharmacodinamically effective, the dose strength of 20 mg bisoprolol was

selected for modified release as a sum of 2-8 therapeutic doses.

2.3. Methods of pharmaceutical formulation characterization

2.3.1. Pharmaco-technical properties
The pharmaco-technical properties of the tablets were tested: hardness, thickness and

diameter, by using a Pharma Test Dr. Schleuniger hardness tester (N), and the friability with an

Electrolab EFII friabilator (100 rpm, 4 minutes), according to the method recommended by the

European Pharmacopoeia (2018) and EMEA (2004).

2.3.2. Fourier Transform Infrared Spectra (FTIR)

The measurement of IR absorption spectra was made with an ALPHA spectrophotometer

(Bruker, Germany) for the pure drug and the excipients separately, and then combined.

Processing of the spectra was performed using the Grams 9.1 program (Thermo Fisher

Scientific).

2.3.3. Wide Angle X Ray Diffraction – WAXD

WAXD analysis was performed on a Diffractometer D8 ADVANCE (Bruker AXS,

Germany), using the CuKα radiation (λ=0.1541 nm). The working conditions were 40 kV and 30

mA, 2s/step, and 0.02 degree/step. All diffractograms were recorded in the range 10-90 2θ

degrees, at room temperature.

2.3.4. Thermal analyses

2.3.4.1. Thermogravimetric analyses
Thermogravimetric analysis (TGA) was carried out under constant nitrogen flow (200

ml/min) at a heating rate of 10 oC/min using a Mettler Toledo TGA/SDTA 851 balance. The

heating scans were performed on 3-5 mg of sample in the temperature range 25–600oC.
2.3.4.2. Differential scanning calorimetry
The DSC curves were recorded using a Mettler Toledo DSC1 device with a 10°C /min

heating rate, in a nitrogen flow of 150 ml/min. The sampless weighing between 2.2 and 4.8 mg

were taken in pin-holed aluminum pans and underwent two heating cycles and a cooling cycle

within the 25-150, 200 or 240oC temperature range. The temperature accuracy was ±0.02 °C, and

the heat flow resolution was 0.04 μW. The device was calibrated using indium and the In Check

method stored in the STARe software, according to the instruction manual.

2.3.5. In vitro drug release tests

BSP release from the tablets was investigated through dissolution test on a SR8 Plus

Dissolution test station equipped with a Disoette II carousel for automatic sampling and a V530

spectrophotometer (Hanson Research, U.S.A.). The test was performed over a period of 9 hours,

according to the following protocol: Apparatus 2 (paddle); rotation: 50 rpm. The volume of the

dissolution media for each vessel was 900 ml, as follows: for the first 2 hours 0.1 M HCl solution

was used and for the remaining 7 hours the medium was replaced with pH 6.8 phosphate buffer

solution, in order to mimic the physiological conditions. Samples were collected automatically

every hour and the BSP concentration was determined by spectrophotometric method

(wavelength = 222 nm). The same volume of fresh dissolution medium at the same temperature

was added to replace the amount withdrawn after each sampling. The drug amount of cumulative

release from tablets was calculated with a standard curve prepared using bracketed concentration

of BSP each dissolution medium in a range 10 to 110% of a theoretical concentration of 5

µg/mL.

Different kinetic models were applied (Dash et al., 2010; Bruschi, 2015) to the release

data of test formulations in order to investigate the release mechanism of drug from the sustained

release matrix tablets, using a Matlab 7.9 software.

 The zero-order release model: 𝑭𝒕 = 𝑲𝟎 ∗ 𝒕 , (where 𝑭𝒕 is the released percentage at

time t).The predicted release (forecast) at 𝒕 = 𝟏𝟎 is 𝑭𝟏𝟎 (%) = 𝟏𝟎 ∗ 𝑲𝟎 (%).

The predicted time until complete release is determined by the following formula: 𝑻 = 𝟏𝟎𝟎/𝑲𝟎.

The first-order release model: 𝑭𝒕 = 𝟏𝟎𝟎 ∗ (𝟏 – 𝒆− 𝑲𝟏 ∗ 𝒕 ) (where 𝑭𝒕 is the released

percentage at time t). The predicted release (forecast) at 𝒕 = 𝟏𝟎 is:

𝑭𝟏𝟎 (%) = 𝟏𝟎𝟎 ∗ (𝟏 – 𝒆−𝟏𝟎∗ 𝑲𝟏 )(%)

The predicted time until complete release is determined by the following formula:

𝐓 ≌ 4.6052/𝐊 𝟏

The Korsmeyer-Peppas release model is defined by the equation:

𝑭𝒕 = 𝑲𝒑 ∗ 𝒕𝒏 , where 𝑭𝒕 is the released percentage at time t.

The predicted release (forecast) at 𝒕 = 𝟏𝟎 is:

𝑭𝟏𝟎 (%) = 𝑲𝒑 ⤫ 𝟏𝟎𝒏 (%)

The predicted time until complete release is determined by the folowing formula:

𝑻 = (𝟏𝟎𝟎/𝑲𝒑 )𝟏/𝒏

The Higuchi release model is defined by the equation:

𝑭𝒕 = 𝑲𝑯 ∗ √𝒕 , where 𝑭𝒕 is the released percentage at time t.

The predicted release (forecast) at 𝒕 = 𝟏𝟎 is:

𝑭𝟏𝟎 (%) = 𝑲𝑯 ⤫ √𝟏𝟎 (%)

The predicted time until complete release is determined by the folowing formula:

𝑻 = (𝟏𝟎𝟎/𝑲𝑯 )𝟐
3. Results and discussions
3.1. Pharmaco-technical properties
Uniform, intact, smooth-surfaced tablets could be obtained: their characteristics are

displayed in Table 2. The hardness of tablets did not record high values, nevertheless increasing

tablet hardness could be correlated with the higher proportion of PREC in the formulation, with a

maximum value for F8 (52.3 N). A very low friability was an important characteristics displayed

by all tablets, 4 formulas recording a zero value; this is attributed to the plasticity of the melt

granulation binder PREC. The values of tablet thickness and diameter indicated a good die fill

during the compression process of granules.

Table 2. Pharmaco-technical properties of tablets

Friability (%)
0.00 0.22 0.23 0.21 0.00 0.22 0.00 0.00
n=20
Hardness (N)
34.3 36.66 40.2 43.9 45.8 46.7 49.7 52.3
n=10
Thickness (mm)
2.16 2.12 2.06 2.06 2.00 2.08 2.07 2.10
n=10
Diameter (mm)
9.02 9.01 9.01 8.99 8.99 8.99 9.01 8.99
n=10

3.2. Compatibility study

For a better observation of the changes appearing in the structure of all materials, we

represented in figures only the pure components and two other samples, as an example.

3.2.1. FT-IR analysis

FTIR spectroscopy is a sensitive technique for evaluating the intramolecular interaction

and conformational arrangement which influence the spatial distribution in the structure of

materials. Figure 1 shows the IR spectra in transmittance mode of the pure components and

studied formulation in the 3700-2400 cm-1 and 1800-800 cm-1 spectral region. In order to
evaluate the intermolecular interactions between the formulation components the

calculated/theoretic spectra were performed by using the additivity low (Popescu et al., 2006).

The first spectral region contains the specific bands corresponding to stretching vibration of free

and bonded OH and/or NH groups and to aliphatic and aromatic CH groups. As can be seen from

Figures 1a, 1b, there are no significant differences between the experimental and calculated

spectra of studied formulations in the spectral region between 3000-2700 cm-1 (assigned to the

stretching vibration of aliphatic and aromatic CH groups).

a b
 c d

Figure 1. FT-IR spectra of the studies samples in the 3700-2400 cm-1 (a, b)

and 1800-800 cm-1 (c, d) spectral regions

In the region assigned to the stretching vibration of free and OH and/or NH groups a

small variation in intensity of the bands is observed. It is well-known that hydrogen bonds play

an important role for the interaction of OH and NH groups with carbonyl or ether functionalities.

In order to evaluate the content of OH and NH groups the deconvolution of this region in

component bands was made (Figure 2).

 Figure 2. The deconvolution of experimental IR spectrum of the F6 formulation in the 3700-

3000 cm-1 spectral region

For both calculated and experimental spectra the best fit was obtained with 5 bands,

located at about 3580, 3505, 3435, 3324 and 3247 cm-1. The first 3 bands describe the free and

hydrogen-bonded OH groups, while the last 2 describe the free and H-bonded NH groups.

Depending on the strength of the interaction, the absorption bands of the involved functionalities

undergo more or less extensive band shifts and intensity changes. From Figures 3 (please see

supplementary material) can be observed that the positions of the bands assigned to free OH

groups are located at about 3610 and 3525 cm-1 in the case of F1 and F2 samples, and are shifted

to lower wavenumber at 3590 and 3500 cm-1 for the sample F3 and at about 3580 and 3505 cm-1

for the samples F4-F8. The position of these bands is shifted to higher wavenumber in calculated

spectra for the F1-F3 samples and to lower value for the F4-F8 samples. The position of the band

assigned to bonded OH groups is observed at about 3420 cm-1 and is slowly shifted to higher

wavenumber with increasing the PREC content in the samples. For the calculated spectra of the

F1-F3 samples, this band is situated at about 3500 cm-1 and is shifted to lower value in the case
of the other samples (F4-F8). Regarding the bands assigned to free and bonded NH groups, from

about 3340 and 3240 cm-1, they are shifted to higher value for the F2-F3 samples, but remain

almost constant for the other studied samples (F4-F8). The values for calculated spectra present a

similar trend with the experimental ones, only slowly shifted (with about 7-10 cm-1) to higher

values.

Using the values for band position obtained through the deconvolution process was

possible to evaluate the energy of the hydrogen bonds using the following formulae.

1   0  
EH   
k  0 

where ν0 is the standard frequency of the OH (3650 cm-1) and NH (3474 cm-1) monomers

observed in the gas phase, while ν is the stretching frequency of OH and NH groups observed in

the infrared spectrum of the sample, and 1/k is a constant equal to 2.625*102 kJ.

The values of the energy are of about 5, 10.5, 15.5, 12.5 and 17 kJ for the experimental

spectra for the samples F4-F8, with lower values in the case of the F1-F3 samples. They are

slightly shifted to higher values in case of calculated spectra (Figure 3).

Figure 3. Graphical representation of deconvoluted bands position (a), H bond energy of free and
bonded OH and NH groups (b), and integral areas, % (c)
 Comparing the values corresponding to the integral area of the deconvoluted bands of the

calculated and experimental spectra, it can be observed that the integral area of the bands

corresponding to free OH and NH groups decrease from about 16-14 % and 2.5-2 % in the case

of calculated F4-F8 spectra to about 15-8.5 % and 1.2-0.4 % in the case of the experimental

spectra, respectively. The integral areas of H-bonded OH groups increase in experimental spectra

from 59-66 % to 62-75 % for the F4-F8 samples. This observation is similar to H-bonded NH

groups. This confirm the involvement of the free OH and NH groups in the formation of new H-

bonds between the components of the system. The samples F1-F3 present different trends; the

number of free and bonded OH groups is higher in experimental spectra comparing to calculated

ones, while the number of free and bonded NH groups is higher in the calculated spectra. The

reduced involvement of the free OH groups in the formation of H-bonds between the

components of the system is corelated, in this case, with the reduced hardness of these samples.

In the fingerprint spectral region, between 1800-800 cm-1, all components present a large number

of bands. Thus, the IR spectrum of PREC indicate the main spectral bands: at 1735 cm-1 assigned

to stretching vibration of C=O groups, at 1472 cm-1 assigned to scissoring vibration of CH2

groups, 1393 cm-1 assigned to rocking vibration CH2 groups, 1425 and 943 cm-1 assigned to

bending vibration of OH groups, 1291, 1103 and 1062 cm-1 assigned to stretching vibration of C-

O groups and at 719 cm-1 assigned to rocking vibration of methyl long-chain.

The IR spectrum of HPMC present specific bands at 1647 cm-1 assigned to stretching

vibration of cyclic C-O groups, 1459 cm-1 assigned to asymmetric stretching vibration of CH3

groups, 1377 cm-1 assigned to asymmetric bending of cyclic C-O-C groups, 1118 and 1063 cm-1

assigned to stretching vibration of C-O-C groups and 848 cm-1 assigned to rocking vibration of

CH2 groups. In case of BSP characteristic bands appear at 1613, 1515 and 1466 cm-1 assigned to
stretching vibration of aromatic C-C groups, 1092 and 1048 cm-1 assigned to bending vibration

of C-H groups, 1364, 1314 cm-1 assigned to symmetric bending of CH3 and CH2 groups, 1236

cm-1 assigned to stretching vibration of C-O groups and 1178 cm-1 assigned to stretching

vibration of -N-CH2 group. In the spectra of formulations all component bands were identified,

but the position of some of them is shifted to different wavenumber. Thus, the band of BSP from

1466 cm-1 is shifted to 1472 cm-1, from 1613 cm-1 to 1618 cm-1 and from 1236 cm-1 to 1239 cm-1.

Comparing the experimental and calculated IR spectra of the studied formulations, significant

differences appear for the bands situated at 1381 cm-1 in experimental spectrum which appears at

about 1372 cm-1 in the case of calculated spectra, 1240 cm-1 is shifted at 1235 cm-1 and the band

at 1102 cm-1 is shifted at 1094 cm-1. These bands are assigned to bending vibration of CH3

groups and to stretching vibration of C-O groups. These shifts indicate changes in

conformational arrangement of the molecular chains and/or specific interactions between them.

Principal component analysis (PCA). It is known that principal component analysis

(PCA) is used to visualize interrelationships among the independent variables and is useful in

identifying data outliers, therefore is useful to investigate the difference between spectra and

derive parameters characterizing the interaction between the different components (Xiong et al.,

2016).

The principal component factor 1 (PC1) describes 71%, principal component factor 2

(PC2) describe 19% and principal component factor 3 (PC3) describe 9 % of data the variance,

so 99% of the existed variances in all the studied spectra can be captured using these three

dimensions instead of the initial data. The three-dimensional coordinate system of these PC

scores is plotted in Fig. 4.

 Figure 4. Three dimensional PCA score plots of the studied samples

According to this figure, the PC scores for the calculated and experimental spectra

established some distinct patterns which clearly expressed their differences. It can be observed

that the calculated spectra appear in the same plane (PC1-PC2 plane) like the components, the

position depending on the components composition. This plane can be defined as the non-

interaction reference plane. In this case the variation on PC3 direction is about zero. The

concentration dependence related to these two factors is expressed by an arrangement of the

spectra in a triangle shape like, having in the corners the pure components. In case of

experimental samples a significant variation on PC3 score direction can be evidenced. Thus, the

plane PC2-PC3 can be defined as the interaction plane. The distances between the experimental

spectra of the formulations and the calculated spectra are evidenced in the case of all studied

samples, this indicating the presence of the interactions.

3.2.2. X-ray diffraction

X-ray diffraction is a structural characterization technique which gives information about

formation of intermediate compounds and the mean grain sizes of the two phases (Fig.5). Figure
5a shows the X-ray diffractograms of pure components and F3 and F7 formulations. Figure 5b –

present the X-ray difractograms of the other formulations.

a b
Figure 5. X-ray diffractograms of the studied samples

In the PREC diffractogram five signals were observed at 19.1, 19.5, 22.9, 23.6 and 24.0o

with a corresponding d spacing of 4.654, 4.564, 3.871, 3.766 and 3.703 Å. On the other hand, the

pure HPMC shows two broad bands at 2θ=9.26 and 19.48° (d spacing=9.545 and 4.554 Å)

corresponding to its polymeric semicrystalline nature. BSP is a low molecular crystalline

substance. The corresponding diffractogram contain a large number of distinct and sharp bands

(see Figure 5 and Table 3 present in suplementary materials).

 In the case of the formulations, apart of the bands arising from BSP which are very small

due especially to the low content of this component in the mixture and/or to the loss of

crystalline structure on the preparation process, appear the intense and clear bands from PREC.

Lately, the bands corresponding to HPMC appear like an amorphous background. As can be seen

from Table S2, the values for 2 in the formulations are slightly shifted for both PREC and BSP,

indicating deformation of the crystalline network of these compounds due to the physical

interaction between the system components.

3.2.3. TG/DTG analysis

In order to evaluate the thermal stability of the drug in the obtained formulation and to

establish the interaction between used excipients and BSP, the TGA curves were recorded

(Figure 6a). The first-order derivatives of TGA curves (DTG) (Figure 6b) reveal the

temperatures corresponding to the maximum decomposition rate. The curves show that the

studied samples exhibit one or two distinct and well-separated weight-loss processes (see the

DTG curves – Fig 6b).

 a b

Figure 6. TG (a) and DTG (b) curves of the studied samples

Excepting PREC, in all studied samples, the first process occurs in the temperature

interval between 32-35ºC and 101-111ºC, with a maximum between 57 and 67ºC and a

corresponding mass loss of 1-6 wt% (Table 4). This process is related to the removal of the

physical adsorbed water.

Table 4. TG/DTG results for the studied samples

F3 F7
T PREC HPMC BSP
exp calc exp calc
Ti1, ºC - 35 32 34 34 33 33
Tm1, ºC - 57 59 62 57 67 57
Tf1, ºC - 103 105 101 101 111 111
Δw1, wt% - 6.1 0.5 2.0 3.0 1.1 2.3
Ti2, ºC 201 241 113 136 136 141 141
Tm2, ºC 391 358 327 364 360 364 363
Tf2, ºC 469 431 485 467 467 478 478
Δw2, wt% 93.3 76.5 78.0 85.6 83.2 86.1 85.2
Δwresidue, wt% 3.4 13.15 15.0 8.3 10.0 7.9 9.1
 The second thermogravimetric process for the pure components takes place between 201-

469ºC and 241-431ºC, with a maximum situated at about 358 in case of PREC and at 391ºC for

HPMC. The mass loss in this stage is about 93.3 % for PREC and 76.5 % in case of HPMC. In

this stage the fission of molecules to a variety of low molecular weight products, including

carbon dioxide, carbon monoxide, water, hydrocarbons (acetaldehyde, glyoxal and acrolein) and

hydrogen takes place. For the BSP, the second decomposition process takes place in the

temperature interval between 113-485ºC and present three overlapped stages with a maximum at

327ºC and two shoulders at about 221ºC and 274ºC. In the case of the mixture of the excipients

with the drug, the decomposition process is starting at about 130 – 145 ºC and finishes at about

460- 482 ºC. The temperature corresponding to the maximum rate of the decomposition process

is about the same for all studied samples, of 364ºC.

Comparing the DTG curves for experimental and calculated samples, it can be observed

that the temperature corresponding to the maximum decomposition rate is shifted to higher

values in the case of the experimental data, indicating an increase in the thermal stability of the

compounds. As mentioned before, this behavior can be determined by the physical interactions

taking pace between the system components in the mixing process.

3.2.4. Differential scanning calorimetry (DSC)

For thermal behavior evaluation the studied samples were heated to 200°C, cooled to

30°C and subsequently re-heated to 200°C. The DSC scans of pure drug and individual

excipients and the drug excipient mixture are shown in figure 7.

 Figure 7. DSC curves for the studied samples

A sharp and asymmetric endothermic process is observed in the first heating scans of

PREC at 68.7ºC with the enthalpy of transition (ΔHf) of 189.64 J*g-1 and at 57.9ºC with a

shoulder at 52.6ºC and with the heat of transition (ΔHf) of 116.46 J*g-1 in the second heating

process. At cooling an exothermic process is observed at 52.9ºC with the enthalpy of transition

of 116.71 J*g-1. This can be due to the melting process of PREC. The DSC curves of HPMC

showed 2 endothermic processes in the first heating scan. The first one, is a broad process with a

maximum at 67.1ºC and a enthalpy value of 168.74 J g-1, due to the evaporation of absorbed

moisture and the second one, a small process with a maximum at 179.3ºC with a enthalpy value

of about 0.81 J g-1. These processes are not observed in the cooling and second heating scans.

BSP present two endothermic processes with maxima at 80.9 and 102.3ºC, respectively. These

processes are not observed in the cooling scan. Further, in the second heating scan an exothermic

process is evidenced at 35ºC and an endothermic process at about 100ºC. In the case of the drug

formulation, the values of the temperatures and shape of the curves are slightly changed. The
shapes of the curves are similar with those corresponding to BSP, but the temperatures are

shifted to lower values. Thus, for the first heating scan, the value corresponding to the maximum

temperature of the endothermic process is recorded between 66 and 63 oC. All samples present a

shoulder between 59 and 57 ºC, which is not observed in PREC. During the cooling scan, the

maxima are shifted to 49 - 51 ºC and the values corresponding to the shoulder to 40-44 ºC,

respectively. In the second heating scan, these temperatures appear at about 55-56 ºC for the

maxima and at about 44-45 ºC for the shoulders.

The processes assigned to BSP are not observed in the DSC curves of the drug

formulations. The variation in shape and temperature of PREC can be due to the purity of each

component. The disappearance of the processes assigned to BSP can be due the low amount of

drug in the formulation, to the solubilization of the drug inside the formulation matrix being in

an amorphous form, and/or to the interactions between drug and excipients.

3.3. In vitro drug release

The in vitro drug release from all formulations is presented in Figure 8. So, as seen in the

graphs, all F1-F8 formulation containing BSP showed sustained drug release over 9 hours.
 80

Cumulative release of BSP (%)

70
F1
60
F2
50
F3
40
F4
30 F5
20 F6

10 F7
F8
0
0 1 2 3 4 5 6 7 8 9
pH = 1.2 pH = 6.8 Time (hours)
Figure 8. The release of BSP from matrix tablets

The cumulative drug release from the prepared tablets after 9 hours of dissolution (Rh9,

%) was within the range 54.62-71.08% (Table 5).

Table 5. Release model parameters fitted on the release kinetics of the investigated tablets.
Korsmeyer-Peppas model
Formula/
F1 F2 F3 F4 F5 F6 F7 F8
Parameter
Kp 18.570 21.055 28.609 27.502 23.724 31.385 26.0590 31.267
n 0.520 0.500 0.440 0.420 0.450 0.350 0.430 0.350
R2 0.989 0.982 0.989 0.988 0.991 0.9825 0.995 0.979
F10 61.502 66.582 78.796 73.176 66.864 70.2629 70.138 69.998
T 25.467 22.557 17.187 20.851 24.459 27.4110 22.816 27.707
Higuchi model
Formula/
F1 F2 F3 F4 F5 F6 F7 F8
Parameter
KH 19.2389 21.0551 25.7581 24.1153 21.7351 24.1863 23.0589 24.0883
2
R 0.989 0.982 0.984 0.976 0.986 0.948 0.999 0.941
F10 60.838 66.582 81.454 76.259 68.732 76.483 72.918 76.173
T 27.017 22.557 15.072 17.195 21.167 17.094 18.807 17.234
First order release model
Formula/
F1 F2 F3 F4 F5 F6 F7 F8
Parameter
K1 0.103 0.118 0.162 0.144 0.123 0.142 0.135 0.140
R2 0.922 0.920 0.921 0.867 0.883 0.798 0.888 0.784
F10 64.611 69.395 80.386 76.303 70.861 75.882 74.108 75.572
T 44.342 38.902 28.276 31.990 37.353 32.386 34.088 32.680
Zero order release model
 Formula/
F1 F2 F3 F4 F5 F6 F7 F8
Parameter
K0 7.498 8.194 9.966 9.301 8.422 9.273 8.925 9.227
2
R 0.778 0.753 0.667 0.606 0.686 0.481 0.669 0.460
F10 74.983 81.946 99.664 93.015 84.222 92.736 89.257 92.272
T 13.336 12.203 10.033 10.750 11.873 10.783 11.203 10.837
Kp=Korsmeyer-Peppas constant; n=release exponent; R2= determination coefficient; F10 (%) =
prediction at t=10 h; T=predicted time (h) until complete release. KH= Higuchi constant; K1=
First order release constant; K0= Zero order release constant.

The formulations with the highest HPMC content displayed the slowest drug release (F1

and F2). Increasing the proportion of PREC lead to a more uniform drug release (F7 and F8). All

formulations displayed faster release in the first 4 hours, followed by a much slower release in

the next hours. Studies performed on tablets by Gohal et al. (Gohel et al., 2009) claimed that

HPMC can act as a channeling agent and can increase the release rate of drugs. This may be the

cause of higher release rate of BSP from the F3.

The correlation coefficient R2 values indicated that both the Higuchi and Peppas models

suitably describe the release of BSP from the prepared matrix tablets. The value of release

exponent n of Korsmeyer-Peppas model was found between 0.35-0.44 (for F8-F3), indicating the

Fickian diffusion mechanism of BSP release. Formulas F1 and F2 released BSP by a non-fickian

diffusion (n = 0.5, and 0.52). It is considered that, when n value ranges from 0.45 to 0.89, the

drug is released by a combination of diffusion and erosion mechanisms (Costa et al., 2001;

Panainte et al., 2018). F7 (PREC 40% and HPMC 46.66%) shows the best fitting on the

Korsmeyer-Peppas model (R2 = 0.9959; Kp= 26.0590, n =0.43).

For the Higuchi model the release data obtained according to the determination

coefficient (R2) had high linearity for F1, F2, F3, F5 and F7. That means for this formulations the

drug and excipients dissolve at rates proportional to their solubility and diffusion coefficients in
the dissolution medium. It is mainly the case of solid disperse systems (Craig, 2002; Lobenberg

et al., 2000)

4. Conclusions

This study presents the design and the characterization of extended release tablets of

bisoprolol by using the melt granulation technique, as a new pharmaceutical formulation with

higher compliance for the elderly hypertensive patients.

The differences found in physico-chemical properties of the matrices were not consistent,

but were significant enough to be able to modulate the amount of drug delivered. The FTIR

spectral studies confirm the involvement of the free OH and NH groups in the formation of new

H-bonds between the system components, which do not affect the release of the bioactive

compound. The DTG curves indicating a small increase in the thermal stability of the

compounds determined by the physical interactions taking pace between the system components

in the mixing process. This was also demonstrated by X-ray diffraction.

The present research shows that glyceryl palmitostearate (Precirol ATO5) at suitable

concentrations combined with HPMC polymer can be used effectively to modify the release rates

of bisoprolol in matrix tablets.

The results showed that a matrix tablet prepared with HPMC and PREC is a better system

for sustained release of a highly water-soluble drug like bisoprolol. Among the various

developed formulations of BSP, the formulations F7 appear suitable for further

pharmacodynamic and pharmacokinetic evaluation in a suitable animal model.

The study aims on finding new, improved formulations for modified-release tablets with

bisoprolol fumarate, a widely-used drug for the treatment of hypertension. The key findings of

this research relies on the use of the modern melt granulation technique for obtaining modified-
release tablets by using a combination binder of Precirol ATO5 and hydroxypropyl

methylcellulose in varied proportion, in order to optimize the formulation. The melt granulation

process involves agglomeration of solid particles using lipophilic excipients without the addition

of water, which involves fewer processing steps over wet granulation, at the same time providing

drug stability, uniform granule size and content uniformity. The technique can be easily

implemented for industrial manufacturing of modified-release tablets (Shanmugam, 2015; Patil

et al., 2015). By using a mathemathical software, some of the prepared formulations could be

fitted on the release models applied (Korsmeyer Peppas and Higuchi models); also, the predicted

time of the release was over 20 hours in most of the cases. These data both indicate that

modified-release was achieved in the proposed formulations. Formulating bisoprolol fumarate in

modified release tablets might be beneficial over the conventional tablet formulations, in order to

reduce dosing frequency, and improve patient compliance.

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Declaration of interest

The authors have no relevant affiliations or financial involvement with any organization

or entity with a financial interest in or financial conflict with the subject matter or materials

discussed in the manuscript. This includes employment, consultancies, honoraria, stock

ownership or options, expert testimony, grants or patents received or pending, or royalties.