
==== Front
Ophthalmol Ther
Ophthalmol Ther
Ophthalmology and Therapy
2193-8245
2193-6528
Springer Healthcare Cheshire

39225713
1023
10.1007/s40123-024-01023-9
Original Research
Overview of 37 Tear Substitutes in Europe Based on Various Physicochemical Parameters
Paschier Adrien adrien.paschier@horus-pharma.com

Manuelli Aurélie
Chauchat Laure
Legall Morgane
Rebika Hayette
Sahyoun Marwan
Guerin Camille
Laboratoires Horus Pharma, 22 Allée Camille Muffat, 06200 Nice, France
3 9 2024
3 9 2024
10 2024
13 10 27992812
18 7 2024
20 8 2024
© The Author(s) 2024
2024
https://creativecommons.org/licenses/by-nc/4.0/ Open Access This article is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License, which permits any non-commercial use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by-nc/4.0/.
Introduction

Dry eye disease (DED) is one of the most prevalent eye conditions worldwide, with artificial tears serving as a primary treatment option. Despite their wide availability on the European market, there is a lack of established classifications based on their physicochemical properties. The aim of our study was therefore (i) to develop an analytical method that measures the concentration and the molecular weight (MW) of the hyaluronic acid (HA) in commercialized products, and (ii) to propose an overview based on their various physicochemical parameters.

Methods

The intrinsic viscosity and MW of the HA, as well as osmolarity, pH, rheological profile, and viscosity, were measured or determined. A specific method was developed to measure the average intrinsic viscosity and HA content using a liquid size-exclusion chromatography system. The MW was determined using the Mark–Houwink equation.

Results

Thirty-seven products commercialized in Europe were analyzed, with 21 of them containing HA. The HA MW was lowest (300 kDa) for Thealose®, Thealoz Duo® Gel, and Hyabak®, and highest (1300 kDa) for Vismed® Multi, Vismed® Gel, and Neovis® Gel. The pH values varied between 5.94 for Treovis® and 8.06 for Systane® Ultra. Osmolarity ranged between 148 mOsm/L and 325 mOsm/L for Neovis® and Treovis®, respectively. Viscosity was highly variable, ranging from 0.38 mPas·s for Hylolipid® to 337.47 mPas·s for Thealoz® Duo Gel. Finally, rheological profile analysis revealed different shear-thinning behaviors.

Conclusion

While the perfect eye drop does not exist, a multitude of options are available to choose from. This study improves our understanding of the major tear substitutes available on the European market based on several physicochemical properties. A better understanding and awareness of these parameters is crucial in order to offer the best treatment for patients with DED.

Supplementary Information

The online version contains supplementary material available at 10.1007/s40123-024-01023-9.

Keywords

Dry eye disease
Tear substitutes
Artificial tears
Hyaluronic acid
Molecular weight
Analytic method
Eye drop
Physicochemical parameters
issue-copyright-statement© Springer Healthcare Ltd., part of Springer Nature 2024
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pmcKey Summary Points

Why carry out this study?	
Dry eye disease is one of the most prevalent eye conditions worldwide, with artificial tears serving as a primary treatment option.	
There is currently no established or standardized classification comparing the different tear substitutes available on the market.	
Comparison of the quality of hyaluronic acid can be challenging, as no recommended values exist, and methods of measurement vary among different papers, making inter-study comparison irrelevant.	
What was learned from this study?	
Wide variability in the measured parameters was observed among the different tear substitutes.	
This variability is predominantly determined by the variations in the viscosity and the presence or the absence of hyaluronic acid.	
A new analytical method was developed that compared the different concentrations and molecular weights of hyaluronic acid inside the eye drops.	

Introduction

Dry eye disease (DED) is a chronic multifactorial disease [1] of the tears and the ocular surface and is one of the most prevalent eye conditions worldwide. Its definition has undergone several modifications since 1995, culminating in the latest update by the International Dry Eye Workshop in 2017 as “a multifactorial disease of the tears and ocular surface that results in symptoms of discomfort, visual disturbance, and tear film instability with potential damage to the ocular surface. It is accompanied by increased osmolarity of the tear film and inflammation of the ocular surface” [2].

Currently, the global prevalence of DED is estimated at between 5% [3] and 57% [4] in the general population, and is more common in women and in the elderly [5].

Treatment of DED is aimed at restoring and stabilizing the tear film, minimizing dryness, and leading to improved ocular surface protection [6]. In 2007, Baudouin et al. [7] proposed the concept of a “vicious cycle of inflammation” as a core driver in the pathophysiology of DED, including tear film instability, hyperosmolarity, corneal and conjunctival cell apoptosis, and chronic inflammation. Although breaking the vicious cycle (with anti-inflammatory or secretagogue agents [8]) was reported as an important step in the management of DED, the mainstay of treatment remains artificial tears or lubricant eye drops [9].

Since 1986 and the assessment by Mengher et al. [10] of the benefits of hyaluronic acid (HA) on tear film stability, HA has become a standard component in many artificial tears [11], with preclinical and clinical studies showing acute and long-term therapeutic benefits. HA is a natural glycosaminoglycan present in the human body, composed of a sequence of repeating disaccharide units of glucuronic acid and N-acetyl-D-glucosamine. Its molecular weight (MW) varies based on the number of disaccharide units forming the molecule, ranging from less than 100 kilodaltons (kDa) to over 1000 kDa.

Studies have shown that high MW HA (HMWHA, > 1000 KDa [11–15]) has greater therapeutic potential, including better hydration, improved mucoadhesion to the cornea [13], and anti-inflammatory properties [15]. In contrast, low MW HA (LMWHA, < 500 KDa [16–18]) tends to act as a potent pro-inflammatory molecule [17]. Comparison of the quality of HA can be challenging, as no recommended values exist, and methods of measurement vary among different papers, making inter-study comparison irrelevant.

Other interesting factors for eye drops can include viscosity, osmolarity, pH, and rheological profile. High viscosity increases the retention time of an eye drop on the ocular surface, while a pseudoplastic profile improves the spreadability of the tear substitute, thereby enhancing the wettability and efficacy of the eye drop.

While many commercialized artificial tears are available on the European market, no classification has been established to differentiate them based on these parameters.

The aims of our study were therefore (i) to develop an analytical method measuring the MW and the concentration of the major HA-based eye drops indicated for the treatment of DED and commercialized in Europe, and (ii) to measure and determine various parameters (HA intrinsic viscosity and MW, pH, rheological profile, and viscosity) of artificial tear substitutes containing or not containing HA, and finally, proposing an updated overview based on these parameters.

Methods

In this study, ethical approval was not necessary because the research did not involve human participants, human data, or animals.

Products

Thirty-seven eye drops commercially available on the European market were evaluated. Twenty-one of them contained HA of various MW and concentrations. The tear substitutes were selected by general consensus among the authors and ophthalmologists to provide a comprehensive range of formulations recommended for DED management. These include formulations with varying complexity, including medicines and medical devices, products with or without HA, solutions to emulsions, oil phase and gels, preservative-free and preserved formulations, simple formulations, and formulations with risk of interference. This approach allows for representing the versatile therapeutic arsenal available across the European territories, with a focus on France, Spain, Belgium, the Netherlands, and Luxembourg.

The evaluated eye drops are listed in Table 1. Table 1 List of eye drops commercially available on the European market that were evaluated

Brand name	Company	HA concentration claim (%)	Type of product	Preserved	Format	
Acuolens®	Alcon	–	Solution	–	SD	
Aquoral®	Esteve	0.40	Solution	–	10 mL (MD)	
Artelac® Rebalance	Bausch & Lomb	0.15	Solution	Yes	10 mL (MD)	
Artelac®	Bausch & Lomb	–	Solution	–	SD	
Cationorm®	Santen	–	Emulsion	–	10 mL (MD)	
Celluvisc®	Abbvie	–	Solution	–	SD	
Clinadry® Multi	ID Phar	0.40	Solution	–	SD	
Dulcilarmes®	Horus	0.05	Solution	–	10 mL (MD), SD	
Elixya®	Bausch & Lomb	0.15	Solution	–	10 mL (MD)	
Fluidabak®	Théa	–	Solution	–	10 mL (MD)	
Hyabak®	Théa	0.15	Solution	–	10 mL (MD)	
Hylo® Fresh	Ursapharm	0.03	Solution	–	10 mL (MD)	
Hylo® Lipid	Ursapharm	–	Oil phase	–	10 mL (MD)	
Lacrifluid®	Europhta	–	Solution	–	SD	
Matrix 3® Ocular	Brill Pharma	0.15	Solution	–	10 mL (MD)	
Neovis®	Horus	0.18	Solution	–	15 mL (MD)	
Neovis® Gel	Horus	0.30	Gel	–	15 mL (MD)	
Neovis® Total Multi	Horus	0.18	Emulsion	–	15 mL (MD), SD	
Nereya®	Bausch & Lomb	0.24	Emulsion	–	10 mL (MD)	
Nutrivisc®	Alcon	–	Solution	Yes	SD	
Optiben®	Cinfa	0.20	Solution	Yes	10 mL (MD)	
Optive® Fusion

Optava® Fusion

	AbbVie	0.10	Solution	Yes	10 mL (MD)	
Optive® Plus	AbbVie	–	Solution	–	10 mL (MD)	
Optive®	AbbVie	–	Solution	Yes	10 mL (MD)	
Refresh®	AbbVie	–	Solution	–	SD	
Treovis®	S-LAB	0.15	Solution	–	10 mL (MD)	
Systane® Balance	Alcon	–	Emulsion	Yes	10 mL (MD)	
Systane® Complete	Alcon	–	Solution	–	10 mL (MD)	
Systane® Hydration	Alcon	NC	Solution	Yes	10 mL (MD)	
Systane® Ultra	Alcon	–	Solution	–	10 mL (MD)	
TheaLipid®	Théa	–	Emulsion	–	10 mL (MD)	
Thealose®	Théa	0.15	Solution	–	15 mL (MD)	
Thealoz® Duo Gel	Théa	0.15	Gel	–	SD	
Treovis®	Horus	0.15	Solution	–	10 mL (MD)	
Vismed® Gel Multi	TRB	0.30	Gel	–	15 mL (MD)	
Vismed® Multi®	TRB	0.18	Solution	–	15 mL (MD)	
Vitadrop®	Densmore	0.15	Solution	Yes	10 mL (MD)	
HA hyaluronic acid; SD single dose; MD multi-dose; NC not claimed

Intrinsic Viscosity/MW and HA Content

HA is a polymer available as a mixture of chains of different lengths. The measured MW is therefore a description of the distribution of different chain lengths present in the mixture. The mean MW to describe the distribution can be assessed focusing on the number of chains, their weight, or their volume, resulting in different values to describe the distribution. Very wide variation in the MW of the same HA has been observed in literature reports, largely depending on the measurement method and the calibration system used [18]. Using HA-based calibration provides a more reliable result than other types of calibration, such as polyethylene oxide.

On the other hand, MW is not usually measured by HA suppliers, and no harmonized method for measurement exists. However, HA MW is correlated with intrinsic viscosity (Mark–Houwink equation), which is a standard measurement method used as a quality control according to the European Pharmacopoeia (EP) [19]. Consequently, the intrinsic viscosity of the HA raw material was used as a calibration system to determine the intrinsic viscosity of the different tested HA-based products.

MW was then derived from intrinsic viscosity results using the Mark–Houwink equation:η=κMα,

where η is the intrinsic viscosity, M is the MW, and κ and α are solvent-polymer-dependent fitting parameters (constants used from the Japanese Pharmacopeia XVII [20]).

Recovery was calculated as the percentage ratio of the measured HA content and the HA concentration claimed for the product.

Six raw material HA powders (Bloomage Biotechnology, EP grade) with intrinsic viscosities ranging from 0.1 m3/kg to 2.5 m3/kg were used as reference standards. N-Acetyl glucosamine was used as reference standard for HA quantification.

The average intrinsic viscosity and HA content were measured externally on a liquid size-exclusion chromatography system (PL aquagel-OH 60 column, 15 µm, 7.5 µm internal diameter × 300 mm length; Agilent, Waldbronn, Germany) equipped with a Shimadzu RID-10A refractive index detector (Duisburg, Germany) with 0.05% sodium azide as mobile phase (flow rate 1 mL/min, run time 17 min).

As HA concentration is a critical parameter for MW determination, the eye drops were diluted in the mobile phase according to their initial HA concentration to reach a 0.35–0.50 mg/mL HA concentration. Depending on the eye drop formulation complexity (i.e., risk for interference of ingredients with the characterization of HA, emulsions), specific sample preparation techniques including precipitation or filtration were applied, when possible, to allow single HA component characterization.

pH

The pH was measured on 20 mL of undiluted samples using a calibrated Mettler Toledo pH meter (Viroflay, France) with combined InLab® Expert Pro 2 m electrodes. The pH meter uses the relationship between the hydrogen H3O+ concentration and the electrochemical potential difference to determine the pH of the solution. The potential difference between the two electrodes is proportional to the pH level of the solution.

Osmolarity

The osmolarity was determined using a calibrated manual type 6 micro osmometer (Löser, Germany) and 100 µL of undiluted sample, measured in triplicate, for which the mean value is reported. The osmometer determines the freezing point of aqueous solutions. The lowering of this point relative to the freezing point of pure water is directly correlated to the osmotic concentration. Indeed, pure water freezes at 0 °C, while an aqueous solution with an osmolality of 1 osmol/kg freezes at −1.858 °C [21].

Rheological Properties (Viscosity and Rheological Profile)

Rheological properties of undiluted products were determined using a rotational rheometer (AR1000; TA Instruments, New Castle, DE, USA) at 25 °C using a 60 mm diameter aluminum cone. The rheological profile was assessed by varying the shear rate from 0 to 500 s–1 in a rising and then descending flow ramp to fully characterize the potential shear-thinning or shear-thickening behavior. The standard viscosity was determined using the same equipment at 25 °C and a fixed shear rate of 100 s–1. Two grams of each product were employed, and the measurements were performed in triplicate, for which the mean value is reported.

For readability purposes, results for all of the measured physicochemical parameters are separated in figures according to the country in which the tested item is commercialized.

Results

HA Content, Intrinsic Viscosity, and MW

For each of the tested HA-containing eye drops, values of the intrinsic viscosity, HA content, and the determined MW are summarized in Table 2. Table 2 Hyaluronic acid (HA) concentration measurement, intrinsic viscosity, and molecular weight of the analyzed products containing HA

Brand name	Measured HA concentration (%)	Recovery between the measured and claimed HA (%)	Intrinsic viscosity, m3/kg (from RT at 50% area)	Molecular weight (MDa)	
Aquoral®	0.42	105	1.5	0.8	
Artelac® Rebalance	0.15	102	1.3	0.7	
Clinadry® Multi	0.43	107	1.4	0.7	
Dulcilarmes®	0.05	94	1.1	0.5	
Elixya®	0.15	102	1.3	0.7	
Hyabak®	0.15	102	0.7	0.3	
Hylo® Fresh	0.03	100	1.4	0.7	
Matrix 3® Ocular	0.15	98	1.7	0.9	
Neovis®	0.18	102	2.1	1.2	
Neovis® Gel	0.31	104	2.2	1.3	
Neovis® Total Multi	0.19	105	1.6	0.9	
Nereya®	0.24	100	1.4	0.7	
Optiben®	0.19	96	1.5	0.8	
Optive®Fusion,

Optava® Fusion

	0.08	79	2.8*	1.7	
Treovis®	0.14	94	2.1	1.2	
Systane® Hydration	0.28	–	1.1	0.5	
Thealose®	0.15	102	0.7	0.3	
Thealoz® Duo Gel	0.15	97	0.7	0.3	
Vismed® Gel	0.31	104	2.2	1.3	
Vismed® Multi	0.18	102	2.2	1.3	
Vitadrop®	0.15	101	1.3	0.7	
*Interference observed → overestimated value

HA content measured in products was globally in line with the products’ HA label claims, ranging from 0.05% to 0.40%. The only exception was Optive® Fusion (labeled Optava® Fusion on the Spanish market), for which the recovery was below 80%.

Wide variability in HA intrinsic viscosities was observed, ranging from 0.7 to 2.2 m3/kg (corresponding to MW from 300 to 1300 kDa).

Based on these results, HA-containing eye drops were classified according to their MW and HA concentration. They are summarized in Fig. 1.Fig. 1 Classification of the major commercialized eye drops in Europe containing hyaluronic acid (HA) according to the claimed HA concentration and measured molecular weight. Values for Optive® Fusion (known as Optava® Fusion in Spain) are overestimated. For readability purposes, results are separated according to the country in which the tested items are commercialized. A France; B Spain; C Belgium, Luxembourg, Netherlands. kDa kilodalton; NC not claimed

pH

Measured pH values ranged between 5.94 for Treovis® and 8.06 for Systane® Ultra. They are summarized in Supplementary Table 1 in the Supplementary Materials.

Osmolarity

All measured osmolarity values (mean interquartile range [IQR]) ranged between 148 [2] for Neovis® and 325 [7] mOsm/L for Treovis®. They are summarized in Supplementary Table 1 and Supplementary Fig. 1.

Viscosity

The range of viscosity (mean [IQR]) for the medical devices available was from 0.38 [0.06] mPas·s for Hylolipid® to 337.47 [1.13] mPa·s for Thealoz® Duo Gel. The results are presented in Supplementary Table 1 and Supplementary Fig. 2.

Rheological Profile

The rheological profile measurements confirmed that all products exhibit shear-thinning behavior. However, the degree of this effect varies depending on the viscosity of the product.

Discussion

Our study analyzed the major tear substitute brands available in the European market in order to propose a new classification based on various measured parameters, including intrinsic viscosity, MW, osmolarity, rheological profile, viscosity, and pH. One major challenge was to find a method that could measure the MW of HA for an entire range of formulations (solutions, emulsions, and gels), with varying potentially interacting excipients and different HA MW and concentrations. Our results confirmed the wide variability in HA MW and concentrations used in the market. There are no official cutoff values for HMWHA and LMWHA. However, the literature seems to agree that MW greater than 1000 kDa is high [11–14] and MW below 500 kDa [16, 17] is low. Based on our analytical method and measurements, Thealose® and Hyabak® were found to have the HA with the lowest MW (300 kDa), whereas Vismed® Gel, Neovis® Gel, and Vismed® Multi were found to have the highest MWHA (1300 kDa). Identification of the MW is a guarantee of the HA quality and efficacy as high- and low-MW HA act as anti- and pro-inflammatory mediators, respectively [22–24]. Masuko et al. [12] and Lennon et al. [14] showed that HMWHA exhibits anti-inflammatory properties in arthritis and lung pathobiology respectively. Also, HMWHA anti-inflammatory properties are of particular interest in DED, because inflammation plays a key role in its pathogenesis [25].

In preclinical studies and animal models, HMWHA has been shown to reduce the expression of genes associated with classically activated macrophages, leading to a decrease in the immunological response [17]. It has also been found to increase mucoadhesion of the tear substitute, resulting in a better retention on the ocular surface [13, 16, 26], and has shown wound-healing and cell-migration properties [27], with a superior cytoprotective effect compared to LMWHA [28, 29]. On the other hand, LMWHA was shown to be pro-inflammatory [17, 30] due to the stimulation of proteins involved in the inflammation process (such as TNF-alpha and IL-6 [17]), but also due to its possible aggregation in tissue injury [31].

In clinical studies, HMWHA was compared to individual optimum artificial tears over a period of 8 weeks in a multicenter randomized trial involving 84 patients [32] and showed a significant improvement of symptoms, without signs of dry eye impairment. A post hoc analysis [33] also showed that eye drop application frequency was significantly reduced in the HMWHA group, which could reduce the socioeconomic burden of the disease. When applied topically in severely dry eyes, HMWHA was also shown to cross the epithelial barrier and to have a neurotrophic effect on the sub-basal nerve plexus [34]. Finally, Beck et al. [35] conducted a randomized comparative study with 11 patients, providing a compelling comparison between HMWHA and autologous serum. Even though their results should be interpreted with caution given the limited number of patients, they highlighted that while no superiority was demonstrated, HMWHA was deemed an acceptable alternative to autologous serum.

In addition to the MW, the HA concentration inside of an eye drop also has to be considered. In fact, a higher HA concentration significantly improves the tear breakup time and the ocular staining compared to a product with the same MW but with a lower concentration [36]. In experimental DED, 0.30% HA artificial tears were more effective than lower concentrations in improving tear film stability and ocular surface staining and irregularity, increasing conjunctival goblet cell density, and decreasing corneal epithelial apoptosis [37]. Calonge et al. [38] compared two concentrations of HMWHA (0.30 and 0.18%) in patients with moderate to severe DED and showed that while both were effective in reducing signs and symptoms, a numerical decrease in the frequency of instillation and significantly faster conjunctival healing were noted for the 0.30% HA group. Also, corneal epithelial wound healing in pterygium excision was faster in the 0.30% HA group than in the 0.18% HA group [39].

Concerning osmolarity, most of the tested products were hypo-osmolar (< 280 mOsm/L) On the other hand, several eye drops, (Treovis®, Optive®, Optive fusion®, Lacrifluid® and Celluvisc®) demonstrated osmolarity superior to 308 mOsm/L, which is known to be the tear film osmolarity [40]. However, Liu et al. [41] determined that no administration discomfort occurs at an osmolarity below 450 mOsm/L. Moreover, it has been observed that the eye recovers its baseline osmolarity within 10 min after isotonic or hypotonic solution administration [42]. Hyperosmolarity of the tear film is a well-known clinical sign of DED [40, 43, 44]. Even though the hypo-osmolarity parameter of an eye drop cannot manage DED on its own [45], it can help to better reduce tear osmolarity and, especially when combined with HMWHA, better alleviate patients’ symptoms.

Rheological analysis demonstrated that all the analyzed products exhibited shear-thinning behavior. However, this was more pronounced in products such as Vismed® Gel and Vismed®, due to their higher viscosities. This analysis also confirms the pseudoplastic nature of HA [46]. The shear-thinning behavior of the tear film has already been documented [47]. The decrease in viscosity during blinking allows for better spreading of the tear film over the ocular surface. However, when the eye is open, the increased viscosity helps to maintain the film on the ocular surface, providing better protection and lubrication. This behavior ensures that the tear film lubricates the eye in the most efficient manner [47, 48]. In that sense, the shear-thinning properties of artificial tears are of interest for patients with DED facing tear film instability, to allow proper spreadability of the eye drops onto the ocular surface.

A broad range of viscosities was observed for the analyzed products, ranging from 0.38 mPa·s for Hylolipid® to 337.47 mPa·s for Thealoz® Duo Gel. It is important to note that the optimal formulation of an eye drop should be viscous enough to ensure long retention time but not excessively so, to avoid causing blurred vision [49]. Additionally, products with excessive viscosity cannot be packaged in multi-dose containers, as is the case with Thealoz® Duo Gel, Lacrifluid®, and Celluvisc®. It is well known that high viscosity allows for a longer retention on the ocular surface, enabling better lubrication of the eye and decreasing the frequency of instillation [47]. This high viscosity can be achieved through the use of different components such as HA, carbomer [50], or carboxymethyl cellulose [51]. While carbomer exhibits better mucoadhesion than cellulosic derivatives, it can increase blurry vision and lead to deposits on the eyelashes during gel drying [52]. HA emerged as a preferred component due to its various beneficial effects described above and to its shear-thinning behavior. As an endogenous component of the ocular surface, its retention time is longer than that of cellulosic derivatives and vinyl polymers [53]. The combination of high viscosity with pseudoplastic behavior allows for longer retention on the ocular surface, better spreadability on the ocular surface during blinking, and improved lubrication and protection of the eye [47].

Concerning pH values, it is well known that a healthy tear film pH is around 7.5 [54]. Moreover, the tear film has a buffering effect, and its high renewal rate helps protect the eye when exposed to slightly acidic (pH > 3.5) or basic solutions (pH < 10.5) [55, 56]. In our study, all measured pH values are considered to be within the physiological range [54].

Achieving an optimal eye drop formulation requires taking into consideration all of these different factors. Considering the current knowledge of the pathophysiology of DED, where tear film is altered and signs include hyperosmolarity, when choosing an eye drop product for patients, the study provides insight into the different studied parameters to support DED management.

Eye drops containing HA present an advantage compared to other polymers, given its endogenous nature and with its viscoelastic and water-absorbent properties, especially with an altered tear film. Higher MW is interesting for increased retention time and anti-inflammatory properties. Additionally, the rheological properties of the eye drop can also increase retention time with higher viscosity, while considering that very viscous solutions and gels can also induce blurring vision and should be preferred at night. Having viscoelastic properties of the eye drop, mimicking the tear film behavior, ensures good integration and spreadability during blinking.

Finally, lower osmolarity seems to be beneficial considering that DED is associated with hyperosmolarity of the tear film, and having a physiological pH ensures good patient tolerance of the eye drop.

However, as our understanding of the pathophysiology of DED continues to improve, eye drop formulations are becoming more complex, and other components with additional pharmacological or mechanical effects are being added, such as antioxidants (alpha lipoic acid, vitamin B12, etc.), osmo-protectants (trehalose), ocular surface modulators (dexpanthenol), or lipid components. The inclusion of preservatives also needs to be carefully considered due to their potential impact on the safety and efficacy of the formulations.

There are some limitations to our current study.

Our challenge was to develop an analytical method capable of characterizing the MW of HA across a large formulation scope. A diverse selection of formulations was chosen, allowing the start of analytical development. Having a common method for all products is interesting but challenging, and interferences with other polymers with the same MW as the measured HA were noted. Sample preparation was critical in this case to allow separation with interfering polymers when possible. When this was not possible, the interference affected the accuracy of our analytical method; this was the case for Optive® Fusion, highlighted with a low HA recovery of 79%. The HA peak overlapped with sodium carboxymethylcellulose present in the formulation. As a result, only a partial integration of the HA peak could be performed (of the portion with the highest MW), resulting in a low recovery and an overestimation of the MW for this product. However, having a common method for all tear substitutes, with individual determination of the recovery, is a good way to ensure unbiased evaluation of the results.

Moreover, it is important to note that the list of tested products represents a broad selection available on the European market. However, it is not exhaustive, and it would be interesting to test additional products to provide a complete overview of the tear substitute therapeutic arsenal across Europe.

Additionally, this study investigated several physicochemical parameters of clinical interest in general practice. However, the study cannot be considered as exhaustive for conclusions regarding the clinical recommendations for eye drop efficacy. Indeed, the formulation of the eye drops can have a direct impact on clinical efficacy, with synergic or antagonist effects. In that case, an individual assessment of each formulation should be performed. The use of additional components in the formulation, such as the presence or absence of preservatives, must be considered due to their known toxicity to the ocular surface [57]. The use of antioxidants, such as lipoic acid, osmo-protectants such as trehalose, alternative polymers as viscosifying agents, or lipidic components in emulsions for better integration into the different lacrimal layers, can also affect the benefit on the ocular surface. The study of single and combined effects of the different formulation components is an axis that could be explored as well, as initiated by the research conducted by Černohlávek et al. [58].

Finally, while our study is analytical, a preclinical study on cellular models such as a mucoadhesion study may provide a deeper understanding of component synergy. Moreover, studies combining our physicochemical results with the analysis of the different components in the various eye drops are necessary. This will enable the proposal of recommendations to physicians, helping them to provide better management of patients with DED. Additionally, conducting a clinical study would further clarify the impact of various eye drop parameters on efficacy and safety, leading to a more accurate classification. Our study highlights the existence of variability among the various tear substitutes available. This variability can be based on different parameters such as the osmolarity or the pH, but it is predominantly determined by the variations in the viscosity and the presence or the absence of HA. Furthermore, products containing HA can possess various types of HA based on the MW, which can impact the product’s behavior. While LMWHA is effective, HMWHA has demonstrated greater improvements in DED signs and provided better protection of epithelial cells, ultimately improving patient care.

Conclusion

This study provides a new overview of 37 tear film substitutes available in Europe, based on several parameters including viscosity, rheological profile, pH and MW of HA. While the perfect eye drop does not exist, a multitude of options are available to choose from. Better understanding and awareness of tear substitute formulation is crucial to enabling the best treatment to be offered for patients with DED.

Supplementary Information

Below is the link to the electronic supplementary material.Supplementary file1 (PDF 306 KB)

Acknowledgements

We would like to thank our experienced external laboratory partner for the HA molecular weight method development and the measurement. For confidentiality reasons, the name of the laboratory cannot be disclosed. The laboratory is accredited according to international standards and follows strict protocols to ensure the reliability and accuracy of the results.

Author Contributions

Aurélie Manuelli and Camille Guerin contributed to study conception, design, result discussion, and article outlines. Morgane Legall performed experimental work and contributed to the design and execution of the experiments. Adrien Paschier, Hayette Rebika, Marwan Sahyoun and Laure Chauchat contributed to result discussion and article outlines. All authors reviewed and approved the manuscript. All authors meet the International Committee of Medical Journal Editors (ICMJE) criteria for authorship for this article, take responsibility for the integrity of the work as a whole and have given their approval for this version to be published. The study was sponsored by Horus Pharma (Nice, France).

Funding

Sponsorship for this study and the journal’s Rapid Service Fee were funded by Laboratoire Horus Pharma, Pharmaceutical Company (Nice, France).

Data Availability

The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.

Declarations

Conflict of Interest

All authors (Adrien Paschier, Aurélie Manuelli, Laure Chauchat, Morgane Legall, Hayette Rebika, Marwan Sahyoun, Camille Guerin) were employed by Horus Pharma at the time of the article publication.

Ethical Approval

In this study, ethical approval was not necessary because the research did not involve human participants, human data, or animals.
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