[version 1; peer review: 1 approved with reservations]
No competing interests were disclosed.
Phacoemulsification is one of the most frequent surgeries in the world. However, prolonged use of phacoemulsification machines produces reactive oxygen species which will damage corneal endothelial cells. Ascorbic acid has an antioxidant capacity to neutralize oxidative stress in the anterior chamber. This study will investigate the protective effect of ascorbic acid on corneal endothelial cells in patients with hard nuclear cataracts.
This study is a double-blinded randomized controlled trial. Samples will be divided into three groups, and 500 mg of vitamin C, tid, or placebo will be received for seven weeks. Clinical characteristics, ascorbic acid, malondialdehyde, and total antioxidant capacity of patients in serum and aqueous humor will be measured before and after intervention and phacoemulsification.
Data from this study will reveal the protective effect of oral vitamin C supplementation on the corneal endothelial cells in patients with hard nucleus cataracts.
This trial has been registered at
Cataract is the most probable cause of blindness in Indonesia. Universally, WHO estimated that 51% of blindness in the world is caused by cataracts and there are 20 million people who are blind from cataracts. In 2020, the prevalence of blindness in Indonesia is 1.4% with 81.2%, or three million, being from cataracts.
Phacoemulsification is the most used cataract extraction method, with a high success rate of 80.1% compared to extracapsular cataract extraction (ECCE) and intracapsular cataract extraction (ICCE).
Free radicals bind with antioxidants in tissues. The imbalance between the number of free radicals and antioxidants in the tissue causes oxidative stress.
Corneal endothelial cells (CEC) have little to no regeneration capacity, hence CEC damage is irreversible. Naturally, corneal endothelial cells diminish 0.3-0.6% yearly.
Total antioxidant capacity (TAC) portrays the capacity of a tissue to neutralize oxidants. As much as 73% of TAC in intraocular tissue is ascorbic acid (AA).
To our knowledge, there is yet a clinical study to investigate the protective effect of oral AA in hard nucleus cataracts. This manuscript describes the protocol for a clinical trial to analyze the protective effects of oral ascorbic acid on the corneal endothelial layer undergoing oxidative stress due to phacoemulsification in patients with hard cataracts.
This is a double-blinded, randomized controlled trial investigating the protective effect of ascorbic acid supplementation on corneal endothelial cells undergoing phacoemulsification. The study will follow the SPIRIT (Standard Protocol Items: Recommendations for Interventional Trials) guidelines.
This study is going to be conducted in the Faculty of Medicine, University of Indonesia with samples from a regional hospital in Cianjur, West Java and Serang, Banten, Indonesia.
The primary aim of this study is to analyze the protective effect of oral ascorbic acid supplementation on corneal endothelial cells which undergo oxidative stress due to phacoemulsification in patients with hard nucleus cataracts. This study will investigate clinical parameter changes in all three intervention groups, including endothelial cell density (ECD) loss, central corneal thickness, visual acuity, and cells in the anterior chamber.
This study will investigate differences in MDA levels in aqueous humor, prior to and after phacoemulsification, and compare between groups with vitamin C supplementation and those without. We will also investigate differences in AA, TAC, and MDA levels in serum and aqueous humor at the start of the study and after seven days of supplementation and compare them between the three groups. Furthermore, we will explore the correlation between aqueous humor and serum AA, TAC, and MDA levels.
The target population of this study is patients with hard nucleus cataracts who undergo phacoemulsification. Patients are eligible to be included in this study with the following inclusion criteria: aged above 60 years, have senile cataract in one or both eyes with Lens Opacities Classification System (LOCS) III nuclear opacity grade 4-6 and nuclear color grade 4-6, willing to consume prescription and participate in the study follow up for seven weeks starting from recruitment, and signed the informed consent. Written informed consent was obtained from all participants prior to enrollment in the study.
The exclusion criteria of this study are as follows: patients with vitamin C allergy or sensitivity, undergo intraoperative complications of postoperative infections, have corneal endothelium disorder, history of intraocular surgery, glaucoma, trauma, or other intraocular inflammations, have diabetes mellitus and kidney disorders, or consumes vitamins as daily supplements.
Samples will be dropped out during the study if they did not participate in the study follow-up or consume less than 80% of the prescribed medications.
We screen patients who sign up for a mass cataract surgery event in a regional hospital in Cianjur, West Java, Indonesia. Patients will be asked for information regarding personal identification, history of systemic and ocular diseases, and allergies. They then will undergo ophthalmology examination including uncorrected visual acuity (UCVA) and best corrected visual acuity (BCVA), intraocular pressure using non-contact tonometry (NCT), biomicroscopy slit lamp, and indirect fundoscopy to evaluate anterior and posterior chambers. Lens opacity will then be measured using LOCS III criteria. We will also examine ECD, coefficient of variety (COV), and hexagonality of endothelial cell layers using a specular microscope (REM 4000, Rodenstock, Germany), central corneal thickness (CCT) using Anterior Segment Optical Coherence Tomography (ASOCT), and biometry. Blood samples will be then drawn from patients to measure serum AA, MDA, and TAC.
Patients will be divided into three groups, each with 40 samples. The first group (n=40) will receive oral vitamin C, tid for one week before surgery to four weeks after surgery. The second group (n=40) will receive oral vitamin C, tid for one week before surgery and a placebo after surgery. The last group (n=40) will receive a placebo one week before surgery to four weeks after surgery. Every phacoemulsification is operated by equally experienced surgeons and identical phacoemulsification machines and consumables.
Patients will be asked to come a week after screening. Firstly, they asked the side effects of the supplementation, including gastrointestinal disorders (nausea, diarrhea, epigastric pain). Second blood samples will be then drawn to measure post-intervention serum AA, MDA, and TAC.
During surgery, after the initial incision, 0.3 mL of aqueous humor will be taken using a 1 cc syringe with a 30 G cannula. Phacoemulsification will then be conducted using standard operating procedure to intraocular lens implantation. Another 0.3 mL of aqueous humor will then be taken three minutes after removing the viscoelastic as a post-phacoemulsification sample. All patients will receive topical medications: levofloxacin 5 mg/mL every three hours for one week, prednisolone acetate 10 mg/mL every three hours for 1 week, and tapered down in 4-6 weeks.
Every patient will be evaluated one day, one week, four weeks, and six weeks post-phacoemulsification to measure study parameters and signs of complications, i.e. inflammation or infection. Patients who experience side effects or complications, including toxic anterior segment syndrome (TASS) or endophthalmitis during the duration of follow-up will be exempted from the study and managed according to the standard operational procedure of the Department of Ophthalmology, Faculty of Medicine, University of Indonesia.
Participants receive IDR100,000 at the end of the last follow-up.
Data from this study will be recorded and analyzed by IBM SPSS 20.0. Sample personal identification, including demographic and clinical characteristics, will be reported in a descriptive analysis. Data distribution of numeric variables will be measured using the Saphiro-Wilk test and will be represented in mean and standard deviation if the distribution is normal, and median and range if the distribution is not normal. Nominal data will be reported in frequency and percentage. We will use Pearson’s chi-squared test for categorical data. Independent T-test or Mann-Whitney test will be used for comparing numeric data, and paired T-test or Wilcoxon test will be used for paired numeric data. Pearson’s test will be used to test the correlation between data in aqueous humor and serum.
The findings from this study will be disseminated through presentations at national and international conferences and submitted for publication in peer-reviewed journals. Trial outcomes will also be reported in the trial registry (
The results of this study will provide data on the protective effect of vitamin C supplementation for corneal endothelial cells in patients with hard nucleus cataracts undergoing phacoemulsification. This will initiate further investigation for the alternative management for permanent endothelial decompensation or bullous keratopathy.
At the time of manuscript submission (September 2025), the study is in the data processing phase. Trial registration has been completed (
This study was approved by the the Faculty of Medicine, University of Indonesia – Cipto Mangunkusumo Hospital Ethics Committee with reference number KET-1252/UN2.F1/ETIK/PPM.00.02/2024 (protocol number: 24-08-1198). All participants provided written informed consent prior to participation.
No data are associated with this article as it describes a study protocol. Data generated from the study will be made openly available upon completion.
No underlying data are associated with this article as it describes a study protocol.
The completed SPIRIT 2013 checklist for this study is publicly available in Zenodo: SPIRIT checklist for “The Effects of Vitamin C Supplementation on Corneal Endothelial Damage in Hard Cataract Phacoemulsification: an Oxidative Stress Study on Aqueous Humour and Corneal Endothelial Cell Characteristics, A Randomized, Double-Blinded, Clinical Trial”. Zenodo. DOI:
License:
The study protocol was well designed. The study protocol lacks some details:
1. Level of significance not to mention: Before the statistical analysis of the data, significance level must be defined like P < 0.05 or P < 0.01
2. detailed procedure or citation not available for estimation of AA, MDA, and TCA. For reproducibility of the research, author must be clear defined the assay procedures, techniques, and methods used to measure the biochemical parameters.
3. Oxidative stress markers are measured in serum only, while Phacoemulsification produces free radicals on eyes. so explain any justification or try to evaluate oxidative stress markers in aqueous humor also.
Is the study design appropriate for the research question?
Yes
Is the rationale for, and objectives of, the study clearly described?
Yes
Are sufficient details of the methods provided to allow replication by others?
Partly
Are the datasets clearly presented in a useable and accessible format?
Partly
Reviewer Expertise:
preclinical studies on animal for anticataract drugs
I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above.
We sincerely thank the reviewer for the careful evaluation and valuable feedback on our study protocol. We appreciate these constructive suggestions, which have helped us improve the clarity and methodological rigor of our work.
We agree with the reviewer and will specify the significance level for statistical analysis as P < 0.05. This will be added in the section “Statistical Analysis.”
We appreciate this valuable suggestion. We will provide detailed information and appropriate citations for the methods used to estimate ascorbic acid (AA), malondialdehyde (MDA), and total antioxidant capacity (TCA). These details will be included in the revised version under “Laboratory Analysis.”
We thank the reviewer for this valuable comment. In fact, oxidative stress markers were also measured in the aqueous humour samples. Sampling was performed twice, pre- and post-phacoemulsification. The first sample (0.2 mL of aqueous humour) was collected immediately after corneal incision using a 1 cc syringe (Terumo) with a 30 G cannula. The second sample (0.2 mL) was obtained after intraocular lens implantation and viscoelastic removal, approximately 3 minutes later, using the same technique. The pre-phacoemulsification aqueous humour samples were analysed for ascorbic acid (AA), total antioxidant capacity (TAC), and malondialdehyde (MDA), while the post-phacoemulsification samples were analysed for MDA levels only. These details will be clarified and incorporated in the
We sincerely thank the reviewer for the valuable and constructive feedback, which has strengthened the methodological transparency and scientific rationale of our research. We will prepare a revised version to address these points in detail.