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SAFETY ASSESSMENT OF LUTEIN AND ZEAXANTHIN

SUPPLEMENTATION AND ITS EFFECTS ON BLOOD GLUCOSE
LEVELS, KIDNEY FUNCTIONS, LIVER FUNCTIONS AND BONE
HEALTH - A RANDOMIZED, DOUBLE-BLIND, PLACEBO-
CONTROLLED CLINICAL STUDY

C A ANZAR*1, JOSEPH M V1, DR. SUNDARAM R1, VADIRAJ G B1, PRASAD C P 1, BINEESH
ERANIMOSE1, DR. SHOBHITH JAGADEESH2
1. Olive Lifesciences Pvt Ltd No.165/5, Near NH-4, Nelamangala, Bangalore – 562123, Karnataka,
India
2. Clinical Research Mallikatta Neuro Centre, Mallikatte, Kadri, Mangalore, Karnataka 575002, India.

ABSTRACT
Objective: HbA1c, a major marker for the sugar levels in the blood, is the litmus test for
people who are on the verge of entering the diabetic zone and for those who are already
affected by this disease. Oral hypoglycemic agents are the fine line of treatment in such
cases. Nutraceutical and herbal supplements can be utilized as a prophylactic to keep such
diseases at bay. Lutein, a carotenoid from the marigold flower, is a very well-known
ingredient in the management of eye health. Lutein and zeaxanthin, put together, are
commonly known as macular pigments. These pigments help in filtering the blue light, thus
protecting the eyes from the harmful effects of the blue light emitted from the screens of
electronic gadgets. However, recent studies have demonstrated that these macular pigments
have a significant effect on improving cognition and overall brain health. A randomized,
double-blind, placebo-controlled clinical study was conducted on lutein and zeaxanthin to
determine their effect on cognitive performance. As a safety parameter, HbA1c was also
recorded during the study. At the end of the study, the statistics on the data revealed that
lutein and zeaxanthin have a positive impact on HbA1c levels. It was observed that the
HbA1c of the subjects in the treatment group was significantly lower than that of those in the
placebo group, and the values significantly improved during the treatment duration between
weeks 1 and 5. As a result, the current study examines how lutein and zeaxanthin affect type
2 diabetes mellitus (T2DM), diabetic kidney disease (DKD), non-alcoholic fatty liver disease
(NAFLD), and bone health in healthy individuals between the ages of 35 and 75. Methods:
The blood parameters that were measured in thirty individuals who were randomly divided
into two groups are the basis for the present study. The trial consisted of two parallel
treatment groups and was randomized, double-blind,
placebo-controlled clinical research. Through advertising, healthy participants between the
NOTE: This preprint reports new research that has not been certified by peer review and should not be used to guide clinical practice.
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ages of 35 and 75 were identified in the community. Following screening, 30 participants

were accepted into the trial and randomly assigned using a computer-based randomization
methodology into the two model groups, G1 (Group 1-treated) and G2 (Group 2-placebo).
The HbA1c level for type 2 diabetes was divided into three groups based on numerical values
at various levels: "improved," "no change," or "unfavourable." For instance, if the levels of
HbA1c decreased, they were categorized as "improved," while if they increased, they were
categorized as "unfavorable." The safety profile of the supplement and any potential negative
effects on the kidneys are evaluated using renal function testing. The tests help determine
whether the supplement is altering kidney function markers such as creatinine, blood urea
nitrogen, and serum urea. These factors can show whether the supplement damages the
kidneys or affects their ability to function properly. After evaluating the levels of albumin,
globulin, bilirubin, SGPT, SGOT, ALP, and GGTP, among various liver function markers,
the effects of lutein supplementation on liver function were determined. According to a study
by Takeda et al., taking lutein for four weeks significantly increased the amount of cortical
bone in the femur and the overall amount of femoral bone mass. This improvement was
quantified by dual X-ray absorptiometry and microcomputed tomography (CT) assessments
of bone mineral density. Results: A measure of blood sugar levels called estimated average
glucose level (eAG), which is generated from HbA1c, showed similar patterns. At visit 1, the
treatment group (group 1, Lutein and Zeaxanthin) reported an average eAG of 135.54,
whereas the placebo group (group 2, Placebo) exhibited an eAG of 119.98. Before receiving
treatment, group-1's mean eAG was naturally higher than that of group-2. The mean BUN
values at visit 1 for the treatment group (group 1, Lutein and Zeaxanthin) were 10.83, while
they were 10.13 for the control group (group 2, Placebo). Groups 1 and 2 showed BUN levels
of 11.03 and 10.7, respectively, during visit 5. The mean serum urea levels for groups 1 and 2
were 23.2 and 21.69, respectively. The mean values for groups 1 and 2 at visit 5 appeared to
be 23.62 and 22.91, respectively, after 5 weeks. When creatinine levels were evaluated at
visit 5, they were practically identical to those at visit 1 (1.02 and 0.99 mg/dL for groups 1
and 2, respectively). The mean values for creatinine during visit 1 were 1.03 and 0.985
mg/dL, respectively. At visit 1, the average albumin levels for groups 1 and 2 were 4.32 and
4.61, respectively. The albumin levels in G1 and G2 were
4.62 and 4.77 mg/dL at visit 5, after receiving therapy for 5 weeks. G1 and G2 exhibited total
bilirubin levels of 0.609 and 0.547 mg/dL, respectively, during visit 1. For G1 and G2, the
respective readings on the visit-5 were 0.633 and 0.662. During visit 1, the total bilirubin
levels for G1 and G2 were 0.183 and 0.176 mg/dL, respectively. For G1 and G2, the
respective readings during Visit-5 were 0.217 and 0.219 mg/dL. Throughout this clinical trial,
there were
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no severe adverse effects. Conclusion: Clinical investigations have shown that the Lutein and
Zeaxanthin is safe for bone, kidney, liver, and diabetes health. It was also noted that the
Lutein supplementation helped in managing the HbA1c levels. Thus this study helps in
establishing the positive effects of Lutein supplementation in people with impaired blood
glucose levels. Key words: Lutein and Zeaxanthin, Type-2 diabetes, Kidney function,
Liver health, Bone health

INTRODUCTION
Carotenoids are pigments that are generated from plants and have a number of functions in
human biology (1). The xanthophyll carotenoids (XC), lutein (L), and zeaxanthin (Z) are
primarily deposited in the human macula. The carotenoid family of nutrients includes lutein
and zeaxanthin, which are fat-soluble nutrients. Marigold flowers, egg yolks, corn, and leafy
green vegetables with dark green leaves like kale and spinach contain lutein (2). Zeaxanthin
can be found in higher concentrations in foods that are yellow or orange, including egg yolks,
corn, orange capsicums, tangerines, persimmons, mandarins, and oranges (3, 4). Lutein and
zeaxanthin are pigments that are found in the brain, breast, fat tissue, and eyes. Lutein is
deposited maximum in our brains, despite the fact that it is not the most prevalent carotenoid
in our diet (5). In fact, 66 to 77% of the total carotenoid content in human brain tissue is made
up of lutein and zeaxanthin (6). There is growing interest in the neuroprotective benefits of
lutein and zeaxanthin since they have been found in the hippocampus, cerebellum, frontal,
occipital, and temporal cortices (7-9), and they also have potent antioxidant and anti-
inflammatory capacities (10, 11).
One of the most common chronic diseases, diabetes mellitus, is regarded as a major issue in
the healthcare system. Diabetes is going to be among the primary causes of morbidity and
mortality due to the rising incidence of diabetes mellitus (12). It would appear that a greater
emphasis on this issue is essential given that diabetes is more common in women and that it
also causes metabolic changes that may result in early menopause and increase hormonal
symptoms in women, affecting their health and quality of life (13–15). In order to attain the
goals of treatment and manage diabetic complications, these patients should take
responsibility for cardiovascular risk factors such as hypertension and dyslipidemia and
manage them at the same time as controlling blood sugar (16).
The primary aspect of treatment for DKD (diabetic kidney disease) includes blood pressure
regulation, renin-angiotensin system (RAS) inhibitors, and glycemic control. These methods
have been shown to successfully lower the risk of disease obtain or progress (17, 18).
However,
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individuals with DKD (diabetic kidney disease) who are intolerant to or insensitive to current
pharmacotherapies as well as those with declining renal function but normal-albuminuria
have unmet needs (19–21).
Non-alcoholic fatty liver disease (NAFLD) is a spectrum of hepatic illnesses characterized by
steatosis without competing etiologies, including alcohol usage, chronic drug use, or viral
infections (25). NAFLD is a serious public health concern because it affects around a quarter
of the world's population (26). Given its close connection to insulin resistance and abnormal
adipose tissue, NAFLD prevalence has increased along with obesity and diabetes (27). The
progressive type of NAFLD, known as non-alcoholic steatohepatitis (NASH), can eventually
result in cirrhosis and hepatocellular cancer. As there is currently little evidence to support
medication, such as vitamin E and thiazolidinedione, the mainstays of NAFLD treatment are
weight loss and physical activity (28). There is not a single drug that has been authorized for
the treatment of NAFLD at the moment (30).
Carotenoids, which include lycopene, beta-carotene, lutein, zeaxanthin, and beta-
kryptoxanthin, have generated a lot of interest in the field of human nutrition because they
function as biological antioxidants and help protect the body from reactive oxygen species
(ROS) (31, 32). They also play a protective role in conditions like diabetes and
cardiovascular disease (CVD), affecting cellular signalling pathways and influencing the
expression of specific genes. Dietary carotenoids are primarily accumulated in the liver, from
which they are transported by various lipoproteins for their release into the bloodstream,
where they are then deposited and stored in various organs and tissues, including the kidneys,
adipose tissue, adrenal glands, testes, skin, and prostate (35).
Lutein is a naturally occurring oxygenated carotenoid with clear anti-inflammatory and
antioxidative properties (36). Additionally, it is widely acknowledged that oxidative stress
and the inflammatory response are strongly associated with T2DM and DKD (37). However,
it is yet unknown whether lutein, T2DM, and DKD in older people are correlated. Although
potential therapy targets are obvious, the practitioner is confronted with a bewildering array
of research and single drug prescriptions that are difficult to understand, communicate to
patients, or incorporate into standard diabetic and DR treatments. Among the 30–40
carotenoid metabolites found in human blood, β-carotene, lycopene, lutein, β-kryptoxanthin,
and zeaxanthin share the majority of xanthophylls (such as lutein, zeaxanthin, and
astaxanthin), which contain one or more oxygen atoms (38). Research has been done
extensively on the chemical characteristics of carotenoids, particularly their strong
antioxidant potential. There has recently been an increase in interest in their biological
activities in relation to their possible
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function in the prevention of certain chronic diseases including diabetes and its consequences
(39-41).
Carotenoids are strong antioxidants that can eliminate ROS and improve a cell's capacity to
avoid oxidative stress, which is thought to be an emerging treatment approach for people with
chronic kidney disease (CKD) (42). It can be divided into two main categories: xanthophylls
and carotenes. Xanthophylls, which include lutein, zeaxanthin, and β-kryptoxanthin, are
oxygenated terpenes, whereas carotenes, which include β-carotene, α-carotene, and lycopene,
as well as other less-studied species, are unoxygenated terpenes (43). The ability of each of
these carotenoids to scavenge radicals in three steps—electron transfer, hydrogen abstraction,
and addition—has been demonstrated, and they all serve as the major scavengers of ROS
(44). A nephroprotective effect of β-carotene has been demonstrated in animal investigations
using rats given bromobenzene. There hasn't been any research done yet to determine
whether CKD patient mortality is correlated with carotenoid intake. Therefore, the purpose of
this study is to determine whether CKD patient mortality risk can be reduced by carotenoid
(lutein and zeaxanthin) intake.
These antioxidants (lutein and zeaxanthin) and their metabolites can build up in the liver and
have a favourable impact on hepatocyte metabolism by controlling the cellular oxidative state
in certain liver diseases. A major and expanding clinical issue in both industrialized and
developing nations, non-alcoholic fatty liver disease (NAFLD) is currently regarded as one of
the most prevalent chronic liver disorders in the world.
Lutein (L), a dietary carotenoid that inhibits bone resorption, stimulates bone formation, and
increases bone density—most prominently in cortical bone (44). If lutein and zeaxanthin
really offer protection against bone loss, as the results from the experimental animal data on
young mice suggest, then it would be beneficial to understand the association between lutein
and zeaxanthin status and bone health prior to the onset of the degeneration that occurs
frequently in aging samples.
There have been an increasing number of clinical trials, preclinical trials, and systematic
reviews in the past few years for lutein and zeaxanthin in the treatment of diabetes mellitus
(DM), chronic kidney disease (CKD), Non-Alcoholic fatty liver disease (NAFLD), and bone
health, but no placebo-controlled trials have been identified anywhere. A randomised placebo
controlled clinical study was conducted to evaluate the efficacy of Marigold extract (Lutein
and Zeaxanthin on cognitive performance and moods in adults with stroke and parkinsonism.
As part of the safety evaluation all the participants in this clinical trial were subjected to
undergo regular monitoring of their HbA1c, Liver function and Kidney function tests during
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their tenure of the clinical trial. The ethical clearance was obtained at Mangala Institutional
Ethics Committee vide letter Ref. No. MIEC/V6.1/015 and the trial was registered at CTRI
vide registration Number CTRI/2022/06/043208. One of the objectives of this study was to
study the safety of Lutein and zeaxanthin supplementation on the blood sugar levels through
HbA1c, Liver and Kidney function test
MATERIAL AND METHODS
Trial design
The current study was based on blood parameters taken from 30 people who were randomly
divided into two groups. The Lutein and Zeaxanthin lutein supplement was placed in group 1,
and the placebo were in group 2. The trial consists of two parallel treatment groups in a
randomized, double-blind, placebo-controlled clinical research. Through advertising, healthy
people in the community between the ages of 35 and 75 were recruited. A computer-based
randomization methodology was used to block-randomize 30 participants into the study's two
treatment groups, G1 and G2, after screening. Age and gender distribution throughout the
study arms was equal. Participants were required to meet a number of inclusion criteria,
including the capacity to give informed consent, a diagnosis of the required disease, severity,
or symptom, and a willingness to adhere by all study guidelines. People had to meet
exclusion criteria such as significant renal or hepatic impairment, elective surgery scheduled
during the trial or other procedures requiring general anesthesia, or any significant disease or
disorder that might danger participants or affect the outcome of the trial. Pregnant, lactating,
or intending to become pregnant female trial participants were also excluded. The patient
characteristics were represented in Table 1.
Table 1: Demographic and Clinical Characteristics of Participants at Study Enrolment
G1 (N=16) G2 (N=14) Overall (N=30)
Age
Mean (SD) 57.5 (9.01) 53.7 (9.37) 55.7 (9.22)
Median (Min, Max) 59.5 (38.0, 71.0) 55.5 (38.0, 65.0) 58.0 (38.0, 71.0)
Diagnosis
PD 8 (50.0%) 6 (42.9%) 14 (46.7%)
PD & Stroke 1 (6.3%) 0 (0%) 1 (3.3%)
Stroke 7 (43.8%) 8 (57.1%) 15 (50.0%)
Gender
Female 5 (31.3%) 4 (28.6%) 9 (30.0%)
Male 11 (68.8%) 10 (71.4%) 21 (70.0%)
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Intervention
During the intervention period, participants in the treatment group (G1) received two capsules
per day that included 75 mg of lutein and zeaxanthin encapsulated in MCT, standardized to
lutein at 10 mg and zeaxanthin at 1 mg. In the placebo group (G2), participants administered
capsules that were equivalent to those given to the therapy group. For the course of the study,
the capsules were delivered in sealed bottles containing 60 capsules each. The labels on the
bottles included an extensive list of details, such as the manufacture date, expiration date,
serial number assigned to the trial dose, an explicit declaration that it was only intended for
clinical trial purposes and not for commercial use, and a beneficial warning that it was an
investigational product. The label also advised against taking a higher dosage, suggested
keeping it away from minors and storing it in a cold, dry location, suggested taking the drug
with water to prevent swallowing issues, and advised exercising caution if someone suffers
any discomfort. The blood parameters were taken at visits 1 and 5, and the tables below
illustrate their particulars.

Table 2: Baseline Clinical Parameters of Study Participants (G1, treatment group) and group-
2 (G2, placebo group) during visit-1
G1 (N=16) G2 (N=14) Overall (N=30)
Age
Mean (SD) 57.5 (9.01) 53.7 (9.37) 55.7 (9.22)
Median (Min, Max) 59.5 (38.0, 71.0) 55.5 (38.0, 65.0) 58.0 (38.0, 71.0)
Gender
Female 5 (31.3%) 4 (28.6%) 9 (30.0%)
Male 11 (68.8%) 10 (71.4%) 21 (70.0%)
Bun
Mean (SD) 10.8 (2.88) 10.1 (1.74) 10.5 (2.38)
Median (Min, Max) 10.3 (7.40, 18.9) 9.95 (8.00, 14.5) 10.1 (7.40, 18.9)
Missing 1 (6.3%) 0 (0%) 1 (3.3%)
Urea Serum
Mean (SD) 23.2 (6.18) 21.7 (3.72) 22.5 (5.11)
Median (Min, Max) 22.0 (15.8, 40.5) 21.3 (17.1, 31.0) 21.6 (15.8, 40.5)
Missing 1 (6.3%) 0 (0%) 1 (3.3%)
Glucose Random
Mean (SD) 103 (15.6) 125 (40.6) 114 (31.9)
Median (Min, Max) 107 (77.0, 132) 117 (70.0, 228) 109 (70.0, 228)
Missing 1 (6.3%) 0 (0%) 1 (3.3%)
Creatinine
Mean (SD) 1.03 (0.256) 0.985 (0.205) 1.01 (0.230)
Median (Min, Max) 1.11 (0.720, 1.47) 0.945 (0.720,1.35) 1.07 (0.720, 1.47)
Missing 1 (6.3%) 0 (0%) 1 (3.3%)
HbA1c
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Mean (SD) 6.35 (0.772) 5.81 (0.459) 6.11 (0.696)

Median (Min, Max) 6.30 (5.50, 8.60) 5.60 (5.20, 6.80) 6.00 (5.20, 8.60)
Missing 0 (0%) 1 (7.1%) 1 (3.3%)
eAG
Mean (SD) 136 (22.2) 120 (13.2) 129 (20.0)
Median (Min, Max) 134 (111, 200) 114 (103, 148) 126 (103, 200)
Missing 0 (0%) 1 (7.1%) 1 (3.3%)

Table 3: Clinical parameters in group-1 (G1, treatment group) and group-2 (G2, placebo
group) during visit-5
G1 (N=16) G2 (N=14) Overall (N=30)
Age
Mean (SD) 57.5 (9.01) 53.7 (9.37) 55.7 (9.22)
Median (Min, Max) 59.5 (38.0, 71.0) 55.5 (38.0, 65.0) 58.0 (38.0, 71.0)
Gender
Female 5 (31.3%) 4 (28.6%) 9 (30.0%)
Male 11 (68.8%) 10 (71.4%) 21 (70.0%)
Bun
Mean (SD) 11.0 (2.69) 10.7 (1.69) 10.9 (2.25)
Median (Min, Max) 10.4 (7.60, 16.6) 10.8 (7.80, 13.5) 10.4 (7.60, 16.6)
Urea Serum
Mean (SD) 23.6 (5.76) 22.9 (3.62) 23.3 (4.81)
Median (Min, Max) 22.1 (16.3, 35.5) 23.0 (16.7, 28.9) 22.1 (16.3, 35.5)
Glucose Random
Mean (SD) 101 (31.6) 118 (47.6) 109 (40.1)
Median (Min, Max) 94.0 (60.0, 177) 103 (85.0, 237) 97.5 (60.0, 237)
Creatinine
Mean (SD) 1.02 (0.236) 0.989 (0.265) 1.01 (0.246)
Median (Min, Max) 1.03 (0.650, 1.41) 0.935 (0.680, 1.40) 1.01 (0.650, 1.41)
HbA1c
Mean (SD) 5.94 (0.733) 5.80 (0.419) 5.88 (0.602)
Median (Min, Max) 5.70 (5.40, 8.20) 5.70 (5.40, 6.90) 5.70 (5.40, 8.20)
eAG
Mean (SD) 124 (21.0) 120 (12.0) 122 (17.2)
Median (Min, Max) 117 (108, 189) 117 (108, 151) 117 (108, 189)

1. Lutein and Zeaxanthin Supplementation on Blood Glucose Levels

Low lutein levels were significantly correlated with high glycated hemoglobin (HbA1c)
levels, according to a population-based cross-sectional investigation (45). In a different cross-
sectional study, 1597 Australian adults were used, and the results revealed an inverse
relationship between serum lutein levels and type 2 diabetes (T2DM), as well as a decrease in
fasting blood glucose (FBG), fasting insulin, and post-load plasma glucose levels with an
increase in lutein quintiles (46). In addition, a recent meta-analysis by Jiang et al. (47)
showed
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that high food intake and circulating lutein levels are linked to a decreased risk of T2DM.
Additionally, eight weeks of lutein administration significantly decreased FBG levels in a
streptozotocin-induced rat model of T2DM, improved glucose tolerance, and raised the
antioxidant enzyme activities in the blood, heart, and kidneys (48). Serum lutein levels have
even been suggested as a possible T2DM measure in a recent study (49). The HbA1c level for
type 2 diabetes was divided into three groups based on numerical values at various levels:
"improved," "no change," or "unfavourable." For instance, if the levels of HbA1c decreased,
they were categorized as "improved," while if they increased, they were categorized as
"unfavorable." If there was no change in HbA1c levels, it was categorized as "no change."
This categorization of the clinical parameter allowed for the use of statistical tests designed
for analysing categorical data, such as Fisher's exact test, to investigate the relationship
between the clinical parameter and treatment. This study also measured random blood sugar
levels during the visits.
2. Lutein and Zeaxanthin Supplementation On Kidney Function
Lutein could enhance kidney function, according to several studies. In accordance with an
investigation published in 2014 (50), the antioxidant and anti-inflammatory properties of
lutein have the potential to protect the kidneys from reperfusion from ischemia injury-
induced kidney damage. The use of lutein appears to restore kidney function in rats exposed
to IRI and considerably lower levels of kidney damage markers such as serum creatinine and
blood urea nitrogen. In a recent study (51), serum lutein levels were shown to be substantially
reduced in elderly people with type 2 diabetes mellitus (T2DM) and diabetic kidney disease
(DKD) compared to healthy controls. Lutein concentrations were found to be a diagnostic
sign for T2DM and DKD since they exhibited a negative correlation with a number of
metabolic markers. These findings indicate that lutein may act as a preventative measure
against the development of T2DM and DKD and may serve as a biomarker for the diagnosis
of these conditions. These studies attribute lutein's anti-inflammatory and antioxidant
characteristics to its positive impact on the kidney. Due to the fact that oxidative stress plays
a significant role in kidney damage, lutein's capacity to decrease oxidative stress is suggested
as the mechanism underlying its positive benefits. Several kidney function markers, including
BUN, serum urea, and creatinine, were assessed in the current study's two groups at visits one
and five, which occurred before and after the various therapies, respectively.
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3. Lutein and Zeaxanthin Supplementation On Liver Function

One of the possible side effects of pharmacological therapies was drug-induced liver damage
(DILI). We evaluated liver function in order to carefully assess the effect of our intervention
on participant safety. Blood tests known as liver function tests (LFTs) are performed to assess
the health and function of the liver. Clinical trials need LFTs because liver dysfunction or
damage can be a significant adverse event linked to numerous drugs and treatments.
Monitoring LFTs during a clinical trial can aid in the early detection of potential liver
toxicity, enabling timely intervention and reducing the risk to research participants. In our
clinical trials, the effects of lutein supplementation on liver function are discussed in the
sections that follow. During the clinical trial, a number of tests were measured, including
albumin, globulin, bilirubin, SGPT, SGOT, ALP, and GGTP levels.
4. Lutein and Zeaxanthin Supplementation On bone health
According to a study by Takeda et al., four weeks of lutein treatment significantly increased
the mass of the femoral bones, particularly the cortical bone. This improvement was
quantified by dual X-ray absorptiometry and microcomputed tomography (CT) evaluations of
bone mineral density (52). Male hip fracture risk has been demonstrated to be negatively
linked with dietary intake of the carotenoids α-carotene, β-carotene, and lutein (53). Higher
serum carotenoid levels were linked to better bone mineral density in both men and women,
according to research by Zhang et al. (54), who observed similar outcomes.
Statistical analysis
The paired t test in R programming was used for the statistical comparison of the baseline
characteristics and outcomes between the 2 groups in visits 1 and 2. Descriptive data analysis
was carried out using descriptive statistical methods for all analyses. The complete results of
the statistical study have been provided in several tables. A P value less than 0.05 was used to
determine whether the results were statistically significant in each case.
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RESULTS AND DISCUSSION

RESULTS
1 Effect of Lutein and Zeaxanthin Supplementation on Blood Glucose Levels
In the current study, the values of HbA1c were measured in each group during visits 1 and 5.
The average HbA1c at visit 1 for the treated group (group 1, Lutein and Zeaxanthin) was
6.35, whereas for the placebo group (group 2, placebo), it was 5.8. The mean HbA1c tended
to be on the higher side of group-1 compared to group-2 prior to treatment. Five weeks of
treatment with lutein supplements resulted in a statistically significant reduction in HbA1c
levels. In group 1, the mean HbA1c levels were reduced to 5.9 as measured at visit 5.
However, in group 2 (placebo), the mean HbA1c levels were unchanged. The data are shown
in Tables 4 and 5 and Figure 1. At the conclusion of five weeks, the outcomes were
categorized as "improved," "no change," or "unfavorable." The patients in G-1, 68.8%
exhibited improvement in their HbA1c levels (i.e., their HbA1c levels dropped as a result of
the treatment), compared to only 35.7% of those in G2. Unfavorable changes in HbA1c were
seen in 57% of G-2 participants. Only 18.8% had an unfavorable change in G-1. This data
was subjected to statistical analysis using Fisher’s exact test, and the effect was significant
since the p value was found to be 0.048. The data was determined in Table 6.

Table 4: HbA1c values (mg/dL) during Visit-1 in G1 (treatment group) and G2 (placebo group)
HbA1c G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 6.35 (0.772) 5.81 (0.459) 6.11 (0.696)
Median (Min, Max) 6.30 (5.50, 8.60) 5.60 (5.20,6.80) 6.00 (5.20, 8.60)
Missing 0 (0%) 1 (7.1%) 1 (3.3%)

Table 5: HbA1c values (mg/dL) during Visit-5 in G1 (treatment group) and G2 (placebo
group)
HbA1c G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 5.94 (0.733) 5.80 (0.419) 5.88 (0.602)
Median (Min, Max) 5.70 (5.40, 8.20) 5.70 (5.40,6.90) 5.70 (5.40, 8.20)
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Figure1: Effect of HbA1c values (mg/dL) during Visit-1 and Visit-5 in G1 (treatment group)
and G2 (placebo group)

G1V1 (HbA1c levels measured in group-1 at visit-1); G1V5 (HbA1c levels measured in
group- 1 at visit-5); G2V1 (HbA1c levels measured in group-2 at visit-1); G2V5 (HbA1c
levels measured in group-2 at visit-5). The central line in the box plots represents the median
values, and the rectangular box shows the interquartile range (IQR), which is the middle 50%
of the data. The whiskers extend to the minimum and maximum values of the data, except for
the outliers that fall beyond the whiskers. Statistical significance was calculated using a
paired, two-sided t-test. The * value represents p<0.05 for comparison between HbA1c levels
between G1V1 and G1V5.

Table 6: Effect of Lutein and Zeaxanthin Supplementation on HbA1c after 5 weeks

HbA1c Outcome G1 (N=16) G2 (N=14) Overall (N=30)
Improved 11 (68.8%) 5 (35.7%) 16 (53.3%)
No change 2 (12.5%) 0 (0%) 2 (6.7%)
Unfavorable 3 (18.8%) 8 (57.1%) 11 (36.7%)
Missing 0 (0%) 1 (7.1%) 1 (3.3%)
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1.1 Effect of estimated Average Glucose (eAG) Levels:

Estimated average glucose level (eAG) is a parameter derived from HbA1c; similar trends
were observed. The average eAG at visit 1 for the treated group (group 1, Lutein and
Zeaxanthin) was 135.54, whereas for the placebo group (group 2, Placebo), it was 119.98.
The mean eAG was, of course, on the higher side of group 1 compared to group 2 prior to
treatment. Five weeks of treatment with a lutein supplement resulted in a statistically
significant reduction in the eAG levels. In group 1, the mean eAG levels decreased to 123.92
as measured at visit 5. However, in group 2 (placebo), the mean eAG levels were unchanged.
This is obviously an expected outcome since eAG is a parameter derived from HbA1c.
Tables 7 and 8 and the figure below represent the details.
Table 7: eAG values (mg/dL) during Visit-1 in G1 (treatment group) and G2 (placebo group)
eAG G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 136 (22.2) 120 (13.2) 129 (20.0)
Median (Min, Max) 134 (111, 200) 114 (103, 148) 126 (103, 200)
Missing 0 (0%) 1 (7.1%) 1 (3.3%)

Table 8: eAG values (mg/dL) during Visit-5 in G1 (treatment group) and G2 (placebo group)
eAG G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 124 (21.0) 120 (12.0) 122 (17.2)
Median (Min, Max) 117 (108, 189) 117 (108, 151) 117 (108, 189)

Figure2: Effect of Lutein and Zeaxanthin Supplementation on Estimated Average Glucose

(eAG) levels

G1V1 (eAG levels measured in group-1 at visit-1); G1V5 (eAG levels measured in group- 1
at visit-5); G2V1 (eAG levels measured in group-2 at visit-1); G2V5 (eAG levels measured
in
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group-2 at visit-5). The central line in the box plots represent the median values and the
rectangular box shows the interquartile range (IQR), which is the middle 50% of the data.
The whiskers extend to the minimum and maximum values of the data, except for the outliers
that fall beyond the whiskers. Statistical significance was calculated using paired, two sided
t-test.
* value represents p<0.05 for comparison between HB1AC levels between G1V1 and G1V5
1.2 Effect of Lutein and Zeaxanthin supplementation on Random glucose levels
There were no statistically significant changes for either of the groups between visits 1 and 5.
However, it should be noted that random blood sugar levels can only provide a quick and
easy snapshot of a person's blood sugar level at a specific moment in time. However, they are
not a reliable method for tracking diabetes or diabetes control over time. This is because
random blood sugar levels can be influenced by various factors, such as recent food intake,
physical activity, and stress levels. Hence, the results of random blood sugar levels are not
generally taken into consideration for validating the effects of any intervention. Tables 9 and
10 and Figure 3 show the proper results.

Table 9: Random glucose values (mg/dL) during Visit-1 in G1 (treatment group) and G2
(placebo group)

Random glucose G1 (N=16) G2 (N=14) Overall (N=30)

Mean (SD) 103 (15.6) 125 (40.6) 114 (31.9)
Median (Min, Max) 107 (77.0, 132) 117 (70, 228) 109 (70, 228)
Missing 1 (6.3%) 0 (0%) 1 (3.3%)

Table 10: Random glucose values (mg/dL) during Visit-5 in G1 (treatment group) and G2
(placebo group)

eAG G1 (N=16) G2 (N=14) Overall (N=30)

Mean (SD) 101 (31.6) 118 (47.6) 109 (40.1)
Median (Min, Max) 94.0 (60, 177) 103 (85.0, 237) 97.5 (60.0, 237)
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Figure 3: Effect of Lutein and Zeaxanthin supplementation on Random glucose levels

G1V1 (random blood glucose levels measured in group-1 at visit-1); G1V5 (random blood
glucose levels measured in group-1 at visit-5); G2V1 (random blood glucose levels measured
in group-2 at visit-1); G2V5 (random blood glucose levels measured in group-2 at visit-5).
The central line in the box plots represents the median values, and the rectangular box shows
the interquartile range (IQR), which is the middle 50% of the data. The whiskers extend to
the minimum and maximum values of the data, except for the outliers that fall beyond the
whiskers. Statistical significance was calculated using a paired, two-sided t-test. No
significance was observed for corresponding comparisons.
2. Effect of Lutein and Zeaxanthin supplementation On Kidney Function
2.1 Blood Urea Nitrogen (BUN)
The mean BUN values at visit 1 for group 1 (Lutein and Zeaxanthin) were 10.83, whereas for
placebo group 2 (placebo), they were 10.13. During visit 5, the BUN values for group 1 and
group 2 were 11.03 and 10.7, as expressed in Tables 11 and 12 and Figure 4, respectively.
These changes were not statistically significant as measured by a paired, two-tailed t-test.
Blood urea nitrogen (BUN) is a common laboratory test that measures the amount of urea
nitrogen in the blood.
Table 11: BUN values (mg/dL) during Visit-1 in G1 (treatment group) and G2 (placebo group)
BUN G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 10.8 (2.88) 10.1 (1.74) 10.5 (3.58)
Median (Min, Max) 10.3 (7.40, 18.9) 9.95 (8.00, 14.5) 10.1 (7.40, 18.9)
Missing 1 (6.3%) 0 (0%) 1 (3.3%)
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Table 12: BUN values (mg/dL) during Visit-5 in G1 (treatment group) and G2 (placebo group)

BUN G1 (N=16) G2 (N=14) Overall (N=30)

Mean (SD) 11.0 (2.69) 10.7 (1.69) 10.9 (2.25)
Median (Min, Max) 90.4 (7.60, 16.6) 10.8 (7.80, 13.5) 10.4 (7.60, 16.6)

Figure 4: Effect of Lutein and Zeaxanthin supplementation on Blood Urea Nitrogen (BUN)
levels

G1V1 (BUN levels measured in group-1 at visit-1); G1V5 (BUN levels measured in group-1
at visit-5); G2V1 (BUN levels measured in group-2 at visit-1); G2V5 (BUN levels measured
in group-2 at visit-5). The central line in the box plots represents the median values, and the
rectangular box shows the interquartile range (IQR), which is the middle 50% of the data.
The whiskers extend to the minimum and maximum values of the data, except for the outliers
that fall beyond the whiskers. Statistical significance was calculated using a paired, two-sided
t- test. No significance was observed for corresponding comparisons.
2.2 Serum Urea levels
The serum urea level is a commonly used laboratory test to assess kidney function and
monitor kidney diseases. In the current study, during visit 1, the mean serum urea levels for
group 1 and group 2 were 23.2 and 21.69, respectively. At visit 5, after 5 weeks, the mean
values for groups 1 and 2 were 23.62 and 22.91, respectively. No significant changes in
serum urea levels were observed in either group between the two visits (Tables 13 and 14 and
Figure 5).
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Table 13: Serum Urea (mg/dL) during Visit-1 in G1 (treatment group) and G2 (placebo group)
Urea Serum G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 23.2 (6.18) 21.7 (3.72) 22.5 (5.11)
Median (Min, Max) 22.0 (15.8, 40.5) 21.3 (17.1, 31.0) 21.6 (15.8, 40.5)
Missing 1 (6.3%) 0 (0%) 1 (3.3%)

Table 14: Serum Urea (mg/dL) during Visit-5 in G1 (treatment group) and G2 (placebo group)
Urea Serum G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 23.6 (5.76) 22.9 (3.62) 23.3 (4.81)
Median (Min, Max) 22.1 (16.3, 35.5) 23.0 (16.7, 28.90) 22.1 (16.3, 35.5)

Figure 5: Effect of Lutein and zeaxanthin supplementation on Serum Urea levels

G1V1 (serum urea levels measured in group 1 at visit 1); G1V5 (serum urea levels measured
in group 1 at visit 5); G2V1 (serum urea levels measured in group 2 at visit 1); G2V5 (serum
urea levels measured in group 2 at visit 5). The central line in the box plots represents the
median values, and the rectangular box shows the interquartile range (IQR), which is the
middle 50% of the data. The whiskers extend to the minimum and maximum values of the
data, except for the outliers that fall beyond the whiskers. Statistical significance was
calculated using a paired, two-sided t-test. No significance was observed for corresponding
comparisons.
2.3 Serum creatinine levels
The serum creatinine levels did not show any significant variation in either group during visit
5 in comparison to visit 1, according to the findings of this study. As mentioned in the table
below, mean values for creatinine during visit 1 were 1.03 and 0.985 mg/dL, respectively,
and the levels remained almost the same when measured at visit 5 (1.02 and 0.99 mg/dL,
respectively, for group 1 and group 2) (Tables 15 and 16; Figure 6).
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Table 15: Creatinine levels (mg/dL) during Visit-1 in G1 (treatment group) and G2 (placebo
group)

Creatinine G1 (N=16) G2 (N=14) Overall (N=30)

Mean (SD) 1.03 (0.256) 0.985 (0.205) 1.01 (0.230)
Median (Min, Max) 1.11 (0.720, 1.47) 0.945 (0.720, 1.07 (0.720,
1.35) 1.47)
Missing 1 (6.3%) 0 (0%) 1 (3.3%)

Table 16: Creatinine levels (mg/dL) during Visit-5 in G1 (treatment group) and G2 (placebo
group)
Creatinine G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 1.02 (0.236) 0.989 (0.265) 1.01 (0.246)
Median (Min, Max) 1.03 (0.650, 1.41) 0.935 (0.680, 1.40) 1.01 (0.650, 1.41)

Figure 6: Effect of Lutein and zeaxanthin supplementation on Creatinine levels

G1V1 (creatinine levels measured in group-1 at visit-1); G1V5 (creatinine levels measured in
group-1 at visit-5); G2V1 (creatinine levels measured in group-2 at visit-1); G2V5 (creatinine
levels measured in group-2 at visit-5). The central line in the box plots represents the median
values, and the rectangular box shows the interquartile range (IQR), which is the middle 50%
of the data. The whiskers extend to the minimum and maximum values of the data, except for
the outliers that fall beyond the whiskers. Statistical significance was calculated using a
paired, two-sided t-test. No significance was observed for corresponding comparisons.
3. Effect of Lutein and Zeaxanthin Supplementation On Liver Function
Monitoring LFTs throughout a clinical trial can help identify potential liver toxicity early on,
allowing for prompt intervention and minimizing harm to study participants. The following
sections describe the effects of lutein supplementation on liver function in our clinical trials.
A battery of tests such as levels of albumin, globulin, bilirubin, SGPT, SGOT, ALP, and
GGTP
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were measured during the clinical trial to determine the proper function of the liver. Tables 17
and 18 provide details on the basal levels of these parameters during the study period.

Table 17: Liver function test parameters of the participants during visit-1, (G1 – treatment
group, G2- Placebo group)
G1 (N=16) G2 (N=14) Overall(N=30)

Age
Mean (SD) 57.5 (9.01) 53.7 (9.37) 55.7 (9.22)
Median [Min, Max] 59.5 [38.0, 71.0] 55.5 [38.0, 65.0] 58.0 [38.0, 71.0]
Total protein
Mean (SD) 7.24 (0.373) 7.36 (0.323) 7.30 (0.348)
Median [Min, Max] 7.09 [6.81, 7.94] 7.34 [6.64, 7.91] 7.31 [6.64, 7.94]
Missing 1(6.3%) 0 (0%) 1 (3.3%)
Albumin
Mean (SD) 4.32 (0.251) 4.61 (0.245) 4.46 (0.286)
Median [Min, Max] 4.25 [4.03, 4.91] 4.64 [4.02, 5.10] 4.45 [4.02, 5.10]
Missing 1 (6.3%) 0 (0%) 1 (3.3%)
Globulin
Mean (SD) 2.93 (0.322) 2.74 (0.281) 2.84 (0.312)
Median [Min, Max] 2.86 [2.57, 3.87] 2.72 [2.27, 3.31] 2.78 [2.27, 3.87]
Missing 1(6.3%) 0 (0%) 1 (3.3%)
Albumin-Globulin ratio
Mean (SD) 1.49 (0.174) 1.70 (0.229) 1.59 (0.225)
Median [Min, Max] 1.51 [1.05, 1.76] 1.68 [1.39, 2.25] 1.54 [1.05, 2.25]
Missing 1 (6.3%) 0 (0%) 1 (3.3%)
Total Bilirubin
Mean (SD) 0.609 (0.359) 0.547 (0.315) 0.580 (0.335)
Median [Min, Max] 0.480 [0.240, 1.59] 0.470 [0.240, 1.42] 0.470 [0.240, 1.59]
Direct Bilirubin
Mean (SD) 0.183 (0.0814) 0.176 (0.0803) 0.180 (0.0795)
Median [Min, Max] 0.160 [0.100, 0.410] 0.145 [0.0900, 0.360] 0.155 [0.0900, 0.410]
Indirect Bilirubin
Mean (SD) 0.426 (0.279) 0.371 (0.241) 0.400 (0.259)
Median [Min, Max] 0.315 [0.130, 1.18] 0.310 [0.150, 1.06] 0.310 [0.130, 1.18]
SGPT
Mean (SD) 20.4 (7.36) 31.4 (15.2) 25.5 (12.8)
Median [Min, Max] 18.0 [11.0, 38.8] 30.5 [8.70, 62.0] 23.0 [8.70, 62.0]
SGOT
Mean (SD) 21.5 (5.79) 24.3 (5.88) 22.8 (5.90)
Median [Min, Max] 21.5 [14.0, 35.0] 23.5 [16.2, 41.0] 22.0 [14.0, 41.0]
ALP
Mean (SD) 77.9 (21.1) 70.2 (16.3) 74.2 (19.0)
Median [Min, Max] 74.0 [49.0, 123] 65.0 [46.0, 108] 67.0 [46.0, 123]
Missing 1 (6.3%) 0 (0%) 1 (3.3%)
GGTP
Mean (SD) 25.9 (18.8) 33.3 (22.0) 29.5 (20.4)
Median [Min, Max] 22.0 [11.0, 90.0] 26.5 [11.0, 83.0] 25.0 [11.0, 90.0]
Missing 1 (6.3%) 0 (0%) 1 (3.3%)
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Table 18: Liver function test parameters of the participants during visit-5, (G1 – treatment
group, G2- Placebo group)
G1 (N=16) G2 (N=14) Overall(N=30)

Age
Mean (SD) 57.5 (9.01) 53.7 (9.37) 55.7 (9.22)
Median [Min, Max] 59.5 [38.0, 71.0] 55.5 [38.0, 65.0] 58.0 [38.0, 71.0]
Total protein
Mean (SD) 7.54 (0.400) 7.73 (0.343) 7.62 (0.381)
Median [Min, Max] 7.47 [6.97, 8.25] 7.59 [7.33, 8.32] 7.59 [6.97, 8.32]
Missing 0 (0%) 1 (7.1%) 1 (3.3%)
Albumin
Mean (SD) 4.62 (0.281) 4.77 (0.200) 4.69 (0.254)
Median [Min, Max] 4.56 [4.26, 5.14] 4.74 [4.45, 5.15] 4.69 [4.26, 5.15]
Missing 0 (0%) 1 (7.1%) 1 (3.3%)
Globulin
Mean (SD) 2.92 (0.401) 2.96 (0.322) 2.94 (0.362)
Median [Min, Max] 2.79 [2.21, 3.90] 3.03 [2.47, 3.48] 2.80 [2.21, 3.90]
Missing 0 (0%) 1 (7.1%) 1 (3.3%)
Albumin-Globulin
ratio
Mean (SD) 1.62 (0.260) 1.63 (0.206) 1.62 (0.233)
Median [Min, Max] 1.61 [1.12, 2.15] 1.66 [1.35, 2.01] 1.65 [1.12, 2.15]
Missing 0 (0%) 1 (7.1%) 1 (3.3%)
Total Bilirubin
Mean (SD) 0.633 (0.426) 0.662 (0.427) 0.646 (0.419)
Median [Min, Max] 0.505 [0.300, 2.04] 0.420 [0.230, 1.45] 0.500 [0.230, 2.04]
Missing 0 (0%) 1 (7.1%) 1 (3.3%)
Direct Bilirubin
Mean (SD) 0.217 (0.0993) 0.219 (0.0993) 0.218 (0.0975)
Median [Min, Max] 0.195 [0.110, 0.500] 0.180 [0.100, 0.420] 0.190 [0.100, 0.500]
Missing 0 (0%) 1 (7.1%) 1 (3.3%)
Indirect Bilirubin
Mean (SD) 0.416 (0.333) 0.443 (0.333) 0.428 (0.327)
Median [Min, Max] 0.325 [0.190, 1.54] 0.280 [0.100, 1.06] 0.310 [0.100, 1.54]
Missing 0 (0%) 1 (7.1%) 1 (3.3%)
SGPT
Mean (SD) 28.2 (14.2) 30.2 (13.7) 29.1 (13.7)
Median [Min, Max] 27.0 [14.0, 61.0] 27.0 [10.1, 55.0] 27.0 [10.1, 61.0]
Missing 0 (0%) 1 (7.1%) 1 (3.3%)
SGOT
Mean (SD) 26.1 (13.9) 27.0 (8.70) 26.5 (11.6)
Median [Min, Max] 23.3 [13.0, 72.0] 26.0 [17.4, 51.0] 25.0 [13.0, 72.0]
Missing 0 (0%) 1 (7.1%) 1 (3.3%)
ALP
Mean (SD) 87.2 (25.7) 68.4 (18.7) 78.8 (24.4)
Median [Min, Max] 80.0 [48.0, 133] 67.0 [43.0, 115] 75.0 [43.0, 133]
Missing 0 (0%) 1 (7.1%) 1 (3.3%)
GGTP
Mean (SD) 29.6 (12.5) 34.8 (23.5) 31.9 (18.1)
Median [Min, Max] 25.5 [11.0, 49.0] 28.0 [10.0, 79.0] 27.0 [10.0, 79.0]
Missing 0 (0%) 1 (7.1%) 1 (3.3%)
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3.1 Total protein

Abnormally low levels of total protein may indicate a problem with protein digestion or
absorption or a liver or kidney disorder. In our clinical trial, the initial and end-of-study levels
of total protein are as represented in Tables 19 and 20.

Table 19: Total protein levels (mg/dL) during Visit-1 in group-1 (G1, treatment group) and
group-2 (G2, placebo group)
Total Protein G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 7.24 (0.373) 7.36 (0.323) 7.30 (0.348)
Median (Min, Max) 7.09 (6.81, 7.94) 7.34 (6.64, 7.91) 7.31 (6.64, 7.94)
Missing 1 (6.3%) 0 (0%) 1 (3.3%)

Table 20: Total protein levels (mg/dL) during Visit-5 in group-1 (G1, treatment group) and
group-2 (G2,placebo group)

Total Protein G1 (N=16) G2 (N=14) Overall (N=30)

Mean (SD) 7.54 (0.400) 7.73 (0.343) 7.62 (0.381)
Median (Min, Max) 7.47 (6.97, 8.25) 7.59 (7.33, 8.32) 7.59 (6.97, 8.32)
Missing 0 (0%) 1 (7.1%) 1 (3.3%)

Figure7: Effect of Lutein and Zeaxanthin supplementation on Total protein levels

G1V1 (total protein levels measured in group-1 at visit-1); G1V5 (total protein levels
measured in group-1 at visit-5); G2V1 (total protein levels measured in group-2 at visit-1);
G2V5 (total protein levels measured in group-2 at visit-5). The central line in the box plots
represents the median values, and the rectangular box shows the interquartile range (IQR),
which is the middle 50% of the data. The whiskers extend to the minimum and maximum
values of the data, except for the outliers that fall beyond the whiskers. Statistical significance
was calculated using a paired, two-sided t-test. *** indicates p<0.001, ## indicates p<0.01
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3.2 Serum Albumin and globulin levels

As indicated in the two tables below, the average albumin levels in groups 1 and 2 at visit 1
were 4.32 and 4.61, respectively. Following treatment for 5 weeks, at visit 5, the albumin
levels in G1 and G2 were 4.62 and 4.77 mg/Dl, respectively (Tables 21 and 22 and Figure 8).
Serum globulin levels were also measured during the clinical study, and the levels are as
indicated below for G1 and G2 during visits 1 and 5 (Tables 23 and 24 and Figure 9).

Table 21: Serum albumin levels (mg/dL) during Visit-1 in group-1 (G1, treatment group) and
group-2 (G2, placebo group

Albumin G1 (N=16) G2 (N=14) Overall (N=30)

Mean (SD) 4.32 (0.251) 4.61 (0.245) 4.46 (0.286)
Median (Min, Max) 4.25 (4.03, 4.91) 4.64 (4.02, 5.10) 4.45 (4.02, 5.10)
Missing 1 (6.3%) 0 (0%) 1 (3.3%)

Table 22: Serum albumin levels (mg/dL) during Visit-5 in group-1 (G1, treatment group) and
group-2 (G2, placebo group
Albumin G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 4.62 (0.281) 4.77 (0.200) 4.69 (0.254)
Median (Min, Max) 4.56 (4.26, 5.14) 4.74 (4.45, 5.15) 4.69 (4.26, 5.15)
Missing 0 (0%) 1 (7.1%) 1 (3.3%)

Figure 8: Effect of Lutein supplementation on Albumin levels

G1V1 (Albumin levels measured in group 1 at visit 1); G1V5 (Albumin levels measured in
group 1 at visit 5); G2V1 (Albumin levels measured in group 2 at visit 1); G2V5 (Albumin
levels measured in group 2 at visit 5). The central line in the box plots represents the median
values, and the rectangular box shows the interquartile range (IQR), which is the middle 50%
of the data. The whiskers extend to the minimum and maximum values of the data, except for
the outliers that fall beyond the whiskers. Statistical significance was calculated using a
paired, two-sided t-test. *** indicates p<0.001
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Table 23: Serum globulin levels (mg/dL) during Visit-1 in group-1 (G1, treatment group)
and group-2 (G2, placebo group)

Globulin G1 (N=16) G2 (N=14) Overall (N=30)

Mean (SD) 2.93 (0.322) 2.74 (0.281) 2.84 (0.312)
Median (Min, Max) 2.86 (2.57, 3.87) 2.72 (2.27, 3.31) 2.78 (2.27, 3.87)
Missing 1 (6.3%) 0 (0%) 1 (3.3%)

Table 24: Serum globulin levels (mg/dL) during Visit-5 in group-1 (G1, treatment group)
and group-2 (G2, placebo group)
Globulin G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 2.92 (0.401) 2.96 (0.322) 2.94 (0.362)
Median (Min, Max) 2.79 (2.21, 3.90) 3.03 (2.47, 3.48) 2.80 (2.21, 3.90)
Missing 0 (0%) 1 (7.1%) 1 (3.3%)

Figure 9: Effect of Lutein and Zeaxanthin supplementation on Globulin levels

G1V1 (globulin levels measured in group-1 at visit-1); G1V5 (globulin levels measured in
group-1 at visit-5); G2V1 (globulin levels measured in group-2 at visit-1); G2V5 (globulin
levels measured in group-2 at visit-5). The central line in the box plots represents the median
values, and the rectangular box shows the interquartile range (IQR), which is the middle 50%
of the data. The whiskers extend to the minimum and maximum values of the data, except for
the outliers that fall beyond the whiskers. Statistical significance was calculated using a
paired, two-sided t-test. # indicates p<0.05
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3.3 Albumin Globulin ratio

Tables 25 and 26 and Figure 10 indicate the values of the albumin-to-globulin ratio (AG
ratio) for two groups during visits 1 and 5.

Table 25: Albumin to Globulin ratio during Visit-1 in group-1 (G1, treatment group) and
group-2 (G2, placebo group)
Albumin-Globulin G1 (N=16) G2 (N=14) Overall (N=30)
ratio
Mean (SD) 1.49 (0.174) 1.70 (0.229) 1.59 (0.225)
Median (Min, Max) 1.51 (1.05, 1.76) 1.68 (1.39, 2.25) 1.54 (1.05, 2.25)
Missing 1 (6.3%) 0 (0%) 1 (3.3%)

Table 26: Albumin to Globulin ratio during Visit-5 in group-1 (G1, treatment group) and
group-2 (G2, placebo group)

Albumin-Globulin G1 (N=16) G2 (N=14) Overall (N=30)

ratio
Mean (SD) 1.62 (0.260) 1.63 (0.206) 1.62 (0.233)
Median (Min, Max) 1.61 (1.12, 2.15) 1.66 (1.35, 2.01) 1.65 (1.12, 2.15)
Missing 0 (0%) 1 (7.1%) 1 (3.3%)

Figure 10: Effect of Lutein and Zeaxanthin supplementation on Albumin Globulin ratio levels

G1V1 (Alb/Glob ratio measured in group-1 at visit-1); G1V5 (Alb/Glob ratio measured in
group-1 at visit-5); G2V1 (Alb/Glob ratio measured in group-2 at visit-1); G2V5 (Alb/Glob
ratio measured in group-2 at visit-5). The central line in the box plots represents the median
values, and the rectangular box shows the interquartile range (IQR), which is the middle 50%
of the data. The whiskers extend to the minimum and maximum values of the data, except for
the outliers that fall beyond the whiskers. Statistical significance was calculated using a
paired, two-sided t-test. * indicates p<0.05
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3.4 Total Bilirubin

The total bilirubin levels for G1 and G2 were 0.609 and 0.547 mg/dL, respectively, during visit
1. The corresponding values during visit 5 were 0.633 and 0.662 for G1 and G2, respectively.
The comparisons are represented in Tables 27 and 28 and graphically in Figure 11.

Table 27: Bilirubin (total) levels (mg/dL) during Visit-1 in group-1 (G1, treatment group)
and group-2 (G2, placebo group)
Total bilirubin G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 0.609 (0.359) 0.547 (0.315) 0.580 (0.335)
Median (Min, Max) 0.480 (0.240, 1.59) 0.470 (0.240, 1.42) 0.470 (0.240, 1.59)

Table 28: Bilirubin (total) levels (mg/dL) during Visit-5 in group-1 (G1, treatment group)
and group-2 (G2, placebo group)
Total bilirubin G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 0.633 (0.426) 0.662 (0.427) 0.646 (0.419)
Median (Min, Max) 0.505 (0.300, 2.04) 0.420 (0.230, 1.45) 0.500 (0.230, 2.04)
Missing 0 (0%) 1 (7.1%) 1 (3.3%)

Figure 11: Effect of Lutein and Zeaxanthin supplementation on Total-Bilirubin levels

G1V1 (bilirubin-total levels measured in group-1 at visit-1); G1V5 (bilirubin-total levels

measured in group-1 at visit-5); G2V1 (bilirubin-total levels measured in group-2 at visit-1);
G2V5 (bilirubin-total levels measured in group-2 at visit-5). The central line in the box plots
represents the median values, and the rectangular box shows the interquartile range (IQR),
which is the middle 50% of the data. The whiskers extend to the minimum and maximum
values of the data, except for the outliers that fall beyond the whiskers. Statistical significance
was calculated using a paired, two-sided t-test. No significance was observed for
corresponding comparisons.
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3.5 Direct bilirubin

Tables 29 and 30 showed the total bilirubin levels for G1 and G2 were 0.183 and 0.176
mg/dL, respectively, during visit 1. The corresponding values during visit 5 were 0.217 and
0.219 mg/dL for G1 and G2, respectively. The comparisons are represented graphically in
Figure 12.

Table 29: Bilirubin (direct) levels (mg/dL) during Visit-1 in group-1 (G1, treatment group)
and group-2 (G2, placebo group)
Bilirubin direct G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 0.183 (0.0814) 0.176 (0.0803) 0.180 (0.0795)
Median (Min, Max) 0.160 (0.100, 0.410) 0.145 (0.0900, 0.360) 0.155 (0.0900, 0.410)

Table 30: Bilirubin (direct) levels (mg/dL) during Visit-5 in group-1 (G1, treatment group)
and group-2 (G2, placebo group)
Bilirubin direct G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 0.217 (0.0993) 0.219 (0.0993) 0.218 (0.0975)
Median (Min, Max) 0.195 (0.110, 0.500) 0.180 (0.100, 0.420) 0.190 (0.100, 0.500)
Missing 0 (0%) 1 (7.1%) 1 (3.3%)

Figure 12: Effect of Lutein and Zeaxanthin supplementation on Bilirubin direct levels

G1V1 (bilirubin-direct levels measured in group-1 at visit-1); G1V5 (bilirubin-direct levels

measured in group-1 at visit-5); G2V1 (bilirubin-direct levels measured in group-2 at visit-1);
G2V5 (bilirubin-direct levels measured in group-2 at visit-5). The central line in the box plots
represents the median values, and the rectangular box shows the interquartile range (IQR),
which is the middle 50% of the data. The whiskers extend to the minimum and maximum
values of the data, except for the outliers that fall beyond the whiskers.
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3.6 Indirect bilirubin:

Tables 31 and 32 show the indirect bilirubin levels for G1 and G2 during visits 1 and 5.

Table 31: Bilirubin(indirect) levels (mg/dL) during Visit-1 in group-1 (G1, treatment group)
and group-2 (G2, placebo group)
Bilirubin indirect G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 0.426 (0.279) 0.371 (0.241) 0.400 (0.259)
Median (Min, Max) 0.315 (0.130, 1.18) 0.310 (0.150, 1.06) 0.310 (0.130, 1.18)

Table 32: Bilirubin (indirect) levels (mg/dL) during Visit-5 in group-1 (G1, treatment group)
and group-2 (G2, placebo group)

Bilirubin indirect G1 (N=16) G2 (N=14) Overall (N=30)

Mean (SD) 0.416 (0.333) 0.443 (0.333) 0.428 (0.327)
Median (Min, Max) 0.325 (0.190, 1.54) 0.280 (0.100, 1.06) 0.310 (0.100, 1.54)
Missing 0 (0%) 1 (7.1%) 1 (3.3%)

3.7 SGPT and SGOT:

The mean values of SGPT in different groups of participants during visits 1 and 2 are
presented in tables 33 and 34 and Figure 13. The mean values of SGOT in different groups of
participants during visits 1 and 2 are presented in tables 35 and 36 and Figure 14.

Table 33: SGPT levels (IU/L) during Visit-1 in group-1 (G1, treatment group) and group-2
(G2, placebo group)
SGPT G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 20.4 (7.36) 31.4 (15.2) 25.5 (12.8)
Median (Min, Max) 18.0 (11.0, 38.8) 30.5 (8.70, 62.0) 23.0 (8.70, 62.0)

Table 34: SGPT levels (IU/L) during Visit-5 in group-1 (G1, treatment group) and group-2
(G2, placebo group)

SGPT G1 (N=16) G2 (N=14) Overall (N=30)

Mean (SD) 28.2 (14.2) 30.2 (13.7) 29.1 (13.7)
Median (Min, Max) 27.0 [14.0, 61.0] 27.0 [10.1, 55.0] 27.0 [10.1, 61.0]
Missing 0 (0%) 1 (7.1%) 1 (3.3%)
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Figure 13: Effect of Lutein and Zeaxanthin supplementation on SGPT levels

G1V1 (SGPT levels measured in group-1 at visit-1); G1V5 (SGPT levels measured in group-
1 at visit-5); G2V1 (SGPT levels measured in group-2 at visit-1); G2V5 (SGPT levels
measured in group-2 at visit-5). The central line in the box plots represents the median values,
and the rectangular box shows the interquartile range (IQR), which is the middle 50% of the
data. The whiskers extend to the minimum and maximum values of the data, except for the
outliers that fall beyond the whiskers. Statistical significance was calculated using a paired,
two-sided t-test. No significance was observed for corresponding comparisons.

Table 35: SGOT levels (IU/L) during Visit-1 in group-1 (G1, treatment group) and group-2
(G2, placebo group)
SGOT G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 21.5 (5.79) 24.3 (5.88) 22.8 (5.90)
Median (Min, Max) 21.5 (14.0, 35.0) 23.5 (16.2, 41.0) 22.0 (14.0, 41.0)

Table 36: SGOT levels (IU/L) during Visit-5 in group-1 (G1, treatment group) and group-2
(G2, placebo group)

SGOT G1 (N=16) G2 (N=14) Overall (N=30)

Mean (SD) 26.1 (13.9) 27.0 (8.70) 26.5 (11.6)
Median (Min, Max) 23.3 (13.0, 72.0) 26.0 (17.4, 51.0) 25.0 (13.0, 72.0)
Missing 0 (0%) 1 (7.1%) 1 (3.3%)
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Figure 14: Effect of Lutein and Zeaxanthin supplementation on SGOT levels

G1V1 (SGOT levels measured in group-1 at visit-1); G1V5 (SGOT levels measured in
group- 1 at visit-5); G2V1 (SGOT levels measured in group-2 at visit-1); G2V5 (SGOT
levels measured in group-2 at visit-5). The central line in the box plots represents the median
values, and the rectangular box shows the interquartile range (IQR), which is the middle 50%
of the data. The whiskers extend to the minimum and maximum values of the data, except for
the outliers that fall beyond the whiskers. Statistical significance was calculated using a
paired, two-sided t-test. No significance was observed for corresponding comparisons.
3.8 Alkaline phosphatase (ALP):
In Tables 37 and 38 and Figure 15 below, the mean ALP levels increased from 77.9 to 87.2
IU/L in G-1 between visits 1 and 5. Whereas in G-2, the mean ALP levels were 70.2 and 68.4
IU/L for visits 1 and 5, respectively. These changes were statistically significant as estimated
by a paired, two-tailed t-test.

Table 37: ALP levels (IU/L) during Visit-1 in group-1 (G1, treatment group) and group-2
(G2, placebo group)
ALP G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 77.9 (21.1) 70.2 (16.3) 74.2 (19.0)
Median (Min, Max) 74.0 (49.0, 123) 65.0 (46.0, 108) 67.0 (46.0, 123)
Missing 1 (6.3%) 0 (0%) 1 (3.3%)
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Table 38: ALP levels (IU/L) during Visit-5 in group-1 (G1, treatment group) and group-2
(G2, placebo group)

ALP G1 (N=16) G2 (N=14) Overall (N=30)

Mean (SD) 87.2 (25.7) 68.4 (18.7) 78.8 (24.4)
Median (Min, Max) 80.0 (48.0, 133) 67.0 (43.0, 115) 75.0 (43.0, 133)
Missing 0 (0%) 1 (7.1%) 1 (3.3%)

Figure 15: Effect of Lutein and Zeaxanthin supplementation on ALP levels

G1V1 (ALP levels measured in group-1 at visit-1); G1V5 (ALP levels measured in group-1
at visit-5); G2V1 (ALP levels measured in group-2 at visit-1); G2V5 (ALP levels measured
in group-2 at visit-5). The central line in the box plots represents the median values, and the
rectangular box shows the interquartile range (IQR), which is the middle 50% of the data.
The whiskers extend to the minimum and maximum values of the data, except for the outliers
that fall beyond the whiskers. Statistical significance was calculated using a paired, two-sided
t- test. P<0.05.
3.9 Gamma Glutamyl Transpeptidase (GGTP)
The mean GGTP levels for G1 were 25.9 and 29.6 IU/L for visits 1 and 5, respectively.
Similarly, for G2, the GGTP levels were 33.3 and 34.8 IU/L for visits 1 and 5, respectively.
The differences were not statistically significant (Tables 39 and 40 and Figure 16).
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Table 39: GGTP levels (IU/L) during Visit-1 in group-1 (G1, treatment group) and group-2
(G2, placebo group)

GGTP G1 (N=16) G2 (N=14) Overall (N=30)

Mean (SD) 25.9 (18.8) 33.3 (22.0) 29.5 (20.4)
Median (Min, Max) 22.0 (11.0, 90.0) 26.5 (11.0, 83.0) 25.0 (11.0, 90.0)
Missing 1 (6.3%) 0 (0%) 1 (3.3%)

Table 40: GGTP levels (IU/L) during Visit-5 in group-1 (G1, treatment group) and group-2
(G2, placebo group)
GGTP G1 (N=16) G2 (N=14) Overall (N=30)
Mean (SD) 29.6 (12.5) 34.8 (23.5) 31.9 (18.1)
Median (Min, Max) 25.5 (11.0, 49.0) 28.0 (10.0, 79.0) 27.0 (10.0, 79.0)
Missing 0 (0%) 1 (7.1%) 1 (3.3%)

Figure 16: Effect of Lutein and Zeaxanthin supplementation on GGTP levels

G1V1 (GGTP levels measured in group-1 at visit-1); G1V5 (GGTP levels measured in
group- 1 at visit-5); G2V1 (GGTP levels measured in group-2 at visit-1); G2V5 (GGTP
levels measured in group-2 at visit-5). The central line in the box plots represents the median
values, and the rectangular box shows the interquartile range (IQR), which is the middle 50%
of the data. The whiskers extend to the minimum and maximum values of the data, except for
the outliers that fall beyond the whiskers. Statistical significance was calculated using a
paired, two-sided t-test, and there were no significant differences between either of the groups
for visits 1 and 5. And here the Table 41 were showing the criteria for classifying the
outcome of study in liver injury
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Table 41: The criteria for classifying the outcome of study in liver injury
Type of injury Enzymatic profile
Hepatocellular ALT > 2ULN
Serum ALT/Serum
Alk. Phos ≥ 5*
Cholestatic Alk Phos ≥ 2ULN
Serum ALT/Serum
Alk Phos ≤ 2*
Mixed ALT > 2 ULN
Serum ALT/Serum Alk
Phos between 2 and 5*

Based on the above results LFT to represent hepatocellular injury in the ALT levels should be
twice more than the upper limit of normal (ULN) and similarly there are specific criteria for
identifying cholestatic and mixed types of injury. In our studies, most of the individuals either
in the group-1 or group-2 did not show increase corresponding to more than twice of ULN for
these parameters. In addition, for neither of the groups the LFT parameters showed any
statistically significant increase.
DISCUSSION
Numerous research evaluated the impact of lutein on normal and diabetic rats. It was
discovered that diabetic rats with dose-dependent extract ingestion had lower blood glucose
levels than their healthy equivalents. According to the Hosseini et al. study, when compared
to the values at the start of the trial and those of the control group, the levels of FBS, HbA1c,
total cholesterol, and triglycerides dramatically decreased in the diabetic patients receiving
lutein treatment (38). The American Diabetes Association advises using HbA1c with a cut-
point 6.5% for diagnosing diabetes as an alternative to fasting plasma glucose since it offers a
reliable marker of chronic glycemia and corresponds well with the risk of long-term diabetes
complications (56). Additionally, HbA1c is an excellent predictor of lipid profile, offering the
additional benefit of recognizing cardiovascular risk in diabetic patients (57).
So, when the Lutein and zeaxanthin were compared to placebo, they reduced the HbA1c
levels. The analysis of HbA1c levels, estimated average glucose levels (eAG), and random
glucose levels showed good results in newly diagnosed diabetic patients. These results
suggest that Lutein and zeaxanthin can be a beneficial supplement for diabetic patients with
type 2 diabetes. Hypertension, aging, hemodynamic dysregulation, proteinuria, and a high
consumption of dietary protein are some of the conditions that may accelerate kidney
damage. A significant risk factor for the advancement of CKD, cardiovascular disease, and
all-cause mortality has been found to be proteinuria, which is unrelated to kidney
disease. Effective proteinuria
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management is important as a key therapy response indicator in a number of renal disorders.

Kidney function tests are utilized in this clinical research to evaluate the safety profile of the
supplement and any potential kidney side effects. The tests aid in figuring out whether the
supplement is having an impact on kidney function indicators including creatinine, blood
urea nitrogen, and serum urea. A common laboratory test to determine the level of urea
nitrogen in the blood is the blood urea nitrogen (BUN) test. The waste product urea is
expelled by the kidneys and is created when proteins are broken down in the liver. The study
results indicate that treatment with lutein did not increase blood urea nitrogen. Urea is
primarily eliminated via the kidneys, with the remaining portion excreted through the
gastrointestinal tract. Elevated levels of urea in the blood may indicate renal impairment or
other conditions such as dehydration, high protein diets, and upper GI bleeding. In contrast,
decreased levels of urea may indicate low-protein diets, starvation, or severe liver disease.
The serum urea level is a commonly used laboratory test to assess kidney function and
monitor kidney disease. Serum creatinine level is another parameter commonly measured in
blood tests to assess kidney function, as the kidneys are responsible for filtering and excreting
creatinine from the body. Elevated serum creatinine levels can indicate decreased kidney
function or damage to the kidneys, while low levels may indicate decreased muscle mass or
other medical conditions. However, the interpretation of serum creatinine levels should be
done in conjunction with other clinical information, such as the patient's age, sex, weight, and
medical history. The serum creatinine levels did not show any significant variation in either
of the groups during visit 5 in comparison to visit 1, according to the findings of this study.
These parameters can indicate whether the supplement is causing damage to the kidneys or
affecting their ability to function correctly. Therefore, it is essential to monitor kidney
function parameters regularly during clinical trials.
In India, a significant number of laboratories rely on RI (reference intervals) derived from
literature that primarily focuses on Western populations. However, the applicability of these
RI is questionable due to disparities stemming from differences in diet, lifestyle,
environmental conditions, and ethnicity-related variations. Keeping this in mind, a recent
study was conducted on 2,021 healthy individuals in India to provide reference intervals (RI)
for certain liver- specific biochemical parameters (58). The study measured several blood
parameters and reported reference ranges for several liver-specific parameters. According to
this study, bilirubin had a wide variation in the observed RI (0.30–1.30) compared to the
reported RI (0– 1.2). Similarly, SGOT showed a wide variation (13-52.80) compared to the
reported RI (0-40). SGPT and ALP also had wide variations in the observed RI (10–68 for
SGPT and 107–361.80
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for ALP) compared to their reported standard RI (0–50 for SGPT and 110–310 for ALP).
However, GGT, a marker for biliary disease and alcoholism, remained within the reported
standard RI (0–50, 5.00–50.60). The total protein test is useful in identifying various health
conditions, such as liver disease, kidney disease, and malnutrition resulting from inadequate
nutrient intake. Abnormally low levels of total protein may indicate a problem with protein
digestion or absorption or a liver or kidney disorder. In both groups, the total protein levels
did not decrease, indicating that the treatment did not have any negative effects. In fact, in
both conditions, the total protein levels increased, and this increase was statistically
significant. However, the interpretation of an increase in total protein in a physiological
context is complex. From the results, we can only infer that the total protein levels did not
reduce during the treatment. Serum albumin is a protein that is produced by the liver and is
found in the blood. It plays an essential role in maintaining the osmotic pressure of the blood
and helps to transport various substances, including hormones, drugs, and fatty acids. Low
levels of serum albumin can indicate liver or kidney disease, malnutrition, or inflammation.
Monitoring serum albumin levels during a clinical trial is critical in order to evaluate the
effects of the intervention on liver function. There are several drugs that can lower serum
albumin levels, either by reducing albumin production or increasing its breakdown. In both
groups, the albumin levels did not decrease, indicating that the treatment did not have any
negative effects. In fact, in G1, albumin levels increased, and this increase was statistically
significant. However, the interpretation of an increase in total protein in a physiological
context is complex. From the results, we can only infer that the total protein levels did not
reduce during the treatment. In either G1 or G2, during the clinical trial, the serum globulin
levels did not drop below 2 mg/dL, indicating that the treatment did not have negative effects.
Albumin is the most abundant protein in the blood and plays a crucial role in maintaining
oncotic pressure, transporting various substances (such as hormones and drugs), and
regulating fluid balance. Globulins, on the other hand, are a diverse group of proteins
involved in immune function, blood clotting, and transporting other substances. The A/G
ratio is used as a marker to evaluate liver and kidney function, nutritional status, and certain
medical conditions. In liver disease, impaired albumin production leads to a decreased A/G
ratio. Kidney damage affects protein filtration, altering the A/G ratio. Inflammation and
infection can raise certain globulin levels, increasing the A/G ratio. Total bilirubin, indirect
bilirubin, and direct bilirubin are the essential markers of liver health and function, as
elevated levels of total bilirubin can indicate liver damage or dysfunction, as well as other
medical conditions affecting the liver.
medRxiv preprint doi: https://doi.org/10.1101/2023.09.22.23295947; this version posted September 29, 2023. The copyright holder for this preprint
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Both SGPT (Serum Glutamate Pyruvate Transaminase, or ALT) and SGOT (Serum Glutamic
Oxaloacetic Transaminase, or AST) are liver enzymes commonly used in liver function tests.
While both enzymes provide valuable information about liver health, their interpretations and
clinical significance differ. SGPT (ALT) is primarily found in the liver, and elevated levels
typically indicate liver damage or injury. It is considered more specific to liver function,
making it a reliable marker for hepatocellular damage such as viral hepatitis, liver cirrhosis,
or drug-induced liver injury. SGOT (AST), on the other hand, is present not only in the liver
but also in other organs like the heart, skeletal muscles, and kidneys. Therefore, elevated
SGOT levels may not solely reflect liver-specific issues. Increased SGOT levels can be
associated with liver damage but can also be caused by conditions affecting other organs or
tissues.
ALP (alkaline phosphatase) is a hydrolase enzyme that is widely distributed in various
tissues, including the liver, bone, intestine, and placenta. In the context of liver function tests
during clinical safety validation, ALP is primarily used as a marker for hepatobiliary system
evaluation. Elevated ALP levels can indicate cholestasis, obstructive liver diseases, or hepatic
cell injury. It is particularly useful in distinguishing between hepatocellular and cholestatic
liver disorders. ALP measurement aids in assessing liver health, monitoring disease
progression, and determining the potential hepatotoxic effects of drugs. In clinical practice,
ALP serves as a valuable biochemical parameter to enhance the diagnostic accuracy and
safety assessment of liver function. GGTP is a microsomal enzyme found in various cells,
including hepatocytes, biliary epithelial cells, renal tubules, the pancreas, and the intestine. It
plays a role in transporting peptides across the cell membrane and is involved in glutathione
metabolism. Although GGTP is present in renal tissue, its activity in the serum is primarily
associated with the hepatobiliary system. Acute viral hepatitis and alcoholism can
significantly increase the levels of GGTP. Additionally, GGTP can serve as an early marker
of oxidative stress.
Measurements of bone mineral density and clinical risk factors are now utilized to determine
who is at risk for osteoporosis. ALP is an enzyme that is made in the liver, bones, intestines,
and kidneys and gets into the bloodstream. Total serum ALP levels as a bone-forming
biomarker have been shown in studies to be able to predict how well medication therapy for
osteoporosis is working (59, 60). Today, there have been some documented clinical trials on
how people with osteoporosis respond to herbal treatment. A recent Cochrane systematic
review showed that most included studies (85 of 108) had small sample sizes, ranging from
20 to 120, and that the overall quality of the evidence was generally low. The use of herbal
medicine among people with osteoporosis is therefore not supported by clear evidence, and
larger sample sizes and more thoroughly planned research are needed. It should be mentioned
medRxiv preprint doi: https://doi.org/10.1101/2023.09.22.23295947; this version posted September 29, 2023. The copyright holder for this preprint
(which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.
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that alkaline phosphatase (ALP) is a crucial enzyme that is essential for the development of
hard tissue, especially mineralized tissue, as well as for liver function. According to research,
extracellular pyrophosphate, which is an inhibitor of mineral formation, and inorganic
phosphate local rates are both increased by ALP, which promotes mineralization (61). Total
alkaline phosphatase levels are actually shown to be a measure of increased osteoblastic
activity and bone conversion status (62). According to a study by Takeda et al., four weeks of
lutein treatment significantly increased the mass of the femoral bones, particularly the cortical
bone. This improvement was quantified by dual X-ray absorptiometry and micro computed
tomography (CT) assessments of bone mineral density (63). Male hip fracture risk has been
demonstrated to be negatively linked with dietary intake of the carotenoids α-carotene, β-
carotene, and lutein (64). Similar findings were made by Zhang et al (65) who discovered that
higher serum carotenoid levels were associated with greater bone mineral density in both men
and women. Lutein, which is well known for its antioxidant abilities, contributes to bone
remodelling by acting as an antioxidant. Our research suggests that supplementing with
Lutein and zeaxanthin may have a beneficial effect on bone formation. However, more
research is need to fully understand this occurrence.
CONCLUSION
In this study, the effects of Lutein and zeaxanthin on blood sugar levels, kidney, liver, and
bone health are described in depth. Five weeks of lutein supplementation improved the levels
of HbA1c. When examining the safety and effectiveness of supplements in clinical studies,
kidney function tests are essential. This is so that waste and extra fluid can be removed from
the body, a task that the kidneys are extremely important for performing. The build-up of
toxins in the body due to impaired renal function can have negative impacts on health. It is
possible that lutein supplementation is not harmful to renal function, according to parameters
such as blood urea nitrogen, serum urea levels, and creatinine levels. Our clinical
investigation revealed that the Lutein and zeaxanthin had a good impact on these kidney
function markers and was safe. The liver function tests suggest that using lutein supplements
may not pose any hazards for the liver. The SGOT and SGPT are the key factors in
identifying the levels of liver damage. Alkaline phosphatase levels increased when lutein was
administered. However, the level of the ALP increase was not sufficiently significant to be
categorized as a liver damage case. Lutein supplementation is known to be beneficial for
bone growth. Considering the role of ALP (Alkaline Phosphatase) in bone mineralisation, the
small change in ALP in this study indicates that Lutein may have effect in improving the
bone health and yet at the same time the increase in the values of ALP was not statistically
significant proving the safety aspects of lutein.
medRxiv preprint doi: https://doi.org/10.1101/2023.09.22.23295947; this version posted September 29, 2023. The copyright holder for this preprint
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However, it is hoped that through this meticulous validation, the findings of this randomized
controlled trial would demonstrate the effectiveness and safety of Lutein and zeaxanthin for
bone health and serve as a solid justification for conducting additional research for better
activity.
medRxiv preprint doi: https://doi.org/10.1101/2023.09.22.23295947; this version posted September 29, 2023. The copyright holder for this preprint
(which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.
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