Influence of multiple compression phases during tableting of spray dried Saccharomyces cerevisiae on microbial survival and physical–mechanical tablet properties

Abstract

The viability of probiotic microorganisms is essential for their health-promoting effects and must be preserved in the best possible way during the production of the final dosage form, such as tablets. This applies to both drying and tableting. Saccharomyces cerevisiae is spray-dried with suitable protective additives, which were identified in a previous study in which also the influence of the formulation during tableting was investigated. One aspect that has not yet been addressed is the effect of multiple compression, as it is typical with pre- and main compression when using rotary tablet presses. To investigate this, tablets are compressed up to five times. It is shown that when tablet strength and survival are considered together, the application of a pre- and main pressure does not have a significant effect. This facilitates the transferability of findings of compaction studies with a single compression phase. In addition, the data allow to consolidate the mechanism of inactivation of microorganisms during tableting found in previous studies by the same authors. This is based on the porosity reduction, whereby it is shown in the present study that it is irrelevant how this reduction is achieved (change in compression stress or the number of compression cycles).

Introduction

The processing of viable microorganisms into tablets is of particular importance in the context of probiotic microorganisms (Klayraung et al., 2009). These microorganisms provide the patient with health benefits when taken in viable form and in sufficient doses (Joint FAO/WHO Working Group, 2002). Dry formulations are preferred due to their better storage stability in unrefrigerated storage and handling (Santivarangkna, 2016). However, the further processing of dried microorganisms into tablets is a challenging process step due to the compressive and shear stresses involved (Vorländer et al., 2020). Nevertheless, it is usually favored over the administration of loose powders or powders filled into capsules, as microorganisms in loose powders are more exposed to the harsh conditions in the stomach (Klayraung et al., 2009) and the production of capsules is more cost-intensive than tableting.

Gentle processes and suitable formulations are required to dry microorganisms in a life-preserving manner, typically by freeze drying, fluidized bed drying or spray drying (Broeckx et al., 2016). Despite process parameters that are as gentle as possible (especially low temperatures), high losses in the viability of microorganisms are sometimes observed since water molecules enable the correct conformation of various biological structures (Ananta et al., 2005, Crowe et al., 1987, Oliver et al., 1998). To counteract this, protective additives are added to the cell suspension before drying. These stabilize essential biological structures (theories of vitrification, water replacement and preferential hydration), whose conformational change would otherwise be accompanied by irreversible inactivation of the microorganisms, in particular through the loss of the integrity of the cell membrane (Belton and Gil, 1994, Broeckx et al., 2016, Cordone et al., 2007, Wolkers and Oldenhof, 2021). Numerous studies have dealt with the identification of effective protective additive formulations in the drying of microorganisms (Broeckx et al., 2016). A universal formulation does not yet exist. However, studies have shown that a combination of trehalose and skimmed milk powder can protect Saccharomyces cerevisiae cells from lethal damage during freeze drying, fluidized bed spray granulation and spray drying (Vorländer et al., 2023a, Vorländer et al., 2023d, Vorländer et al., 2020).

The further processing of dried microorganisms into tablets has also already been investigated in numerous studies. In some cases, aspects such as protection against bile juices or storage stability have already been addressed (Klayraung et al., 2009). However, it is first necessary to know the damage mechanisms during densification in order to be able to counteract these in a targeted manner (Vorländer et al., 2023b). Survival during tablet production has already been investigated in several studies. Various aspects have been brought into focus. These include, in particular, the formulation and its deformation behavior (Ayorinde et al., 2011, Blair et al., 1991, Byl et al., 2018, Fassihi and Parker, 1987, Plumpton et al., 1986a), but also kinetic factors of compression such as dwell time and consolidation time (Fassihi and Parker, 1987, Vorländer et al., 2023c) or the geometry of the tablets produced. The damage to the microorganisms can be of a thermal or mechanical nature (Chesworth et al., 1977). Studies with cells of different sizes indicate that the mechanical component is predominant, as larger microorganisms show lower survival (Plumpton et al., 1986b). Earlier studies by the authors of the present publication have recently shown that tablet porosity is essentially decisive for the inactivation of microorganisms, with the change in tablet porosity showing a correlation across formulations (Vorländer et al., 2023d, Vorländer et al., 2023b).

Previous studies on the tableting of viable microorganisms have generally used hydraulic presses or compaction simulators, applying compression profiles with a single main pressure. An important aspect for the transfer of the results of these studies to an industrial scale has not yet been taken into account. The high production speeds when using rotary tablet presses usually require the use of a pre-pressure and a main pressure in order to prevent the formation of tablet defects in the best possible way (Hansen and Kleinebudde, 2021, Mazel and Tchoreloff, 2020, Patel et al., 2006). In the production of tablets with probiotic microorganisms, the microorganisms contained are subjected to multiple stresses in this case. The applied pre-compression pressure is typically significantly lower than the main pressure (Patel et al., 2006). With a low pre-compression stress, densification is significantly lower there. Nevertheless, the question remains to what extent this first densification is associated with additional damage to the microorganisms.

In order to answer this question, spray-dried yeast cells are mixed with various fillers in the present study. The mixtures are densified with a compaction simulator at different compression stresses. The same compression stress is applied once, twice or five times. This extremization compared to a low initial pressure and the actual main pressure in rotary tablet presses is intended to reveal influences that would otherwise remain undetected. At the same time, the findings should further strengthen the understanding of the physical damage mechanisms during tableting.

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2. Materials and methods

2.1. Spray drying

Baker’s yeast Saccharomyces cerevisiae (Lallemand-DHW GmbH, Vienna, Austria) as a model organism was spray dried as established elsewhere (Vorländer et al., 2023d). In brief, a suspension with a cell dry weight concentration (CDW) of 50  g/L and a concentration of 50  g/L trehalose dihydrate (FormMed HealthCare AG, Frankfurt am Main, Germany) and 50  g/L skimmed milk powder (Carl Roth GmbH + Co. KG, Karlsruhe, Germany) was prepared and spray-dried after one hour of incubation at room temperature. Spray drying was conducted in co-current (ProCepT 4 M8-TriX, PROCEPT nv, Zele, Belgium). Before the cell suspension was sprayed, water was sprayed for at least 15 min to bring the entire system to a state of equilibrium (temperature and humidity). The flow rate was adjusted so that the same mass flow of vaporizable water was sprayed as during the spraying of the cell suspension. In order to limit the thermal stress on the product, the product was removed after every 60 min of drying time. The inlet temperature was 100 °C, the volume flow of the drying air 0.3  m3/min, the mass flow of the cell suspension 2  g/min, the nozzle diameter 1.2  mm and the nozzle pressure 1.5  bar. In addition, 0.12  m3/min air was supplied to the cyclone to reduce the separation limit.

2.2. Preparation of powder blends

The spray-dried product was mixed (3D shaker mixer TURBULA, Willi A. Bachofen AG at 49  min−1, 5  min) in a mass ratio of 1:3 with the fillers dicalcium phosphate (DCP, DI-CAFOS A150, kindly provided by Chemische Fabrik Budenheim KG, Budenheim, Germany), isomalt (ISO, GalenIQ 721, kindly provided by BENEO GmbH, Mannheim, Germany), lactose (LAC, Granulac 70, kindly provided by MEGGLE GmbH & Co. KG, Wasserburg am Inn, Germany) or microcrystalline cellulose (MCC, Vivapur 102, kindly provided by J. Rettenmaier & Söhne GmbH + Co KG, Rosenberg, Germany). In the case of DCP, ISO and LAC, 1 wt-% of magnesium stearate (MgSt, MAGNESIA GmbH, Lüneburg, Germany) was added as a lubricant and mixed for a further 2  min. MCC required no lubrication as ejection forces were low and tool wear is correspondingly low even without addition of MgSt, which is known to negatively affect the tensile strength of MCC tablets (Puckhaber et al., 2022).

2.3. Preparation of tablets

A compaction simulator (Styl’One evolution, MEDELPHARM, Beynost, France) was used to produce the tablets. This is instrumented with force and displacement sensors and enabling the calculation of in-die porosity data during compaction. It was equipped with flat, round punches with a diameter of 11.28  mm. The die was filled manually in order to keep mass fluctuations as low as possible. The target mass of the tablets was 450  mg. The compression was displacement-controlled with a generic, symmetrical trapezoidal compression profile with a constant speed of 45  mm s−1 of the upper and lower punch and dwell times between 30 and 40  ms. The compression height was adjusted so that compression stresses in the range of 25 to 300  MPa were applied. Initially, tableting was carried out with a singular compression with and without pre-compression. The pre-compression stress was 10 % of the main compression stress. For another batch, compression was performed once in one test series and twice or five times in the two other test series. In the case of multiple compressions, the tablet remained in the die between the compression repetitions and was only ejected after the last compression. In order to achieve the respective target compression stress with an increasing number of compression repetitions, the compression height was reduced accordingly with each compression repetition. In contrast to multiple compression, the results of compaction with and without pre-compression show hardly any differences and are therefore not considered in more detail in this manuscript. For reference purposes, most diagrams from results and discussion section are shown in the appendix analogously for compaction with and without pre-compression (Suppl. 1 – Suppl. 6).

Karl Vorländer, Arno Kwade, Jan Henrik Finke, Ingo Kampen, Influence of multiple compression phases during tableting of spray dried Saccharomyces cerevisiae on microbial survival and physical–mechanical tablet properties,
International Journal of Pharmaceutics, 2024, 124948, ISSN 0378-5173, https://doi.org/10.1016/j.ijpharm.2024.124948.