INTRODUCTION:

The pharmaceutical business has been studying natural plant alternatives as an excipient for decades¹. The food, cosmetic, pharmaceutical, and nutraceutical sectors are the main consumers of herbal products. Botanical extracts and semi-finished herbal goods are in high demand. Among the Opuntia varieties, desert plants are particularly notable. Since ancient times, O. elatior has been used as medicine².

In south India, thrives in the wild. It also has a pear bush that produces berries all year long³. It is best suited for dry environments and made to rapidly absorb and store rainwater from erratic downpours. The growing components of these plants are commonly referred to as pads, joints, and cladodes⁴.

The modified stems, which have an ovoid or elongated form and a length of 20–30cm, take the place of leaves in the photosynthetic process. The inner portion, which is composed of white medullar parenchyma, is where water is primarily kept⁵. When leaves become spines, they develop into tiny barbs known as areoles⁶. On these boulders, prickly areole-producing plants flourish. The biggest ones are 2cm long and have sharp edges. Every internode contains 35 areoles⁷. [The mining, phytochemical/ biological assessment of O. elatior cladodes have been studied extensively⁸. Additionally, despite earlier studies not reporting any investigation, a comparison of temperature and time effect on percent yield was carried out using Design of Experiment software ⁹.

Factorial Design (FD):

The influence of one variable is typically studied at a time in traditional research methods because it is possible to manipulate it statistically. However, only one influencing element can be investigated at once¹⁰. Attempting to combine these two factors will result in inaccurate findings because of their interdependence. Multivariate analysis includes the design of experiments (DOE) as a component^11,12. DOE is still regarded as a treaty with some elements, but not all of them. Screening and optimising replies is one of DOE's goals¹³.The imitations investigate potential combinations of the factor values. All input variables are referred to as FD at both the "high" (+1) and "low" (-1), or FD, levels¹⁴. It is recommended to use a fractional FD/ or Plackett-Burman design (PBD), as the number of runs will be greatly increased above 5 factors¹⁵. When there are four to five factors, the screening goal is FD^16,17, and the response surface goal is CCD or BBD. For screening factors containing five or more factors, FDs or PBDs are appropriate, followed by response surface goals^18,19. Most widely used DOE software is Design-Expert/Statistica/Fusion/Minitab. The dependent variable is created by adding the independent mutable to the anticipated output^20,21.

MATERIALS AND METHODS:

Raw Material:

The plants used to make the Opuntia elatior cladodes were found in the dry hills near Anantapur, Andhra Pradesh, India. A 2 x 6cm sample of cladodes was gathered. It was recognized and verified by the botanical department at Sri Krishnadevaraya University in Anantapur. The Herbarium received an Exemplar (SKBD/21/029) that was placed.

Extraction of Mucilage:

Cleansed and prepared for mucilage separation were the cladodes from Opuntia elatior. Manually separating the medullar covering from the medullar parenchyma and crushing it. 200cc of water were added to 100g of parenchyma. The substance was first filtered through a home sieve and stirred at 100rpm for 60–90min at 40–50^oC^22,23. After precipitation with 95 percent ethanol, the mucilage was extracted from the supernatant. Following that, 100ml of the filtrate were mixed with 200ml of ethanol. The mucilage was dried at 40°C for six hours to produce a powder. Three copies of the extracted data were made^14,24,25.

RESULTS AND DISCUSSION:

As mentioned earlier, mucilage can serve as an inexpensive and natural replacement for many industrial processes. Research is therefore needed to regulate its efficiency. Various extraction methods have been examined. A temperature and time impact been evaluated.

Mucilage Extraction:

Thermal extraction produced mucilage yields from fresh cladodes that varied from 59.1 to 61.5%. (Table 1). Quadratic polynomials for O. elatior cladodes were calculated based on the investigational design data and the mucilage % discovered for each condition, and were then verified by ANOVA analysis.

Initial risk assessment:

Q8 and Q9 of ICH state that the Quality Target Product Profile (QTPP) is critical for quality by design (QbD). Additionally, it is crucial to separate unbiased viewpoints from biased ones at the beginning of the product development process. A quality product must meet the QTPP's standards to be considered quality. The QTPP and CQAs for extraction are healthy due to past surveys and appraisals of the literature.

Experimental design:

Using a central composite design (CCD), quadratic response surfaces were created and evaluated in this research to screen the effects of temperature and exposure time (ET) on the extraction of contents from the cladodes of O. elatior. Design-Expert software (11.0) was used by Stat-Ease Inc. to evaluate the factors. The key, interface, and quadratic properties of independent variables on dependent variables were evaluated using the resulting quadratic model.

Y = B⁰+B¹X¹+B²X²+B¹²X¹X²+B¹X¹²+B²X²²

Here, Y is the dependent variable, X1 and X2 are the independent variables, and B⁰, B¹, and B² are the regression values of the independent variables and their mutual interfaces. (Temperature and ET). The yield of O. elatior cladodes served as the dependent variable and reaction. Table 1 expresses the core composite design experimental designs.

Table 1: The experimental design by Design-Expert software for reviewing the impact of temperature and exposure time for the extraction of contents from the cladodes of O. elatior

Trials	Factor 1 (A) Time (min)	Factor 2 (B) Temperature (^oC)	Response (% yield)
1	60	40	60.6
2	90	40	61.5
3	60	50	59.3
4	90	50	60.5
5	53.7868	45	60.5
6	96.2132	45	61.2
7	75	37.9289	60.6
8	75	52.0711	59.1
9	75	45	59.7

The factors were assessed using the Response Surface Methodology. (Time and temperature). (RSM). The best conditions for mucilage extraction were identified by the highest values of the response surface of the temperature and time curves. Utilizing Stat-Ease-Design Expert V.11, the untested designs were created and approximated. One-way analysis of variance (ANOVA) was used to carry out the statistical validation, which had a 95% degree of confidence. A central composite design with constraints was used to create star points, and a factorial design was used to construct models. The speech produced the following reaction:

Yield=+59.70+0.3862A-0.5527B+0.0750AB+0.6062A² + 0.1062B².

The equation allows one to predict the outcome for specific levels of each component. Factors with a high level are typically classified as +1, while those with a low level are typically coded as -1. The coded equation can be used to control the factors' relative significance by comparing their coefficients. Quadratic and linear models were proposed as fit summaries for O. elatior cladodes' response to percent yield. (Table 2).

Table 2: Fit Summary for the response (%Yield) of O. elatior cladodes

Source	Sequential p-value	Adjusted R²	Predicted R²
Linear	0.0309	0.5816	0.3209	Suggested
2FI	0.8039	0.5047	0.2062
Quadratic	0.0392	0.9048		Suggested
Cubic	0.4063	0.9528

Additionally, the ANOVA for Quadratic model for the response (% yield) O. elatiorcladodes were shown in table 3.

Table 3: ANOVA for Quadratic model for the response (% yield) for O. elatior

Source	Sum of Squares	df	Mean Square	F-value	p-value
Model	5.11	5	1.02	16.20	0.0222
A-Time	1.19	1	1.19	18.92	0.0224
B-Tempt.	2.44	1	2.44	38.73	0.0084
AB	0.0225	1	0.0225	0.3566	0.5924
A²	1.07	1	1.07	16.95	0.0260
B²	0.0328	1	0.0328	0.5205	0.5228

Having an F-value of 16.20 designates that the model is significant. There is only a 2.22% chance that an F-value with this size will be considered significant if their P-values are less than 0.05 are significant. A, B, and A² are significant model terms. If the value is greater than 0.10, then the model terms must not be significant. If you have many insignificant variables, it may be advantageous to reduce model terms (Table 1).

The linear term for time has a positive coefficient for young cladodes, demonstrating a rise in mucilage extraction over time. Large changes in time, either in one way or the other, are associated with higher extraction yields, according to a minimum in the response surface. (Positive quadratic terms). The interface between the two factors regulates the amount of mucilage that can be extracted, even though the linear term for temperature has no effect on mucilage yield. The quadratic terms show that large temperature variations, whether positive or negative, boost extraction yield. The rate of mucilage extraction does not seem to be substantially impacted by time during the time period considered (60-90 min). The interfaces between variables have no discernible impact on mucilage withdrawal when the sureness intervals are taken into consideration. The interfaceimpact of temperature and ET on the percent yield from the cladodes of O. elatior were uttered in the figure 1, whereas the contour plot and 3D response plots were signified in figure 2.

The effect of temperature and time on yield was resolute using Response Surface Methodology (RSM). The maximum value of the response surface designates the idealsituations for mucilage extraction. Using Stat-Ease-Design Expert V.11, experimental designs were generated and evaluated using an ANOVA at a confidence level of 95%. By generating models with circumscribed central composite designs based on a factorial design with fewer star points, we created models with circumscribed star points.

To demonstrate a retort at dissimilar levels of each factor, the equation in the form of coded factors may be used. Coded equations can be used to control the percentage involvement of each factor to an outcome by comparing their constants. The high values of the factors are assumed to be +1 and the low values as -1.

CONCLUSION:

Design of the experiment was carried out using QbD software called Design Expert to decide the effect of temperature and exposure time on the extraction of Opuntia elatior cladodes. The reading concluded that the cladodes can be extracted at a temperature of 40^oC and the exposure time is directly relative to the temperature.

ACKNOWLEDGEMENTS:

The authors are thankful to the Department of Industrial Pharmacy of the college for the encouragement and support.

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Received on 17.07.2022 Modified on 27.12.2022

Research J. Pharm. and Tech 2023; 16(12):5760-5764.

DOI: 10.52711/0974-360X.2023.00932