1 Department of Pharmacology, Faculty of Pharmacy, Ziauddin University, Karachi, Pakistan
2 Department of Pharmacognosy, Faculty of Pharmacy, Ziauddin University, Karachi, Pakistan
Corresponding Author Email:
Objective: Ageing is the natural process of getting old and involve deleterious phenomenon like glycation and oxidative stress. The quality of life can be improved and maintained for longer duration of time by inhibiting these harmful processes. In this regard, the dietary substances, especially spices hold tremendous potential.
Methods: The present work was designed to assess the anti-ageing potential of five spices i.e. cumin seed, cinnamon, ginger, paprika and garlic, using anti-glycation and anti-oxidant assays along with HPLC fingerprint of the most potent extract.
Results: Our data showed that among all spices, the ethanolic extract (70%) of cumin seeds showed the most significant inhibition of glycation (80%) and oxidative stress (95%) at the tested dose of 1mg/ml. The Venn diagram (set at a threshold of 50%) showed that among all tested extracts, only cumin seed was able to produce 50% inhibition in both assays. The HPLC fingerprint of cumin seed extract was also obtained as the crude reflection of the phytochemicals residing in it.
Conclusion: Hence, the cumin seed presents itself as a promising functional food for the management of age-associated diseases and diabetic complications attributed to the deleterious phenomenon of glycation.
Keywords: Glycation; Oxidative stress; Cumin seeds; Functional food
Senescence or ageing is the natural phenomenon of getting old, which is jeopardizing the human race in an unprecedented manner. The world needs balanced demography to run its machinery. However, the number of old age people shall surpass youth by the year 2050; an event that is unprecedented in known human history (Suzman and Beard, 2011). The scenario is alarming and stresses upon the need to take appropriate measures to increase quality of life over time. Several underlying pathological mechanisms have been reported for the process of ageing (Campisi et al., 2019). Among which, one of them attributes senescence to the buildup of garbage in the body (Sergiev et al., 2015) such as Advance Glycation End products (AGEs) (Ross, 2015) and oxidative stress (imbalance between oxidants and antioxidants) (Pole et al., 2016). Glycation (Maillard reaction) involves interaction between carbohydrates; especially reducing sugar and other bio-molecules i.e. nucleic acids, proteins or lipids, which after a long process produces AGEs (Goldin et al., 2006). The levels of N(6)-carboxymethyl lysine (CML), the predominant AGEs product, co-relates with the phenomenon of ageing, age-linked diseases such as learning & memory impairment (Southern et al., 2007, Igase and Igase, 2018), kidney injury (Sun et al., 2016), non-alcoholic fatty liver diseases (Leung et al., 2016) and complications associated with diabetes mellitus (Ramasamy et al., 2005). The AGEs binds to its receptor known as Receptor for Advanced Glycation End products (RAGE), thereby leading to activation of Nuclear Factor-kappa B (NF-кB) (Haslbeck et al., 2005); the transcription factor famous for oxidative stress. The stress in this case is specifically termed as glycative stress (Yonei et al., 2020).
In quest of impeding this deleterious phenomenon of glycation, several inhibitors (natural and synthetic) were reported (Reddy and Beyaz, 2006). Despite of their significant effectiveness (e.g. aminoguanidine), they could not make it to the bed side due to their potential to produce adverse effects (Thornalley, 2003, Campbell, 1996). Under this milieu, the field of nutritional pharmacology comes to rescue. This is an emerging field in pharmacy, which uses the concept of functional foods i.e. the food resources which along with nutritional value also offer medicinal value. This concept was formulated by Hippocrates i.e. let food be thy medicine and medicine be thy food (Hasler, 2002). The benefit of using this concept is that it offers fast and inexpensive solutions without needing detailed toxicological assessments as part of drug development process. Furthermore, this type of approach suits well for the indications which grow insidiously.
Dietary spices hold promising potential as functional food and need comprehensive investigations (Dearlove et al., 2008). In this regard, cumin is the famous spice used in folklore for the management of tooth ache, gastrointestinal, inflammatory and neurological disorders (Benelli et al., 2018). Other biological actions attributed to cumin are antioxidant, anticancer, stimulant and carminative (Sharma et al., 2001). Cumin is actually the dried seed obtained from Cuminum cyminum L., a medicinal plant of Apiaceae family, which is native to the Eastern Mediterranean and South Asia (Singh et al., 2017, Zarandi et al., 2017).
Currently, there is no treatment option available in the allopathic system of medicine for the management of glycation. Keeping this in view, the present study was designed to evaluate functional food i.e. spices for their ability to inhibit the deleterious phenomenon of glycation.
MATERIALS & METHODS
Selection of Spices
Five spices i.e. cumin seed, cinnamon, ginger, paprika and garlic were selected for the present study.
Preparation of plant extract
Briefly, 1 kg of each spice was separately soaked in ethanol (70%, 3L) for three days at room temperature, followed by rotary evaporation to yield thick extract, which was freeze dried and stored in refrigerator till further use.
The in vitro AGE inhibition assay was performed by incubating BSA (10 mg/ml) and fructose (50mM) together for 24 hours at 60 ˚C. The spice extract was used at the concentration of 1mg/ml. The AGEs were detected using its innate fluorescence (Khan et al., 2017). ) All results were tested in triplicate. The percentage inhibition of AGEs was measured using the following formula
% Inhibition = [1-(fluorescence of test)] / (fluorescence of control) x 100
Free radical scavenging activity of functional foods was assessed by 1-1diphenyl-2-picryl-hydrazyl (DPPH) as reported earlier (Shen et al., 2010). Briefly, DPPH (0.1 mM in methanol) was added (100ul) to spice extracts (1mg/ml, 300ul), vortexed and allowed to stand at room temperature for 30 minutes. The absorbance was measured at 517nm using UV spectrophotometer. Percent free radical scavenging activity was calculated using the following formula (Aiyegoro and Okoh, 2010):
DPPH Scavenging activity (%) = [(Abs control – Abs sample) / Abs control] x 100
The Venn diagram was prepared at 50% threshold to observe the effect of spice extracts on both anti-glycation and anti-oxidant assays.
The High Performance Liquid Chromatography (HPLC) was used to obtain the fingerprint of the extract. Following were the working conditions:
Extract Strength: 50µg/ml
Column: Hibar® 250-4, 6 LiChrospher® 100 RP-18e (5um) column.
Detector: DAD (SPD-M20A) set at 250nm.
Mobile phase: Methanol (50%)
Flow rate: 0.5ml/min
Injection volume: 10µl
The data is shown as mean ± SEM of percent change.
The results obtained in the present study are as follows:
Among the various spices, the cumin seeds extract showed the most promising anti-glycation action with the percent inhibition of 80% (Table-1). Cinnamon, ginger, paprika and garlic showed the percent inhibition of 30%, 25%, 21% and 10% respectively.
Table-1 Effect of Various Spice Extracts on Inhibition of Glycation
|S. No.||Functional Food||Scientific Name||AGE Inhibition (%)|
|1||Cumin seed||Cuminum cyminum||80|
|4||Paprika (red)||Capsicum annum||21|
|Aminoguanidine 3mM (positive control)||71|
Among the various spices, the cumin seeds extract showed the most promising anti-oxidant action with the percent inhibition of 95% (Table-2). Cinnamon, ginger, paprika and garlic showed the percent inhibition of 87%, 80%, 85% and 41% respectively
Table-2 Effect of Various Spice Extracts on Free Radical Scavenging
|S. No.||Functional Food||Scientific Name||Free Radical Scavenging (%)|
|1||Cumin seed||Cuminum cyminum||95|
|4||Paprika (red)||Capsicum annum||85|
|Ascorbic acid 1 mM (positive control)||95|
Among all tested extracts, only cumin seeds showed 50% inhibition in both glycation and anti-oxidant assays as shown in the Venn diagram (Figure-1).
Figure-1 Venn diagram at Threshold of 50 percent
The HPLC finger print of cumin seed extract is shown in Figure-2.
Figure-2 HPLC Fingerprint of Cumin Seed Extract
Ageing or senescence has been attributed to capacity loss and emergence of diseases in living beings. Both of which affect the quality of life. The deleterious mechanism like glycation has been reported to underlie the process of ageing and has been the pharmacological target of anti-ageing substances. In this regard, dietary interventions especially through spices appear to be of medicinal value and their evaluation against the said indication was the primary objective of the present study.
Dietary modulation has always been an important tool for the management of ailments, which are either of chronic nature or develop insidiously. Nutritionists often recommend use of specific constituents (Solfrizzi et al., 1999), diet (Berr et al., 2009) or ask to follow specific dietary patterns (Yannakoulia et al., 2015) to manage such situations. Furthermore, the diet based interventions were found to have greater patient compliance. In this context, the dietary spices hold a promising position and their consumption is negatively co-related with ageing and associated ailments especially in our part of the world, which is famous for spicy meals (Kannappan et al., 2011). Keeping this in view, the five commonly used spices were selected (Cumin seed, cinnamon, ginger, paprika and garlic) and their ethanolic extracts (70%) were prepared followed by assessment of their anti-glycation activity. Our data showed that cumin seeds extract was most potent in inhibiting the process of glycation with 80% inhibition (Table-1). Rest of the spices, did not produce a significant inhibition i.e. cinnamon, ginger, paprika and garlic showed the percent inhibition of 30%, 25%, 21% and 10% respectively. This is suggestive of anti-ageing potential of cumin seeds. Search of literature revealed that oxidative stress is also an important factor responsible for deteriorating bodily faculties with age. On similar lines, the spices were reported to hold anti-oxidant potential as well (Srinivasan and nutrition, 2014). Keeping this in view, all extracts were also tested for their ability to scavenge free radicals. It is of note that cumin seeds were again found to be the most potent with inhibition of 95% (Table-2). With an exception of garlic, the other spices i.e. cinnamon, ginger and paprika, also showed promising results with the percent inhibition of 87%, 80% and 85% respectively. The Venn diagram, at threshold of 50% revealed that among all tested extracts, only cumin seed was able to produce more than 50% inhibition in both assays. This suggests that along with inhibition of deleterious phenomenon of glycation, the cumin seeds extract additionally possesses anti-oxidant potential, which shall add value to its medicinal properties against ageing and associated diseases. Additionally, the HPLC fingerprint of cumin seed was also obtained for crude reflection of medicinal constituents residing in it. This shall also be useful for future comparisons in similar studies involving cumin seeds extract.
In conclusion, our data demonstrates that the cumin seed presents itself as the promising functional food for the management of age-associated diseases and diabetic complications attributed to the deleterious phenomenon of glycation.
AIYEGORO, O. A. & OKOH, A. I. 2010. Preliminary phytochemical screening and in vitro antioxidant activities of the aqueous extract of Helichrysum longifolium DC. BMC Complementary and Alternative medicine, 10, 21.
BENELLI, G., PAVELA, R., PETRELLI, R., CAPPELLACCI, L., CANALE, A., SENTHIL-NATHAN, S. & MAGGI, F. 2018. Not just popular spices! Essential oils from Cuminum cyminum and Pimpinella anisum are toxic to insect pests and vectors without affecting non-target invertebrates. Industrial Crops and Products, 124, 236-243.
BERR, C., PORTET, F., CARRIERE, I., AKBARALY, T. N., FEART, C., GOURLET, V., COMBE, N., BARBERGER-GATEAU, P. & RITCHIE, K. 2009. Olive oil and cognition: results from the three-city study. Dementia and geriatric cognitive disorders, 28, 357-364.
CAMPBELL, I. L. 1996. Exacerbation of lymphocytic choriomeningitis in mice treated with the inducible nitric oxide synthase inhibitor aminoguanidine. Journal of neuroimmunology, 71, 31-36.
CAMPISI, J., KAPAHI, P., LITHGOW, G. J., MELOV, S., NEWMAN, J. C. & VERDIN, E. J. N. 2019. From discoveries in ageing research to therapeutics for healthy ageing. 571, 183-192.
DEARLOVE, R. P., GREENSPAN, P., HARTLE, D. K., SWANSON, R. B. & HARGROVE, J. L. J. J. O. M. F. 2008. Inhibition of protein glycation by extracts of culinary herbs and spices. 11, 275-281.
GOLDIN, A., BECKMAN, J. A., SCHMIDT, A. M. & CREAGER, M. A. J. C. 2006. Advanced glycation end products: sparking the development of diabetic vascular injury. 114, 597-605.
HASLBECK, K.-M., SCHLEICHER, E., BIERHAUS, A., NAWROTH, P., HASLBECK, M., NEUNDÖRFER, B., HEUSS, D. J. E., ENDOCRINOLOGY, C. & DIABETES 2005. The AGE/RAGE/NF-κB pathway may contribute to the pathogenesis of polyneuropathy in impaired glucose tolerance (IGT). 113, 288-291.
HASLER, C. M. J. T. J. O. N. 2002. Functional foods: benefits, concerns and challenges—a position paper from the American Council on Science and Health. 132, 3772-3781.
IGASE, M. & IGASE, K. J. G. S. R. 2018. Cognitive impairment and glycative stress. 5, 45-49.
KANNAPPAN, R., GUPTA, S. C., KIM, J. H., REUTER, S. & AGGARWAL, B. B. 2011. Neuroprotection by spice-derived nutraceuticals: you are what you eat! Molecular neurobiology, 44, 142-159.
KHAN, S. A., HAIDER, A., MAHMOOD, W., ROOME, T. & ABBAS, G. J. P. B. 2017. Gamma-linolenic acid ameliorated glycation-induced memory impairment in rats. 55, 1817-1823.
LEUNG, C., HERATH, C. B., JIA, Z., ANDRIKOPOULOS, S., BROWN, B. E., DAVIES, M. J., RIVERA, L. R., FURNESS, J. B., FORBES, J. M. & ANGUS, P. W. J. W. J. O. G. 2016. Dietary advanced glycation end-products aggravate non-alcoholic fatty liver disease. 22, 8026.
POLE, A., DIMRI, M. & DIMRI, G. P. J. A. M. S. 2016. Oxidative stress, cellular senescence and ageing. 3.
RAMASAMY, R., VANNUCCI, S. J., YAN, S. S. D., HEROLD, K., YAN, S. F. & SCHMIDT, A. M. J. G. 2005. Advanced glycation end products and RAGE: a common thread in aging, diabetes, neurodegeneration, and inflammation. 15, 16R-28R.
REDDY, V. P. & BEYAZ, A. 2006. Inhibitors of the Maillard reaction and AGE breakers as therapeutics for multiple diseases. Drug discovery today, 11, 646-654.
ROSS, S. M. J. H. N. P. 2015. Sugar-induced aging: the deleterious effects of excess dietary sugar intake. 29, 114-116.
SERGIEV, P., DONTSOVA, O. & BEREZKIN, G. J. A. N. 2015. Theories of aging: an ever-evolving field. 7.
SHARMA, P., YELNE, M. & DENNIS, T. 2001. Database on medicinal plants used in Ayurveda, Central Council for Research in Ayurveda and Sidda. New Delhi, 2, 538-549.
SHEN, Q., ZHANG, B., XU, R., WANG, Y., DING, X. & LI, P. 2010. Antioxidant activity in vitro of the selenium-contained protein from the Se-enriched Bifidobacterium animalis 01. Anaerobe, 16, 380-386.
SINGH, R. P., GANGADHARAPPA, H. & MRUTHUNJAYA, K. 2017. Cuminum cyminum–A popular spice: An updated review. Pharmacognosy Journal, 9.
SOLFRIZZI, V., PANZA, F., TORRES, F., MASTROIANNI, F., DEL PARIGI, A., VENEZIA, A. & CAPURSO, A. 1999. High monounsaturated fatty acids intake protects against age-related cognitive decline. Neurology, 52, 1563-1563.
SOUTHERN, L., WILLIAMS, J. & ESIRI, M. M. J. B. N. 2007. Immunohistochemical study of N-epsilon-carboxymethyl lysine (CML) in human brain: relation to vascular dementia. 7, 35.
SRINIVASAN, K. J. C. R. I. F. S. & NUTRITION 2014. Antioxidant potential of spices and their active constituents. 54, 352-372.
SUN, H., YUAN, Y. & SUN, Z. J. B. R. I. 2016. Update on mechanisms of renal tubule injury caused by advanced glycation end products. 2016.
SUZMAN, R. & BEARD, J. J. W. H. O. 2011. Global health and ageing. Bethesda, MD: US Department of Health and Human Services.
THORNALLEY, P. J. 2003. Use of aminoguanidine (Pimagedine) to prevent the formation of advanced glycation endproducts. Archives of biochemistry and biophysics, 419, 31-40.
YANNAKOULIA, M., KONTOGIANNI, M. & SCARMEAS, N. 2015. Cognitive health and Mediterranean diet: just diet or lifestyle pattern? Ageing research reviews, 20, 74-78.
YONEI, Y., YAGI, M. & TAKABE, W. J. G. S. R. 2020. Stop the “Vicious Cycle” induced by Glycative Stress. 7, 13-21.
ZARANDI, M. H., ALLAHDADI, I., KHALAJ, H. & LABBAFI, M. 2017. Cuminum cyminum L. Seed Science and Technology, 6.